AN ABSTRACT OF THE THESIS OF
Carlos L. Ballare
for the degree of
Doctor of Philosophy
in Crop Science presented on July 10, 1992.
Title:
Photomorphogenic Processes in the Agricultural
EnvAr,nment
Abstract approved:
Redacted for Privacy
Steven R. Radosevich
I review recent advances in photomorphogenic research
and use information derived from physiological experiments
to explain patterns of space occupation and competition
among neighboring plants. I found compelling evidence that
the presence of surrounding vegetation influences plant
growth and development not only because it modifies the
availability of light energy and materials, but also because
it affects aspects of the environment that are specifically
used by seeds and plants to obtain information about the
proximity of neighbors (e.g. red:far-red ratio, thermal
regime, photon-fluence-rate gradients). The significance of
recent findings on signal perception by plants is discussed
in connection with the development of mechanistic models of
plant competition.
I used a photomorphogenic mutant of cucumber (Cucumis
sativus L.) to explore the roles of the photoreceptor
phytochrome B in morphological responses to UV-B radiation
and in processes that influence morphological development in
patchy light environments. The studies on UV-B responses
provided strong circumstantial evidence that some
morphological effects of UV-B on green plants can be
separated from unspecific damage and involve the action of a
UV-B photoreceptor. Experiments in natural and artificial
light/shade mosaics showed that, when compared over periods
of several weeks, wild-type (WT) plants are far more
efficient than phytochrome-B-deficient mutants (lh) at
invading and maintaining their shoots in light gaps. The
reasons for this difference were multiple and appeared to
result from several actions of phytochrome B on WT plants.
These actions include: (i) establishment of a phototropic
system that responds to lateral red:far-red gradients,
(ii)
modulation of gravitropic responses of stems, (iii) delay of
tendril production and reduction of stem-bending responses
to mechanical stresses exerted by tendrils, and (iv)
switching plants grown under full light from the "shadeavoiding" (default) developmental program to the one that
results in a phenotype with short internodes and reduced
apical dominance.
In the final section of this thesis I discuss the
agricultural significance of photomorphogenic research.
Three areas are considered: photocontrol of seed
germination, morphological and pigmentation responses to UVB radiation, and photomodulation of plant morphology in
canopies.
Photomorphogenic Processes in the Agricultural Environment
by
Carlos L. Ballare
A THESIS
submitted to
Oregon State University
in partial fulfillment of
the requirements for the
degree of
Doctor of Philosophy
Completed July 10, 1992
Commencement June 1993
APPROVED:
Redacted for Privacy
Pro essor of Crop and Soil Science in charge of major
Redacted for Privacy
Head of department of Crop and Soil Science
Redacted for Privacy
Dean of Grad
Schoo
Date thesis is presented July 10, 1992
Typed by
Carlos L. Ballare
ACKNOWLEDGEMENTS
I wish to express here my sincere gratitude to the
individuals and institutions that contributed to the success
of this endeavor. I am deeply indebted to Dr. Steven R.
Radosevich for his gracious advise in all the steps that led
to the completion of my degree, and for his hospitality and
support during my stay at Oregon State University. He and
Dr. Mary Lynn Roush significantly contributed to broaden my
scientific background and forced me to review most of my
ideas on ecological research and the relationships between
science and society. I am also grateful to Dr. Joe B. Zaerr
for his advise, for discussions on aspects of science and
education, and for helping me with the translation of German
papers. He and Dr. Radosevich gave me opportunities to
lecture in their Tree Physiology and Weed Ecology classes.
My appreciation is also extended to Dr. Terri Lomax, for
serving as a committee member and for organizing a class
based on seminars during the fall of 1990 where I learned a
great deal on molecular aspects of plant development. I had
also the good fortune of having Dr. Leslie Fuchigami as the
Graduate Council Representative in my committee. Les was
collaborative in every respect and I am most grateful for
his cordiality and encouragement. The cooperation of Dr.
Paul W. Barnes during the first months after my arrival to
Corvallis was very stimulating. Paul allowed me to work in
the facilities of the U. S. Environmental Protection Agency
and spent many hours discussing with me aspects of
photobiology and physiological ecology.
I thank the Consejo Nacional de Investigaciones
Cientificas y Tecnicas (CONICET) of Argentina for granting
me an External Fellowship that made this visit to the United
States possible and to the Dept of Forest Science, O. S. U.,
for additional financial support.
I am deeply indebted to Claudio M. Ghersa and M.
Alejandra Ghersa, two Argentinean colleagues and friends who
helped me and my family in many aspects of our daily life in
Corvallis. The Ghersa's place was also the locus of many
stimulating discussions on science and ecology. Tim Brewer,
Abul Hashem, Andre Laroze, Doris Prehn, the Olivas,
Thomanns, Barbara and Gil Vera, and Barbara Yoder were among
the friends and mates that contributed to make our stay more
enjoyable.
I also wish to thank several O.S.U. staff members for
their competence and cordiality, specially Lu Berger,
Gretchen Bracher, Fred Friedow, Bud Graham, Randy Hopson,
and Mayvin Sinclair.
My greatest appreciation goes to my wife, Ani, and to
our son Nicolas, for their love and strength. This work is
dedicated to them and to my parents, Pedro and Dora, and my
sister Cecilia.
CONTRIBUTION OF AUTHORS
Co-authors of individual papers are identified on the
first page of each chapter. The following is brief list of
their contributions to each manuscript.
Paul W. Barnes advised me on techniques used for UV-B
irradiation and dosimetry, collaborated in the design and
execution of the experiments presented in chapter 3, in the
discussion of hypotheses and results, and edited successive
drafts of the manuscript.
Richard E. Kendrick corresponded with me on some of the
ideas presented in chapters 3 and 4 and reviewed the
manuscripts.
Steven R. Radosevich advised me on many steps along the
completion of this thesis. He contributed to the ideas
discussed in chapters 2, 4, 5, and 6 and edited early
versions of the manuscripts presented in chapters 4, 5, and
6.
Rodolfo A. Sanchez discussed with me several of the
thoughts and hypotheses presented in chapter 6 and commented
on an early draft of that paper.
Ana L. Scopel was involved in the discussion of the
hypotheses of chapters 4, 5, and 6, collaborated in the
design and execution of experiments, and reviewed and
improved successive drafts of the manuscripts.
TABLE OF CONTENTS
page
CHAPTER 1. GENERAL INTRODUCTION
CHAPTER 2. LIGHT GAPS: SENSING THE LIGHT OPPORTUNITIES
IN HIGHLY DYNAMIC CANOPY ENVIRONMENTS
Introduction
Sensing and responding to light
opportunities
Competition for light during early
succession
Signals, decisions, and models of
plant competition
Summary
References
CHAPTER 3. PHOTOMORPHOGENIC EFFECTS OF UV-B RADIATION
ON HYPOCOTYL ELONGATION IN WILD-TYPE AND STABLEPHYTOCHROME-DEFICIENT MUTANT SEEDLINGS OF CUCUMBER
Abstract
Introduction
Materials and Methods
Results
Discussion
References
1
6
7
8
31
34
38
49
64
65
66
68
71
73
83
CHAPTER 4. PHYTOCHROME-MEDIATED PHOTOTROPISM
IN DE-ETIOLATED SEEDLINGS. OCCURRENCE AND ECOLOGICAL
SIGNIFICANCE
Abstract
Introduction
Materials and methods
Results
Discussion
References
111
CHAPTER 5. FORAGING FOR LIGHT IN HORIZONTALLYPATCHY ENVIRONMENTS BY WILD-TYPE AND PHYTOCHROME-BDEFICIENT CUCUMBER PLANTS
Abstract
Introduction
Materials and Methods
Results
Discussion
Conclusions
References
115
116
118
120
123
130
135
146
CHAPTER 6. PHOTOMORPHOGENIC PROCESSES IN THE
AGRICULTURAL ENVIRONMENT
Abstract
Introduction
Photocontrol of weed seed germination
in arable land
87
88
89
91
94
97
150
151
153
153
Plant responses to solar UV-B
Photomorphogenic signals in process of
plant competition
R:FR-signals and phytochrome
Fluence-rate signals
Implications for crop improvement
Summary and outlook
References
157
162
162
164
166
173
176
LIST OF FIGURES
Figure
2.1 Spectral distribution of daylight and wavelengths
absorbed by known plant photoreceptors.
2.2
2.3
2.4
2.5
2.6
2.7
2.8
3.1
3.2
3.3
page
41
Effects of sunlight pulses on the germination of
buried seeds of Datura ferox under field
conditions.
42
Elongation rates of the first internode of Datura
ferox seedlings growing in even canopies of
different densities under natural radiation and
effect of density on PAR interception by individual
seedlings.
43
Effect of the proximity of a green grass canopy
on the elongation of the first internode of
Sinapis alba seedlings growing under full sunlight.
44
Effects of neighboring seedlings on the spectral
distribution of the radiation scattered within the
stems of Datura ferox seedlings in an even canopy
of LAI = 0.72.
45
Effects of increasing density in even canopies of
dicotyledonous seedlings on light interception by
leaves and the light environment of the stems.
46
Elongation responses of Datura ferox first
internodes when seedlings were placed in the
center of an even canopy of LAI -0.9 with the
internodes surrounded by annular cuvettes
containing distilled water (clear filter) or a
CuSO4 solution that absorbed FR radiation and
maintained the R:FR ratio at ca. 1.1 (FR-absorbing
filter).
47
The effect of green filters that reduced the R:FR
ratio of side-light on the azimuthal orientation of
main shoots of Portulaca oleracea seedlings grown
in the field.
48
Spectral photon distributions measured at plant
level under control (polyester-filtered) and UV-B
(cellulose-acetate filtered) lamp systems.
78
Effect of 6-h exposures to UV-B on hypocotyl
elongation, cotyledon area expansion and dry
matter accumulation in de-etiolated lh and WT
cucumber seedlings.
79
Recovery from UV-B inhibition of hypocotyl
3.4
3.5
4.1
4.2
4.3
4.4
4.5
4.6
4.7
4.8
5.1
5.2
elongation in de-etiolated lh and WT cucumber
seedlings.
80
Effects of GA3 on hypocotyl elongation growth in
de-etiolated lh and WT cucumber seedlings exposed
to UV-B.
81
Effects of covering different parts of the seedling
with a UV-B-excluding polyester filter on hypocotyl
elongation in de-etiolated cucumbers exposed to
UV-B for 6 h.
82
Spectral photon distributions of the light sources
used in the controlled-environment experiments on
phototropism.
103
Effect of the proximity of a green canopy on the
orientation of the hypocotyl of WT seedlings at
solar noon.
104
Time courses of hypocotyl bending in WT and lh
seedlings growing under white light with or
without exposure to unilateral FR light.
105
Effects of increasing the fluence rate of
unilateral FR light on hypocotyl curvature in WT
and lh seedlings.
106
Effects of increasing the fluence rate of bilateral FR light on hypocotyl elongation in WT
and lh seedlings (the latter with or without
application of GA3).
107
Effect of shading the hypocotyl during irradiation
with unilateral FR light (121.7 gmol m2 s4) on
hypocotyl curvature in WT seedlings.
108
Effects of the proximity of a green maize canopy
and B-absorbing acetate filters on the spectral
photon distribution of the light flux measured
parallel to the ground.
109
Effects of the proximity of a green maize canopy
and B-absorbing acetate filters on the orientation
of the hypocotyl of WT and lh seedlings at solar
noon.
110
Morphological development of WT and lh plants
under conditions of uniform illumination in the
controlled environment.
138
Graviotropic responses of mature WT and lh plants
under conditions of full light in the controlled
environment.
139
5.3
Spatial distribution of nodes produced by WT and lh
plants after four weeks of growth at the border of
a maize canopy in the field.
140
5.4
Internode production, spatial distribution of main
stems and tendril dynamics in WT and lh plants
grown for 6 weeks at the border of simulated
canopies in a controlled environment.
141
Angular orientations of leaves and main-stem
internodes of WT and lh plants grown for 5 weeks
at the border of artificial canopies in the
controlled environment.
142
Time courses of artificial gap invasion by six WT
and six lh shoots in the controlled environment.
143
Effects of tendril attachment to supports on
tendril morphology and internode orientation.
144
Changes of internode curvature after severing the
attached tendril in lh plants.
145
5.5
5.6
5.7
5.8
LIST OF TABLES
Table
4.1
5.1
page
Effects of the proximity of green plants on the
light flux measured parallel to the ground.
102
Stem length and angle of inclination toward the
artificial light gaps in WT and lh-mutant
seedlings.
137
PHOTOMORPHOGENIC PROCESSES IN THE AGRICULTURAL ENVIRONMENT
CHAPTER 1
GENERAL INTRODUCTION
Life on Earth is dependent on the process of
photosynthesis performed by plants. Apart from providing the
energy for photosynthesis, light has a dramatic influence on
the development of living organisms. Light is a vehicle of
information. Many life forms, but particularly
photoautotrophic plants, have evolved fascinating systems to
obtain information about their environment by "measuring"
certain aspects of the impinging radiation. The acquisition
of information by higher plants is the principal focus of
subsequent chapters, but I will give some general examples
here. Seeds of weedy species do not germinate at random
through time. Light is one of the environmental factors that
signals seeds to germinate. A specific photoreceptor in the
seeds (phytochrome) can detect short light pulses when the
surrounding soil is disturbed by tillage, or changes in the
spectral distribution of radiation when a tree-fall opens a
gap in the canopy. When seedlings or sprouts penetrate
through the soil toward the surface their morphology is
completely different from the one we see after emergence.
The stems are elongated, the leaves rudimentary, and tissues
contain little or no chlorophyll. It is light, perceived by
specific photoreceptors while the seedlings are breaking
through the soil surface, that triggers all the changes in
morphology and physiology that make the seedling competent
to photosynthesize. The enzymes that are responsible for CO2
fixation in the chloroplasts are not active all the time.
2
The reducing pathway is shut off every night. It is the
activity of the photosystems, fueled by sunlight, that
changes the chloroplast environment and activates the
photosynthetic catalysts. The morphology of plants is
tremendously plastic. Plants grown with plenty space are
often profusely branched, short, and form extended canopies
(i.e. umbrella-type trees). Plants of the same genotype
grown in a crowded population are commonly less branched and
form vertically-extended canopies or crowns (i.e. lolly-pop
type trees). It is light, detected by specialized
photoreceptors, that informs plants about the proximity of
neighboring individuals and elicits changes in morphological
development. Finally, many plants go through seasonal cycles
of vegetative and reproductive activity. The duration of the
photoperiod, detected by internal mechanisms re-phased by
light signals, is one of the factors that informs plants
about seasonal changes.
Photomorphogenesis is a term used to refer to all
regulatory actions of light on plant growth and development
that are independent of photosynthesis. A milestone in the
study of photomorphogenesis was the discovery, forty years
ago, of the plant photoreceptor phytochrome, by a scientific
group led by H. A. Borthwick and S. B. Hendricks at the
U.S.D.A. Plant Industry Station, Beltsville. We know today
that phytochrome is not a single pigment. In Arabidopsis,
for example, three different phytochromes and five different
phytochrome genes have been discovered over the last few
years. In addition, we have learned that other
photoreceptors, such as the blue/ultraviolet-A-, and the
ultraviolet-B-absorbing photoreceptors, are involved in the
control of photomorphogenesis.
How do these photoreceptors act? The first step is
light absorption by the photoreceptor (e.g. phytochrome)
3
accompanied by some change in the properties of the pigment
molecule. This alteration initiates a transduction process,
that is a series of molecular events that culminate in
altered growth or development. The precise sequence for any
response is not yet known, but the gap in understanding of
how the "black boxes" work between light absorption and
observable response has been dramatically narrowed during
the last few years. Phytochrome-mediated processes are being
studied intensively. One of the things that we have learned
is that there is more than just one primary mechanism of
phytochrome action. In some cases it is clear that plant
responses mediated by phytochrome do not involve changes in
gene expression (e.g. chloroplast orientation in the alga
Mougeotia). Yet, in others (probably the majority of
phytochrome responses), light absorption by phytochrome
leads to altered expression of selected genes. There is
little evidence for a direct interaction between phytochrome
and the genome and, possibly, there is more than one
mechanism whereby phytochromes affect transcriptional
activity.
Two areas of photomorphogenic research have made rapid
progress during the past two decades. The explosion of
molecular biology gave photobiologists a whole spectrum of
new techniques to study the molecular properties of
photoreceptors and to unravel signal transduction chains. A
significant example is the recent characterization of
several phytochrome genes and molecular species, which
opened the possibility that different phytochromes have
different mechanisms of action and physiological roles. On
the other hand, many plant physiologists and ecologists
became interested in the significance of photomorphogenesis
in the natural environment.
This thesis is focused on the ecological significance
4
of photomorphogenesis in the agricultural environment. In
Chapter 2, I review some recent developments on how seeds
and growing plants perceive the light environment and
discuss the role of information acquired via
photomorphogenic pigments on mechanisms of competition in
plant communities. The style of chapter 2 is deliberately
provocative. Its last section contains a great deal of
speculation, and I invite the reader to take some of the
ideas expressed there as an evolving conceptual model, from
which constructive discussion and experimentation could be
derived.
In the next three chapters I explore the involvement of
photomorphogenic mechanisms and phytochrome on selected
responses of plants to the light environment. Each chapter
was written as an individual manuscript and can be read
independently of the others. The experiments reported in
Chapter 3 constitute and attempt to understand morphological
(more specifically stem-elongation) responses of plants to
ultraviolet-B radiation. This is an issue of importance for
agriculture given the increased concern about stratospheric
ozone depletion and its potential effects on ultraviolet-B
levels received by plant communities. Although there is
considerable information on effects of ultraviolet-B on
plant growth, the underlying mechanisms are still poorly
understood. Chapter 4 is concerned with the ecological
significance of plant phototropic responses and, by using a
phytochrome-B-deficient mutant of cucumber, I investigated
the role of phytochrome in phototropism. The major
conclusions of that series of experiments are that (i)
phytochrome functions as a phototropic sensor in green
plants and (ii) phototropic responses to red:far-red
gradients created by neighboring plants allow young
seedlings to actively project leaves into light gaps when
colonizing patchy canopies. This study is expanded upon in
5
Chapter 4, where I compared wild type and phytochrome-B-
deficient plants of cucumber in regard to their efficiency
to invade light gaps in heterogeneous canopies. Several
critical actions of phytochrome B on plant morphological
development that are of key importance in the processes of
"foraging" for light in patchy environments are identified
and discussed. The evidence derived from these physioecological studies with a photomorphogenic mutant has
significant implications for our perceptions about
mechanisms of competition among neighboring plants.
In the last Chapter, I review recent advances in the
understanding of the ecology of photomorphogenesis from the
perspective of an agronomist. Three major areas are covered:
photocontrol of seed germination, responses to solar
ultraviolet-B, and morphological and developmental responses
in plant canopies. The chapter closes with a discussion on
implications of recent discoveries for the improvement of
cropping systems. One of the points emphasized in that
review is the importance to pursue photo-physiological
research under field conditions. Such an approach is
urgently needed if the goal is to discover how the
expression and agronomic significance of photomorphogenic
responses is affected by other factors of the crop's
environment.
6
CHAPTER 2
LIGHT GAPS: SENSING THE LIGHT OPPORTUNITIES IN HIGHLY
DYNAMIC CANOPY ENVIRONMENTS
Carlos L. Ballar6
Depts of Forest Science and Crop and Soil Science, Oregon
State Univ., Corvallis, OR 97331-5705, USA, and Dept de
Ecologia, Facultad de Agronomia, Univ. de Buenos Aires,
(1417) Buenos Aires, Argentina.
Exploitation of Environmental Heterogeneity by Plants:
Ecophysiological Processes Above and Below Ground. Edited by
M. M. Caldwell and R. W. Pearcy, Academic Press, In review.
7
I. INTRODUCTION
In natural environments the amount of photosynthetic
light energy received at a given point in space is extremely
variable over time. Part of this variability is associated
with seasonal changes, part is due to the activity of nearby
vegetation and, at a finer time scale, to weather factors
and changes in solar angle during the course of the day.
Because plant fecundity and carbon gain are likely to be
positively correlated in most situations, the ability to
exploit light opportunities in temporally and spatially
patchy environments should be regarded as a major
determinant of evolutionary success. The first (obvious?)
condition for a plant to exploit a pulse of light
availability is that it must be growing at the right time.
That is germination or sprouting must be synchronized with
the onset of periods of high light availability. The second
condition is that the plant has to manage to position and
orient its light absorbing surfaces so as to maintain high
rates of light capture during the growing period. Finally,
the photosynthetic apparatus must be adapted to operate
under the prevailing light environment.
Plants exhibit a fascinating variety of mechanisms for
acquiring information about the light environment. These
mechanisms operate at several stages controlling, inter
allia, the timing of seed germination, the transition to
the photoautotrophic stage, the spatial orientation of
branches and leaves, the elongation of stems and hence the
height of leaf insertion, the production of new branches or
tillers, and the performance of the photosynthetic
apparatus.
The consequences of these mechanisms, that is the
association between germination flushes and canopy openings
or soil disturbances, or between plant morphology and canopy
density and structure, have been the subject of previous
reviews or are covered by other authors. This chapter deals
8
with the processes involved in the perception of light
opportunities by plants in systems characterized by rapid
changes of plant cover over time, such as those dominated by
herbaceous species in early secondary succession. In the
first and major part of the paper I concentrate on what is
known about the nature of the environmental signals and
sensory systems that regulate physiological and
morphological responses to changes in light environment. The
focus is on germination responses of seeds and morphological
reactions of plants of early successional species. Then I
present an overview of the likely roles of informationacquiring systems for plants that exploit patchy and highly
dynamic light environments. Finally, I discuss the necessity
of considering these systems in mechanistic models of plant
competition. Most of the examples I use in the discussion
are derived from research on herbaceous species from
agricultural plant communities.
II. SENSING AND RESPONDING TO LIGHT OPPORTUNITIES
A. Environmental signals and plant receptors
Seeds and growing plants use many environmental cues to
obtain information about the prevailing light climate. These
cues are factors of the environment that, under natural
conditions, are directly or indirectly associated with
important aspects of the flux of photosynthetically active
radiation (PAR), such as its intensity, direction, and
likelihood of future change. Examples of these cues are the
amplitude of the daily fluctuation of soil temperature
sensed by a seed, the differential intensity of blue light
received at opposite flanks of a stem, or the ratio between
the fluxes of red and far-red radiation. The way in which
seeds and plants perceive these signals is considered in
this section. As it will be seen light signals, perceived by
specific photoreceptors play in many cases a fundamental
role in the detection of light opportunities. Therefore, a
brief description of plant photoreceptors is necessary here.
As far as we know today, higher plants contain at least
three families of photoreceptors (e.g. Mohr 1986) (Fig.
2.1):
1. Phytochromes, which absorb radiation over a wide range
from the ultraviolet (UV) to the far-red (FR) with maxima
9
around 660 nm for the Pr form, and 730 nm for the Pfr form
2. A series of blue (B) / UV-A absorbing pigments
3. A little characterized receptor for UV-B radiation.
For the purposes of this paper we can assume that these
receptors are devices that transform light into biological
signals (see Smith 1982, for a discussion of partial
processes involved in light perception by plants). It should
be kept in mind that, in addition to the specific
photoreceptors listed above, the two photosystems in
chloroplasts (PSI and PSII) may act as sensors of variations
in light environment and play a role in processes such as
the long-term photosynthetic acclimation to total irradiance
(e.g. Chow et al. 1990). More indirectly, plants may
accommodate to variations in light conditions by sensing and
reacting to changes in levels of photosynthetic products,
which may act as internal developmental signals (see an
example in Section II.B.5.).
Phytochrome is the best characterized of all plant
photoreceptors and, because it is involved in many of the
responses discussed in this chapter, a brief overview of its
major photochemical properties is included in this section.
Detailed coverage can be found in several recent books (e.g.
Kronenberg and Kendrick Eds. 1986, Furuya Ed. 1987, Attridge
1990). Phytochromes are now known to be a family of proteins
associated with a very similar chromophore (Sharrock and
Quail 1989, Furuya 1989). Phytochrome molecules exist in two
relatively stable forms, Pr and Pfr, with absorption maxima
in the R and FR regions of the electromagnetic spectrum,
respectively Fig. 2. 1). Each form is converted into the
other upon absorption of light. Because the absorption
spectra of Pr and Pfr overlap to a large extent, the pigment
cannot be pushed to pure Pr or Pfr. Thus, under conditions
of continuous illumination, a photoequilibrium (0) between
the two forms is reached that may be characterized by the
proportion of phytochrome molecules that are in the Pfr
form, i.e. the Pfr/P ratio. Values of 0 may vary from 0.02
under monochromatic FR to 0.86 under monochromatic R (e.g.
Mancinelli 1988). If the pigment is exposed to polychromatic
light, as is the case under natural conditions, the Pfr/P
ratio at 0 is highly dependent on the ratio of R to FR
10
photon fluxes in the incident radiation (R:FR ratio) (Smith
and Holmes 1977). Thus, 0 values obtained by exposing
purified phytochrome preparations to natural light fields
typically vary from ca. 0.6 (direct sunlight; R:FR = 1.15)
to 0.15 (light filtered through dense leaf canopy; R:FR =
0.2). The effects of light treatments on a number of
physiological functions are well correlated with their
effects on 0. Values of 0 can be estimated analytically
(Hartmann and Cohnen-Unser 1972) using spectral irradiance
data and published values of phytochrome photoconversioncoefficients (e.g. Smith 1986, Mancinelli 1988). The same
model can be used to calculate the rate of cycling between
Pr and Pfr, which might modulate certain responses to total
irradiance (Section II.B.3.), and to estimate how much Pfr
is formed after exposures to light that are too short to
drive the pigment to photoequilibrium (e.g. light pulses
perceived by seeds during soil cultivations, Section
II.B.1.)(see Cone and Kendrick [1985], Scopel et al.
[1991]). It should be noted here that physiological
responses to different aspects of the light environment
(e.g. total irradiance, R:FR ratio) might be mediated by
different molecular species of phytochrome.
B. Plant responses to light opportunities
1. Seed Germination.
Successional environments are characterized by
transient increases in light availability at the soil
surface, which are caused by disturbances that eliminate or
reduce the existing vegetation. Canopy gaps may be created
by natural agents (i.e. fire, storms, or herbivores) or by
man in managed ecosystems. Gaps may be of very different
sizes, from minute ones created by small herbivores foraging
on herbaceous vegetation to those involving hundreds of
hectares caused by tillages in cultivated areas. Frequently,
disturbance is followed by pulses of germination of seeds of
colonizing species.
The environmental changes brought about by disturbances
that are responsible for triggering seed germination are, as
can be anticipated, many and diverse. To a large extent the
subject is reviewed by Vdsquez-Yanez (1993); therefore,
coverage here will be selective and restricted to the
11
perception by seeds of light opportunities in cultivated
lands.
In arable lands, light at the soil surface level varies
in a more or less cyclical fashion due to the managing
practices associated with the production of a particular
crop. The major disturbances, and hence the major causes of
light opportunities in these systems are grazing, mowing,
and soil cultivation. Several factors have been proposed to
work as environmental signals in the mechanisms whereby
seeds detect these events, including soil temperature
changes, light, changes in the soil atmosphere, and
modification of chemical composition of the soil solution.
These are briefly considered in the remainder of this
section.
a. Light opportunities created by canopy removal
One of the major consequences of canopy removal is to
modify the thermal regime of the topsoil. In particular, the
daily thermal amplitude (i.e., temperature
temperaturemin) in the upper strata is much larger in bare
soil than under a dense vegetation canopy. Seeds of weed
species usually germinate better at alternating than at
constant temperatures (e.g. Aldrich 1984). This observation
led Thompson et al. (1977) to propose that, under natural
conditions, increased temperature fluctuations may signal to
seeds when canopy gaps occur. The best evidence for the
operation of a gap sensing-mechanism dependent on soil
temperature fluctuations was provided by Benech Arnold et
al. (1988) working with Johnsongrass (Sorghum halepense).
They found that germination of this weed was greater in
experimental canopy openings than under undisturbed
vegetation, and were able to demonstrate that the promotive
effect of clipping could be reproduced under the canopy by
manipulating the soil temperature regime to mimic the
temperature fluctuations that occurred in the adjacent bare
plots.
In other cases, the environmental factor that informs
seeds about the occurrence of openings in the leaf canopy
appears to be light itself. From an optical point of view,
green leaves are made of a strongly scattering material with
12
well defined absorption bands (Knipling 1970). Flavonoids
and photosynthetic pigments absorb most of the radiation in
the UV and visible wavelengths (i.e. 290-700 nm), and water
absorbs strongly in the infrared (> 1200 nm). There is very
little absorption between 700 and 1200 nm so that most of
the FR emerges from either side of the leaf in the form of
scattered radiation. This accounts for the low R:FR ratios
that are observed in transmittance and reflectance spectra
of green plant organs and in light measurements taken
beneath vegetation canopies.
Classic studies on the photocontrol of seed
germination, carried out mainly on freshly dispersed seeds
or with seeds kept in dry storage for some time, have shown
that, in order to be effective, light must establish a
relatively high level of the Pfr form of phytochrome (see
references in Bewley and Black 1982). Thus, the potential
effect of light can be nullified if the Pfr content is
immediately reduced to about 2 % with a saturating pulse of
FR radiation. Seeds exposed at the soil surface to light
filtered through a leaf canopy (low R:FR ratio) would be
prevented from germinating. Opening the canopy will increase
the R:FR ratio and this would form enough Pfr in the seeds
to promote germination (see Vasquez-Yanez 1993). The
evidence for a gap-sensing mechanism based on perception of
light quality changes in herbaceous plant communities under
natural conditions is scant. Perhaps one of the drawbacks of
a R:FR-driven dormancy mechanism is that seeds would be
required to stay at or very near the soil surface in order
to perceive changes in light environment. Rates of seed
predation have been shown to be overwhelming in abandoned
(Mitelbach and Gross 1984, van Esso and Ghersa 1989) and
intensively cultivated fields (Scopel et al. 1988), which
may prevent accumulation of significant seed banks on the
ground surface.
Besides temperature and light, there are other
environmental factors associated in a predictable way with
variations in above-ground biomass that, on theoretical
grounds at least, might act as effective signals in the
control of seed germination under natural conditions. Pons
(1989) proposed that by responding to NO3 concentration in
13
the soil, seeds might detect gaps in the canopy in
herbaceous communities. Organic acids produced by decaying
vegetation (e.g. Simpson 1990) or reduction in levels of
allelopatic substances (e.g. Angevine and Chabot 1979) might
also act as germination signals, but the evidence for these
mechanisms remains largely circumstantial.
b. Light opportunities preceded by soil disturbances
In agricultural land the event that most frequently
heralds a period of high light availability is disturbance
of the topsoil by tillage operations. That seeds have the
sensory machinery capable of detecting these events is
fairly obvious when one looks at the patterns of seed
germination in arable soils. Tillage during the season(s) of
germination usually results in large cohorts of seedlings of
weedy species (e.g. Chancellor 1964, Roberts and Potter
1980, Ballard et al. 1988b). In some species, seeds must
spend some period of time in the soil before they can be
induced to germinate by exposure to light or temperature
fluctuations. Thus, in these cases, germination seems to
depend on the occurrence of at least two tillage episodes:
one to bury the seeds and other to provide the necessary
environmental signal to the (in some way sensitized) seeds.
This suggests a remarkable degree of adjustment to the
intensively-disturbed agricultural environment (Soriano et
al. 1970). What are the mechanisms that allow such a degree
of synchronization?
Seeds of many species enter the soil bank with a high
degree of primary dormancy that may be imposed by a variety
of factors. During burial, seed populations undergo dramatic
changes in the dormancy status, as gauged from results of
germination tests on exhumed seed samples. In some cases
these changes are cyclical, with dormancy reaching a minimum
by the time of the year when germination usually occurs, and
then increasing again in seeds that remained in the soil
bank after that period (secondary dormancy) (reviews by
Karssen 1982, Baskin and Baskin 1985).
Germination tests carried out on unearthed seed samples in
the laboratory are useful to show variations in the
germination behavior over time, particularly the broadening
14
and narrowing of environmental conditions in which
germination can occur. Nevertheless, these tests may provide
little insight into the nature of the environmental
conditions that actually trigger germination under real
field conditions (see Bouwmeester and Karssen 1989). At the
scale of the individual seed, tillages produce a great
variety of environmental changes, and different species or
ecotypes appear to have evolved to rely on different signals
to detect soil disturbances.
The mechanism that depends on temperature fluctuations
(Section II.B.1.a.) is likely to be important for many
species. Daily thermal amplitude diminishes rapidly with
depth and, particularly at the beginning of the growing
season, seeds buried more than a few cm are subjected to
very small temperature fluctuations in most soils. Tillages,
particularly plowing, can carry seeds from deep to shallow
soil strata, where higher thermal amplitudes may elicit
germination (see Koller 1972, Ghersa et al. 1992).
In many cases light appears to play a decisive role in
the mechanisms whereby buried seeds detect cultivation
episodes. Classic simulation experiments by Sauer and Struik
(1964) and Wesson and Wareing (1969) have shown that, if
steps are taken to exclude the light, fewer seedlings emerge
from disturbed soil samples. More recently, Scopel et al.
(1991) induced germination of buried seeds of Datura ferox,
an annual weed of summer crops, by piping sunlight from the
soil surface without disturbing other microenvironmental
factors during illumination. This demonstrated that, for a
fraction of the seed population, germination in the field
was limited only by the lack of light. The experiment was
carried out during the spring, with large daily temperaturefluctuations even at -10 cm. When seeds were unearthed using
light-tight equipment, exposed to sunlight, and placed again
at the initial depth (simulating the effects of soil
cultivation), germination exceeded 90%. The most significant
aspect of these results is the observation that a short
period of burial produced a dramatic (ca. 10,000-fold)
increase in the sensitivity of Datura ferox seeds to light.
In fact, after a period in the soil, the majority of the
seed population could be induced to germinate in the field
15
by irradiations equivalent to 0.1 to 10 ms of full sunlight
Fig. 2. 2). The amount of Pfr established by these short
irradiations would be on the order of 0.0001 to 0.01 %, that
is 2 to 4 orders of magnitude lower than the level
established by a saturating exposure to pure FR (Scopel et
al. 1991).
It is now well established that many plant responses to
light, particularly in dark-grown material, can be elicited
by irradiations that form only minute amounts of Pfr and,
therefore, can not be reversed by FR. This type of
photoresponse has been termed very-low-fluence (VLF)
response, to distinguish it from the classic, FR-reversible,
low-fluence (LF) response (Mandoli and Briggs 1981, BlaauwJensen 1983, Kronenberg and Kendrick 1986). Seeds of several
species displaying VLF responses have been obtained in the
laboratory by means of artificial pretreatments. Based on
the fluence response curves obtained in laboratory and field
conditions, Scopel et al. (1991) interpreted the
sensitization observed in buried seeds as a natural
transition from the LF to the VLF mode of phytochrome
action. Previous laboratory experiments by Taylorson (1972)
and Baskin and Baskin (1979) also indicate a very high light
sensitivity in seeds retrieved from soil. Theoretical
calculations of the amount of light that individual seeds
would get during soil tillage suggest that VLF responses are
of great importance for the perception of light signals of
soil cultivation (Scopel et al. 1991).
2. Stem phototropism and leaf movements
After germination, the transition to the
photoautotrophic stage, that involves leaf and plastid
development and reduced axis elongation is also controlled
by light acting through photomorphogenic pigments. Once
seedlings have emerged from soil, they have to adjust to a
complex light environment that is characterized by extreme
variations in intensity and direction of the incident rays.
These variations are caused by changes of solar angle during
the course of the day, degree of cloudiness, and light
absorption and scattering by neighboring objects and plants.
In this section, I briefly discuss how plants or individual
organs detect the direction of illumination and how they
16
react with morphological changes that improve light
interception. Typical morphological responses to diurnal
variations of light direction are phototropic movements of
young stems and modifications of leaf inclination and
azimuth.
Phototropism is the bending of elongating shoots
towards the brighter side in an uneven light field. This
bending results from redistribution of growth, i.e.
inhibition on the illuminated flank and parallel stimulation
on the shaded side, with little change in net growth (Rich
et al. 1987 and references therein; see also Briggs and
Baskin 1988, lino 1990). The subject has been reviewed
fairly recently (Firn 1986, lino 1990) and only some general
aspects will be considered here.
Most of our understanding of phototropic mechanisms in
higher plants comes from experiments with etiolated (darkgrown) seedlings. These studies have shown that detection of
the direction of illumination is based on fluence rate
gradients that are created between the illuminated and dark
flanks of the phototropic organ (i.e. coleoptile tip, stem).
These gradients are the consequence of internal light
scattering (mainly at the air / cell-wall interphases) and
absorption (Seyfried and Fukshansky 1983, Vogelmann and
Haupt 1985, Vogelmann 1986). The B / UV-A region of the
spectrum, sensed by one or more as yet unidentified specific
receptors, is used by plant phototropic-systems to obtain
information about changes in light direction (e.g. lino
1990). Most studies indicate that phytochrome does not
function as a detector of the direction of illumination in
etiolated seedlings (see references in lino 1990), although
this idea has been challenged by recent studies with darkadapted corn (Zea maize) (Iino et al. 1984) and etiolated
pea (Pisum sativum) (Parker et al. 1989) seedlings. What is
clear is that phytochrome is involved in accessory processes
(e.g. sensitivity and responsivity adjustment) that may be
essential for the phototropic machinery to operate under
natural light conditions (see Woitzik and Mohr 1989, lino
1990, and references therein).
In light-grown, green seedlings phytochrome appears to
17
participate in the detection of internal light gradients.
Early action spectra obtained using broad-band light sources
show a peak in the R region in addition to the B / UV-A peak
(Atkins 1936). The hypothesis put forward by lino (1990) to
explain how phytochrome may act as a phototropic sensor in
plant receiving polychromatic light is based on the
differential penetration of light of different wavelengths
into green organs. Due to absorption by chlorophyll, the
fluence rate of R light declines abruptly within the first
mm of tissue, while the fluence rate gradient in the FR
region is comparatively much less steep (Seyfried and
Fukshansky 1983, Vogelmann and Bjorn 1984, Vogelmann 1986).
Therefore, in shoots exposed to unilateral light, there
would be a gradient of R:FR ratio and 0 between the
illuminated and shaded flanks of the organ. Since low Os
stimulate elongation (Section II.B.3.), the shaded side
would elongate faster than the illuminated one, and this
will cause the shoot to bend towards the lateral light
source. Although this proposed mechanism is consistent with
the available data we can not yet conclude that it actually
plays a role in determining phototropic responses in nature.
Plants are seldom exposed to truly unilateral illumination.
The "shaded" flank of the stem may actually receive
substantial amounts of diffuse radiation which, in open
conditions, will be unaltered skylight, with even higher
R:FR ratios than direct sunlight (Holmes and Smith 1977a).
Recent results with potted cucumber (Cucumis sativus)
seedlings provide some indication about the involvement of
different photoreceptors in natural phototropism (Ballare et
al. 1991b). Seedlings grown in a glasshouse with no
supplemental lighting naturally bent along the hypocotyl
towards the Northern side (the experiments were carried out
in the Southern hemisphere). The degree of curvature in
seedlings of the lh-mutant, which is deficient in a stable
phytochrome (see Section II.B.3.a.), was the same as in
wild-type individuals. Furthermore, in both genotypes,
bending was totally prevented by removing the B component
from direct sunlight with a cellulose acetate filter. This
suggests that the B waveband, acting through a sensor other
than phytochrome, is essential for phototropic responses, at
least in plants grown in sparse settings. The situation
18
might be different for plants growing in the vicinity of
other vegetation. In fact, light scattering by nearby plants
can substantially increase the amount of FR that impinges
laterally on vertically oriented shoots (Figs 2.5 and 2.6,
Section II.B.3.b.). This will create R:FR gradients across
stems that would be related to the distribution of neighbors
around the plant. For instance, if a plant is growing at the
edge of a gap, the R:FR ratio received at the stem surface
will tend to be lower on the side that faces the canopy than
on the side that faces open space. Under these conditions,
the phytochrome-dependent phototrophic mechanism considered
by lino (1990) might, in addition to the B-driven system,
signal the elongating organ to bend towards the gap.
Stem bending responses to directional light signals are
probably an important component of the mechanisms that help
the plant to forage for light and improve PAR capture in
heterogeneous light environments. The above, admittedly
superficial consideration of phototropism suggests that more
work is needed in order to clarify the contribution of
different photoreceptor systems to the responses observed
under natural conditions.
Apart from being able to rapidly change the direction
of growth of young stems, many plants can adjust the spatial
orientation of their leaves in response to directional light
signals (reviews by Ehleringer and Forseth 1989, Koller
1990). Diaphototropic leaf movements reorient the lamina
throughout the day so as to maintain it nearly perpendicular
to the direct sunlight vector (solar-tracking). Solar
tracking by young leaves is analogous to phototropic stem
movements because reorientation depends on redistribution of
growth between opposite flanks of the petiole. In mature
leaves, reorientation is based on reversible structural
deformations of a specialized tissue, the pulvinus, located
between the lamina and the petiole (or at the base of each
leaflet in species with compound leaves). Information about
changes in the direction of illumination is acquired by
leaves through a specific receptor for B light. The
available information about sites of light perception and
the fascinating complexity of the mechanisms of response
were reviewed recently by Koller (1990).
19
Solar tracking contributes to maintain a high and
relatively constant PAR interception during the day and may
also have some positive influence on photosynthesis via
increased leaf temperature at the extremes of the
photoperiod (Mooney and Ehleringer 1978, Shackel and Hall
1979). The adaptive value of solar tracking has been
frequently considered in relation to the ecology of species
that occur in arid habitats with short growing seasons (e.g.
Mooney and Ehleringer 1978, Ehleringer and Forseth 1980,
1989). Solar tracking is also common in seedlings of pioneer
species of secondary succession, and most likely contributes
to increase plant growth rate during the early stages of
canopy development.
3. Stem elongation
Rapid responses to directional light signals are
important to adjust the orientation of light harvesting
organs to increase PAR interception in temporally variable
environments. In many situations, particularly in rapidly
growing canopies formed by plants of similar stature, there
may be little advantages to change orientation without rapid
growth in height. Not surprisingly, the key feature of
elongation growth in species of early succession is
plasticity and responsivity to local canopy conditions.
The rate of stem elongation is influenced by many
factors, including all those that affect total plant growth
(i.e. PAR, water and nutrient supply), and a number of
others with specific morphogenic activity. Some of these
factors have more than one effect: light, for instance, may
stimulate stem elongation via promoting total growth but,
under certain conditions, increased light intensity leads to
inhibitory morphogenic effects. In the short term at least,
shade-intolerant species with orthotropic stems typically
respond to leaf shading or increased population density by
accelerating internode elongation. Plants with
diagraviotropic stem systems may show the opposite reaction
to shading (i.e. decreased elongation), and elevation of the
foliage depends on changes in shoot inclination (Section
II.B.5.) or increased petiole length (Solangaarachchi and
Harper 1988, Thompson and Harper 1988, Methy et al. 1990).
20
a. Responses to shading in multi-story canopies
Stimulation of stem elongation in the lowest strata of
multi-story canopies may be caused by several
microenvironmental changes brought about by the presence of
a leaf cover. Under controlled conditions, stem elongation
rate is highly sensitive to changes in air humidity,
mechanical disturbances, total irradiance, and spectral
light quality. Although the effects of air humidity (Mc
Intyre and Boyer 1983) and mechanical perturbations (e.g.
movements caused by wind, Neel and Harris 1971, Telewsky and
Jaffe 1986) are well documented, little is known about the
extent to which responses to these variables contribute to
the putative effect of shading on elongation under field
conditions.
Differences in light environment between open and leafcovered sites do certainly influence elongation growth. The
optical characteristics of leaves (Section II.B.1.b.)
determine that, compared to open situations, the radiation
environment beneath canopies is characterized by low levels
of UV and visible wavelengths and reduced R:FR ratios (e.g.
Vezina and Boulter 1966, Holmes and Smith 1977b). All these
factors are known to influence elongation, at least under
controlled conditions.
The UV-B waveband inhibits shoot elongation in many
species. Evidence is accumulating to suggest that this is
not merely a by-product of unspecific growth inhibition, but
a true photomorphogenic response to UV-B (e.g. Steinmetz and
Wellmann 1986, Barnes et al. 1990, Ballard et al. 1991a),
which most likely involves the action of a specific UV-B
receptor. Attenuation of UV-B by a leaf canopy would thus
lead to increased elongation in sensitive species growing in
areas that naturally receive high UV-B levels.
Another aspect of the light environment beneath leaf
canopies that may stimulate elongation is the reduced
fluence rate in the 320-800 nm waveband (which comprises the
UV-A, PAR and FR regions, and that I will call "white light"
for brevity). Short-term experiments with young seedlings
(grown in the light for a few hours) have demonstrated
marked inhibitory effects of increased white light
21
irradiance on hypocotyl elongation (e.g. Meijer 1959). These
effects are mediated by a specific photoreceptor(s)
operating in the B / UV-A region (Gaba and Black 1979,
Thomas and Dickinson 1979, Ritter et al. 1981) and
phytochrome(s) (e.g. Ritter et al. 1981, Wall and Johnson
1981, Holmes et al. 1982, Gaba and Black 1985; see also
Cosgrove 1986). The effects of white light irradiance on
more mature, fully de-etiolated plants are not as clear (see
references in Ballare et al. 1991c), and paradoxically,
there is some controversy about the actual significance of
this variable in the control of stem elongation under leaf
canopies (see Frankland 1986, Child and Smith 1987). Britz
(1990) has recently shown that stem elongation in fully deetiolated soybean plants can be substantially promoted by
reducing white-light fluence rate, and suggested that this
response was, at least in part, mediated by a specific B
light receptor.
The effect of reduced R:FR ratio in promoting stem
elongation beneath vegetational canopies has received
considerable attention in recent years and the subject has
been reviewed by Smith (1982, 1986) and Smith and Morgan
(1983). The work of Smith and associates has substantiated
the following key points. (i) The reduction of R:FR ratio
beneath canopies is a function of the amount of leaf area
and is closely correlated with the reduction of PAR (Holmes
and Smith 1977b). (ii) Changes in R:FR beneath canopies are
within the range of high phytochrome sensitivity, that is
where variations of R:FR lead to approximately proportional
changes in 0 (Smith and Holmes 1977). (iii) In controlled
environments, under light levels comparable to those that
can be found beneath dense canopies (ca. 10 % of full
sunlight), plants of shade-intolerant species respond to
reduced R:FR with an increase of elongation rate that is
quantitatively related to the estimated effect of the R:FR
treatment on 0 (Morgan and Smith 1978, 1979).
The importance of light signals (relative to other
microenvironmental factors) in determining elongation
responses to leaf shading has been tested by using
photomorphogenic mutants. The lh mutant of cucumber (Adamse
et al., 1987) has a longer hypocotyl compared to its near
22
isogenic wild-type (WT), and is deficient in a stable type
of phytochrome (Adamse et al. 1988, Peters et al. 1991,
Lopez et al. 1991), that is in the form that appears to
mediate stem elongation responses to R:FR in light-grown
plants. Seedlings of the lh mutant do not respond to R:FR
treatments (Adamse et al. 1987, Lopez-Juez et al. 1990,
Ballare et al. 1991b). Furthermore, the lack of stable
phytochrome does, in some way, eliminate the elongation
responses to B light mediated by other photoreceptor(s)
(Ballare et al. 1991b; see Mohr 1986 and Gaba and Black 1987
for discussions about interactions among photoreceptor
systems). The lh-mutant is therefore very useful as an
experimental tool because, except for the responses to UV-B
radiation, it appears to be (naturally) blind to
photomorphogenic signals of leaf shading. When WT seedlings
under-story of a
were transferred from full sunlight to the
dense canopy in a glasshouse experiment (Ballare et al.
1991b), the average rate of hypocotyl elongation was
promoted by a factor of ca. 3.5. This reaction was totally
absent in the lh mutant. Applications of gibberellin A3
promoted elongation of lh seedlings in the under-story,
suggesting that the lack of response to shading was not a
consequence of assimilate limitation, but a reflection of
the inability of the mutant to sense variations in the B and
R-FR environment. Qualitatively similar results were
obtained in parallel field experiments during the summer at
Buenos Aires. From these studies one is forced to conclude
that mechanisms controlled by light signals play a major
role in determining short-term elongation responses to
shading (Ballare et al. 1991b).
One of the issues that received little attention in the
above discussion is the effect of the illumination history
on the sensitivity of plants to light stimuli. The responses
to shading in the cucumber experiments were measured two
days after the beginning of treatments. In the long term
(i.e. 1 or 2 weeks) the differences in elongation rate
between "sun" and "shade" plants tended to disappear
(Ballare et al. 1991b). This might reflect either that
shaded plants could not elongate more rapidly because of
assimilate limitations or that they had changed their
pattern of response to shading. The latter does not appear
23
to be an unreasonable possibility, since in cases of
strongly asymmetric competition (i.e. seedlings v.
established plants) the "escapist" response to shading is
likely to be of very limited value to the subordinated
plants. Photosensory adaptation refers to the processes
whereby the sensitivity of a system to light treatments is
compensated for fluctuations in the illumination background
(Galland 1989). Adaptation has been studied in detail in
various systems, but we know relatively little about how
sensitivity of higher plants to changes in irradiance and
R:FR ratio is affected by prolonged shading.
b. Early reactions in even canopies
The assertion that stem elongation responses to massive
shading are unlikely to contribute to the success of pioneer
species in the natural environment is indirectly supported
by many studies on the demography of weeds in secondary
succession. It is a common observation that plants that
emerge late in the season, when the aerial and below-ground
spaces have already been occupied by earlier individuals,
tend to make little growth and usually experience very heavy
mortality (e.g. Black and Wilkinson 1963, Weaver and Cavers
1979, Gross 1980, Peters 1984, Ballare et al. 1987b, Panetta
et al. 1988, Scopel et al. 1988). It would seem, therefore,
that when plants of early successional species become
severely shaded and experience low levels of other
resources, the time for initiating shade-avoidance reactions
is already over. Sophisticated, environmentally controlled
dormancy mechanisms (Section II.B.1.), tend to assure that
germination of pioneer species is restricted to the
beginning of periods of high light availability. Therefore,
competition for light tends to occur among individuals of
relatively similar size, in canopies with little vertical
organization and low species diversity (see Bazzaz 1990). In
this scenario, precise modulation of elongation growth is a
major strategic priority, for slight differences in height
may result in disproportionately large differences in PAR
capture between neighboring plants (Grime and Jeffrey 1965,
Ballare et al. 1988a, see also Section III). There is now
solid evidence that the control of stem elongation in even
canopies entails a great deal of mechanistic sophistication,
and that responses to the proximity of other plants can
24
begin well before the leaves become severely shaded by
neighboring individuals.
Fig. 2.3 shows the elongation responses of seedlings
of the annual weed Datura ferox when they were placed in the
center of even canopies of different densities. The average
leaf area index (LAI) of the canopies varied from 0
(isolated plants) to a maximum of 1.5, so that shading among
neighboring seedlings was very small in these experiments.
However, in that range of LAIs, increasing neighborhood
density led to a marked, rapid increase in stem elongation
rate. In a related study (Ballare et al. 1987a), seedlings
of white mustard (Sinapis alba) were grown on the Northern
(directly illuminated in the Southern hemisphere) side of
grass hedges, half of which had been bleached by spraying
with a herbicide. Seedlings grown in front of green hedges
developed longer internodes than those near bleached
neighbors even though, in both cases, the seedlings were
continuously exposed to unfiltered sunlight (Fig. 2.4). What
follows is a discussion of the role of light signals in
these early elongation responses to the proximity of other
plants.
As pointed out in section II.B.1.a., transmittance and
reflectance spectra of green leaves are very similar and
show a steep rise in the FR region. Indeed, the distinction
between transmitted and reflected light is mainly
operational (based only on the relative positions of the
light source, the leaf, and the light receiver), and should
not obscure the important fact that the two components have
the same origin and, therefore, very similar spectral
properties. Ballare et al. (1987a) hypothesized that the
differential reflection of R and FR radiation, acting
through phytochrome, provides an environmental cue for the
detection of non-shading neighbors in the early stages of
canopy development. Measurements obtained with conventional
light sensors (e.g. Kasperbauer et al. 1984) or cuvettes
containing phytochrome preparations (Smith et al. 1990)
pointed at illuminated plant parts typically show low R:FR
or Pfr/P ratios. However, to predict the effects of
neighboring plants on the state of phytochrome in a
particular organ of an intact, green plant is usually
25
cumbersome. To a large extent this is because plants are
much more complex structures than a flat light receiver.
Individual organs are usually exposed, at the same time, to
light scattered by nearby objects and foliage and to direct
sunlight. Ballare et al. (1989) used a fiber optic probe
implanted in different plant organs and attached to a
spectroradiometer in order to investigate how the quality of
radiation scattered within plant tissues was affected by the
proximity of other plants. One limitation of the fiber optic
approach is that we do not know the internal distribution of
the phytochrome molecules involved in the control of
straight growth. However, it is the only way to visualize
the integrated product of light beams entering the plant
from all possible directions. Fiber optic studies showed
that, in even canopies, the proximity of neighboring plants
is unlikely to bring about a significant spectral change
inside leaves that are almost perpendicular to the direct
light vector if shading among neighbors is small. This is
due to the overwhelming contribution of direct sunlight to
the internal light regime.
The situation is very different in the case of vertical
stems or leaves. In these cases, light reflected by
neighboring plants may cause a substantial increase in the
amount of FR light scattered inside the tissues, even if the
LAI of the canopy is very low (Fig. 2.5). Calculations
showed that spectral changes of the magnitude shown in Fig.
5 may cause a 30 % decrease in the projected value of 0
inside the stem (Ballare et al. 1989, see also Mancinelli
1991).
Stem internodes have some degree of autonomy to respond
to local light-quality conditions (Garrison and Briggs 1975,
Morgan et al. 1980, Casal and Smith 1988a,b). Figure 2.6
shows a detailed picture of the effects of increasing canopy
density on the light environment of the stems in even-aged
populations of Sinapis alba and Datura ferox. Measurements
were taken with the fiber optic probe (Fig. 2.6c), or with
an integrating cylinder (Ballare et al. 1987a) that accepts
side-light received from all compass points (Fig. 2.6b). It
can be seen that the only detectable change when LAI
increases in the very-low range (i.e. LAI between 0 and 1)
26
received by
is a substantial increase in the amount of FR
the stems. The canopy works as an effective FR trap, and the
R:FR ratio at the stem surface typically drops from ca. 0.9
to 0.4 (Ballare et al. 1987a). Other wavelengths are
of the
virtually unaffected in this LAI range. If the LAI
rapid
reduction
in the
canopy increases further, there is a
fluence rates of B, R, and FR radiation received by the
all these drastic
stems. The most significant aspect is that
changes in the stem's light environment take place well
before shading at leaf level becomes a factor. In fact,
measurements of light interception by leaves (Fig. 2.6a)
there is only a
show that even in canopies with a LAI of 2,
small effect of neighbors on light available for
photosynthesis.
How do all these changes in the stems' light
projection
environment influence elongation rate and, hence,
here
some
of new leaves into uncolonized space? I describe
to address
of the experiments which have been carried out
this question. In one series of field experiments (Ballare
species from open
et al. 1987a), seedlings of three annual
habitats, Sinapis alba, Datura ferox, and Chenopodium album
mirrors that
were grown in front of selective-reflecting
provided additional FR so as to mimic the presence of other
seedlings (cf. Figs. 2.5, 2.6), without affecting PAR
receipt at leaf level. The mirrors reduced the R:FR ratio
received by the stems (measured with the integrating
cylinder) from ca. 0.8 to 0.5 and the estimated 0 inside the
with the fiber
stem (calculated from spectral scans obtained
optic probe) from ca. 0.34 to 0.25 (see Ballare et al.
1989). Plants exposed to additional FR produced longer
internodes than the controls grown either without or in
front of mirrors that reflected R and FR light with
have
approximately similar efficiencies. Comparable results
FR
been obtained in experiments were the additional
treatment was applied during the central hours of the
photoperiod only (Ballard et al. 1991c).
In a
of Datura
period of
plants of
series of glasshouse studies, 2-week old seedlings
short
ferox and Sinapis alba were placed for a
formed
by
time in canopies of different densities
similar stature. Transplanting led to a rapid
27
increase in the rate of stem elongation that was related to
the LAI of the surrounding canopy. This reaction to the
proximity of neighbors was much reduced when the internodes
of test plants were covered by a FR-absorbing filter collar
that maintained a high R:FR ratio at the stem level (Fig.
2.7). These experiments demonstrated that in seedling
canopies of very low LAI (i.e. LAI < 1), where interference
among neighbors for PAR capture is almost nil, the R:FR
ratio perceived at the internode level plays a major role
driving adjustments of stem elongation rate to local density
conditions (Ballare et al. 1990).
In canopies with LAI > 1, the efficiency of the FRabsorbing filters was generally reduced, even when they
still maintained a relatively high R:FR (ca. 0.9). It
appeared that some factor, other than the reduced R:FR,
promoted elongation growth in those populations. Light
measurements presented in Fig. 2.6 showed that, when LAI
reaches about 1, PAR interception by leaves continues to be
high, but total irradiance at the stem level begins to
decline rapidly. In most of the experiments that addressed
the influence of total irradiance on elongation growth, the
light treatments were applied to the whole shoot (Section
II.B.3.a.), which is clearly not equivalent to the effects
of neighboring plants in even canopies. In experiments under
natural radiation, annular filters that reduced the fluence
rate of side-light by less than 50 % without influencing PAR
interception by leaves promoted internode elongation
(Ballare et al. 1991c). This response to fluence rate was
not obviously affected by R:FR in the range of ratios
typical of canopies of low LAI (i.e. 0.3 < R:FR < 0.9). This
suggests that, when the canopy begins to close, both R:FRand fluence-rate-dependent mechanisms modulate elongation
responses to neighbors proximity (Ballare et al. 1991c).
What are the photoreceptors responsible for the
perception of changes in fluence rate? Photosynthetic
pigments are unlikely candidates. In fact, shading the stems
with a pink collar that reduced PAR by filtering the green
waveband (with minimal effects on spectral regions absorbed
by phytochromes and B / UV-A receptors), did not result in
increased elongation in Sinapis alba (Ballare et al. 1991c).
28
Phytochrome(s) and B / UV-A sensors have been implicated to
explain the fluence-rate-dependent photoinhibition of
hypocotyl elongation in seedlings de-etiolated for a few
hours (see Section II.B.3.a.) and the same photoreceptors
would seem to control elongation responses to stem shading
in the case of more mature, fully de-etiolated plants
(Ballare et al. 1991c).
4. Branching and production of new ramets
In the long term, most species rely on the production
of new branches for colonization of open space in canopies,
and it is well established that local conditions of crowding
have a dramatic influence on the amount of branches that a
plant produces throughout its life span (e.g. Langer 1966,
Harper 1977). Multiple factors, including responses to
resource levels and morphogenic signals, are most likely
involved in the control of branching and ramet production
under field conditions. The possible involvement of external
morphogenic signals was first suggested by Kirby and Faris
(1972) after observing that, in barley (Hordeum vulgare)
crops, the effects of planting density on the fate of
individual tiller buds appeared to be decided quite early
during the bud --> tiller transition. In this section I
briefly cover the effects of light (sse Casal et al. 1986,
and Hutchings and Slade 1988 for more complete treatment).
Reduced total irradiance has been shown to result in
increased apical dominance and reduced production of new
branches in growth cabinet (e.g. Mitchell 1953), greenhouse
(Slade and Hutchings 1987, Methy et al. 1990) and field
experiments (Bubard and Morison 1984, Thompson and Harper
1988). It is not clear whether this detrimental effect of
shading is a consequence of assimilate limitations (which
might act as an internal signal) or a more direct response
to light fluence-rate mediated by specific photoreceptors.
Direct responses to B and R fluence rates might contribute
to determine that the inhibitory effects of leaf shading on
branching are usually more pronounced than those caused by
roughly similar reductions in PAR obtained with neutral
filters (e.g. Solangaarachchi and Harper 1987). Direct
responses to UV-B fluence rates should also be considered
when interpreting results of field studies, because moderate
29
levels of these wavelengths are known to promote tillering
in some species (Barnes et al. 1990).
Alterations of the ratios between wavelengths (i.e.
Mansfield
R:FR ratio) influence apical dominance (Tucker and
1972) and are certainly involved in the branching responses
to the proximity of other plants. Deregibus et al. (1983)
demonstrated that tillering in grasses is depressed by FR
light acting through phytochrome in a R:FR reversible manner
(see also Casal et al. 1985, Kasperbauer and Karlen 1986,
Casal 1988). Deregibus et al. (1985) also showed that
photomorphogenic mechanisms do play a role in the control of
tillering under field conditions. Working in a natural
grassland, they were able to increase tillering rates during
the growing season by supplementing the radiation received
at the base of Paspalum dilatatum and Sporobolus indicus
plants with small amounts of R light from light emitting
diodes that increased the R:FR ratio.
The interactions between light signals and resource
have
levels in the control of branching and ramet production
not been thoroughly investigated in any system. The
experiments of Casal et al. (1986) with Paspalum dilatatum
show that in grass canopies of low LAI, where mutual shading
other
is small, tillering responses to the proximity of
plants are most likely mediated by the altered R:FR balance.
Grasses were shown to be very sensitive to small reductions
of R:FR (Casal et al. 1987), particularly if the treatments
are applied during daytime (as opposed to end-of-day pulses)
(Casal et al. 1990). Thus, changes in light environment
produced, for example, by the differential reflection of R
and FR light by nearby leaves (e.g. Figs 2.5 and 2.6) may
signal grass plants to reduce tillering rate early in the
development of the canopy (see Casal et al. 1987). This is
probably advantageous for the internal economy of the plant
in environments where late branches have little chance of
contributing to reproduction. It should be noted that the
factors that influence branch or tiller mortality in
canopies are still unclear. The popular belief that branch
senescence is triggered by resource (light) starvation has
been questioned recently by Lauer and Simmons (1989).
30
5. Spatial orientation of branches
The phototropic responses of leaves and stems
considered in Section II.B.2. are examples of rapid,
reversible movements that allow plants to re-arrange their
architecture in continuously changing light conditions.
Plants can also change the orientation (zenith and azimuth
angles) of whole branches in response to more permanent
features of their light environments. Casal et al. (1990)
found that the tillers of annual ryegrass (Lolium
multiflorum) tended to adopt a more erect position in the
canopy as plant density was increased, and there is some
evidence that R:FR signals perceived by phytochrome activate
mechanisms that control shoot inclination (e.g. Casal et al.
1990, Aphalo et al. 1991). Effects of total irradiance may
also be involved in the responses of shoot inclination to
that neutralcanopy density. Grime et al. (1986) reported
shading treatments led to the production of more vertical
shoots in a number of bryophytes that, under natural
conditions, tend to exploit the upper strata of pasture
canopies. Montaldi and co-workers suggested that, in grasses
of
with diagraviotropic stems, the angle of inclination
shoots is modulated by sucrose content (Montaldi 1963,
Willemods et al. 1988), and postulated that the effects of
irradiance and R:FR ratio on inclination are consequences of
the effects of these variables on carbohydrate production
and partitioning (e.g. Beltrano et al. 1991).
An interesting example of how plants use directional
light signals to adjust the orientation of branches in
patchy canopies was provided by Novoplansky et al. (1990).
oleracea, a weed of
They found that seedlings of Portulaca
intensively disturbed areas with a diagraviotropic shoot
neighbors when
system, tended to avoid growing towards their
colonizing bare soil. This appeared to be a consequence of
effects of neighbors on (i) the azimuthal orientation
adopted by the main stem at the time it became horizontal
and (ii), the pattern of lateral shoot initiation. The
after
authors implicated light signals in these responses
observing that seedlings also tended to grow away from green
plastic objects that absorbed PAR and reduced the R:FR
with gray
ratio. Furthermore, when seedlings were confronted
transmission
and green plastics that provided similar PAR
31
with different effects on the R:FR ratio, only a small
proportion of the plants became recumbent toward the green
(low R:FR) sector (Fig. 2.8). Since the green plastic
reflected more FR than the neutral gray, the response
observed in Portulaca might have been a consequence of the
phytochrome-mediated phototropism discussed earlier (Section
II.B.2).
III. COMPETITION FOR LIGHT DURING EARLY SUCCESSION
In Section II, I have condensed the available
information about the mechanisms whereby plants of early
succession perceive and react to changes in light conditions
with emphasis on germination responses of arable weeds and
morphological plasticity of vegetative shoots. In this
Section, I present an overview of how species of early
secondary succession apply the acquired environmental
information to actively locate and exploit light
opportunities in highly dynamic canopy systems.
A. The acquisition of information and the race for light
By definition, secondary successions are generated by
some type of disturbance that brings about an increase in
light availability at the ground surface. Disturbance is
followed by recolonization of the site by individuals of
pioneer species (and cultivated plants in agricultural
environments) that rapidly begin to compete with each other
for access to light and other resources. We can visualize
the process as a race, where the success of an individual
relative to its neighbors depends essentially on (1) its
relative ability to capture light energy at the beginning of
the period of interaction (which is mainly a function of its
relative initial size), and (2) its relative ability to
maintain high rates of light capture as the canopy develops.
Differences in initial size are largely due to
differences in propagule (seed) size and emergence time (or
emergence order). The production of large seeds carrying
large embryos may have a number of associated costs (Harper
et al. 1970, Thompson 1987) and small seeds appear to
prevail among pioneer species in many different systems
(Fenner 1987). As mentioned earlier (Section II.B.1.),
germination does not occur at random through time in
32
habitats subjected to periodic disturbance. Environmental
signals such as light and daily temperature fluctuations
synchronize germination events with variations in light
receipt at the ground surface. Field experiments are
beginning to uncover the complexity of the perceptual
processes involved and are contributing to determine which
are the relevant cues for different species and
environments. Given the importance of relative time of
emergence in determining growth and fecundity (see
references in Section II.B.3.b. and Weiner 1988), one can
hardly dispute the notion that the ability of seeds to input
environmental information into the systems that control the
degree of dormancy is a key element of success in frequently
disturbed ecosystems.
Maintaining high rates of light interception as the
canopy develops is obviously necessary to sustain growth.
The amount of leaf area generated per unit of dry matter
produced (i.e. the leaf area ratio), a quotient that is
strongly influenced by light conditions (Blackman and Wilson
1951, Evans and Hughes 1961), is certainly a major
determinant of the dynamics of light capture. Light
interception per unit leaf area depends on how foliage
pieces are spatially arranged in relation to the pattern of
light availability. Since this pattern is characterized by
major fluctuations imposed by solar movement, atmospheric
factors and rapid growth of neighboring plants, a high
degree of morphological plasticity, based on continuous
acquisition and interpretation of environmental information,
is most likely an essential requirement for species of early
secondary succession. In Section II, I discussed how
information acquired by B / UV-A receptors (and probably by
phytochrome as well) is used to orient leaves and young
growing stems to maximize PAR capture in variable light
environments. Changes in branch inclination (zenith angle)
in response to local conditions of crowding are also known
to occur, and although the picture is still incomplete,
there is clear evidence that photomorphogenic signals are
involved in the controlling mechanisms (Section II.B.5.).
Perception of patchiness in the light environment may lead
to non-random spatial patterns of vegetative spreading in
clonal plants. Phytochrome-perceived light cues are known to
33
mediate tillering responses to canopy density under certain
conditions (Section II.B.4.) and they also appear to
influence the processes whereby plants with diagraviotropic
shoot systems choose a direction for spreading (Section
II.B.5.). Considerable evidence has been accumulated to
suggest that these morphogenic events may take place very
early in the process of space colonization. Thus, the
observed patterns of branching and site occupation reflect
both the limitations imposed by the resources available at
each particular spot of the environmental mosaic, and the
"decisions" that the invading shoots took even before
experiencing discontinuities in resource supply.
Stem elongation is important for maintaining young
leaves within the reach of direct sunlight in rapidly
growing even canopies. Although stem internodes in
herbaceous plants appear to be relatively cheap in terms of
the opportunity value of the carbon invested in their
construction (Ballare et al. 1991d), different types of
indirect costs are most likely associated with the
production of longer stems (e.g. reduced mechanical
strength, increased exposure to herbivores). Therefore, for
each particular plant in a population, there should be an
optimum rate of stem elongation dictated by the rate of
growth in height of the surrounding vegetation (see
discussions by Givinish 1982, and Waller 1988). I have
presented evidence in Section II.B.3.b. that light signals
of different nature, perceived by phytochromes and B / UV-A
sensors, modulate the rate of stem elongation in even
canopies of dicotyledonous seedlings. Furthermore, it was
shown that elongation responses to changes in canopy density
and vertical distribution begin before the amount of light
available to leaves for photosynthesis is affected by
neighboring individuals. These early reactions are, to a
large extent, a consequence of effects of adjacent plants on
the stems' light conditions and the ability of stem sections
to respond to local light signals. Thus, in a sense, when
responding to these local light changes, plants are
"guessing" about future variations in PAR receipt at leaf
level. Morgan and Smith (1979) and Corre (1983) found that
elongation responses to experimental changes in R:FR ratio
are usually more dramatic in pioneer species than in species
34
of shaded habitats, which is consistent with the notion that
these responses are valuable only for plants that compete
for access to light against individuals of similar
morphology in relatively open canopies. Ballare et al.
(1988a) grew seedlings of the arable weed Datura ferox at
three densities in the field and monitored the time course
of stem elongation, leaf area expansion and PAR interception
per plant. Two weeks after the beginning of the study, the
LAI of the densest population was - 2 and shading among
neighboring seedlings was still small. However, a
significant effect of canopy density on seedling height was
already evident, and seedlings transferred from low- to
high-density canopies intercepted one third of the PAR
captured by their local neighbors. These results suggest
that a short lag in stem elongation would be enough to
seriously compromise seedling survival in rapidly growing
communities.
The examples discussed above suggest that the systems
that permit acquisition of information about present and
future changes in PAR availability imposed by abiotic and
biotic factors play a fundamental role as determinants of
competitive success in early successional communities.
Curiously, however, and with few exceptions (e.g. Grime et
al. 1986), the value of information-acquiring systems has
been largely overlooked in experimental and theoretical
approaches to the study of plant interactions. The likely
reasons and potential consequences of this apparent neglect
are discussed in the following section.
IV. SIGNALS, "DECISIONS", AND MODELS OF PLANT COMPETITION
Two major avenues of research have contributed the most
to shape our current perception of the mechanisms of plant
competition. Phenomenological studies, on the one hand,
provided a wealth of information on the consequences of
competition in relatively simple systems like monocultures
and mixtures of cultivated species and arable weeds (reviews
by Willey and Heat 1969, White and Harper 1970, Harper 1977,
Weiner 1988, Firbank and Watkinson 1990, Radosevich and
Roush 1990). These studies examined the effects of crowding
on plant growth, morphology and mortality. Classic
physiological experiments, on the other hand, generated
35
information on plant responses to light and other
environmental resources that are presumably or certainly
affected by changes in canopy density. These two disciplines
grew up rather independently from each other and,
traditionally, there has been very little exchange of ideas
between groups working in descriptive population or
community ecology and plant physiology. Physiological
ecologists historically appeared to be more inclined to
study plant responses to abiotic factors in extreme habitats
(e.g. alpine, desert; see Mooney 1991) than to unravel
physiological mechanisms of plant-plant interactions in more
productive environments.
During the last few years, a number of mechanistic
models of plant competition have been developed by
ecologists and agronomists. These models vary greatly in
complexity and purpose; the common denominator is that they
are designed to predict the outcome of plant competition
from assumptions about the way plants affect and respond to
environmental resources (Goldberg 1990). Models of this kind
are proving to be useful to study competition for single
resources over short periods of time in cultivated plant
associations (e.g. Rimmington 1984), or to predict results
of competitive interactions among plants with contrasting
life forms or allocation patterns along gradients of
resource supply or disturbance intensities (Tilman 1988).
However, their value as analytic tools may be limited when
the goal is to pin-point specific physiological traits that
determine variations in competitive ability.
Most competition models assume that plants are "blind"
organisms. They are assumed to grow over a fixed period at a
rate that is essentially proportional to the instantaneous
rate of resource supply. Developmental and biomass
allocation patterns are most commonly fixed (i.e. genotypeor age-dependent) or, in a few cases, some degree of
phenotypic plasticity is incorporated into the model by
assuming that allocation among organs or parts depends on
resource levels or ratios (e.g. Tilman 1988). Most commonly
plants are forced to co-occur, and the only possible way by
which plants may influence each other's activity is by
decreasing (or increasing) resource availability. These
36
assumptions are false. Previous sections (II.B.1) have
presented an overview of the systems used by seeds to
(see also
acquire information about the aerial environment
specific
Vasquez Yanez 1993). These systems, based on rather
environmental signals, were shown to be responsible for
synchronizing growth activity with periods of increased
with
light availability. In highly disturbed habitats,
differences
in
pulsed supplies of light and other resources,
formation of
ability to coordinate seed germination with the
natural or artificial gaps are likely to be far more
ability
important determinants of differences in competitive
(e.g.
than variations in resource-assimilation physiology
photosynthetic capacity) or allocation patterns.
Information-acquiring systems may help reduce the costs
associated with the production of competitive offspring.
mechanisms to
Thus, weed species that evolved efficient
afford to
detect indicators of light opportunities may
produce small seeds, which are cheap and suitable for
with chances of
dispersal but unlikely to generate seedlings
success in already occupied systems. Models or experimental
species or
designs that assume uniform germination across
part
of
the
genotypes certainly miss a very important
competition story in early successional environments.
regulation of green
Our present understanding of the
also
plant development by specific photoreceptor systems
highlight the significance of information carried by light
in plant
signals as determinants of morphological responses
canopies (Sections II.B.2. to II.B.5.). The signals that
to match the
plants use for accommodating their architecture
definition,
prevailing conditions of illumination are, by
of light
directly or indirectly associated with the amount
ask
available for photosynthesis. Therefore, one can rightly
why it is necessary to recognize the information-acquiring
systems as separate components in models of plant
competition. From a conceptual point of view it is important
in the canopy is
to keep in mind that each individual plant
they alter the
influenced by its neighbors not only because
availability of resources but also because they modify
aspects of the environment that are used by the plant to
The use of
decide among alternative developmental programs.
specific cues to keep track of variations in light supply
37
the plant over
may offer several strategic advantages to
direct monitoring of the resource, the most significant of
which is probably the ability to appropriately time the
responses. In Section II.B.3.b., I discussed how perception
rapid
of R:FR changes by individual internodes may allow
variations
in
light
stem elongation reactions that precede
interception at leaf level. Thus, particularly where light
supply is mainly a function of the size and activity of
likely to
neighboring individuals, natural selection is
favor specific mechanisms of phenotypic response to
environmental factors that are reliable and sensitive
indicators of plant proximity. These mechanisms should,
therefore, be considered as an integral part of the traits
that determine variations in competitive ability.
From a practical point of view, the distinction between
plant responses to resources and signals may be of
importance in applied fields like agronomy or forestry. In a
part of every
crop, neighboring individuals are an important
plant's environment. The way in which each plant responds to
the
proximity cues affects its performance and the yield of
whole population. Elongation growth (Section II.B.3),
tillering (Section II.B.4), developmental timing (e.g.
Mondal et al. 1986), and carbon allocation to reproductive
structures (Heindi and Brun 1983) are all affected by canopy
have
density in field crops, and different types of evidence
effects are
been presented showing that at least part of the
light
indeed photomorphogenic responses to alterations of
climate caused by nearby plants. Some of the response
mechanisms could have been selected because they conferred
competitive advantages under conditions that were different
from the ones normally faced by crop plants in modern
agricultural systems. Moreover, selection did most likely
the
favor physiological responses that were beneficial to
individual plant, rather than to the whole population. If we
accept these possibilities, we should accept that under
certain combinations of planting density and environment,
(or
crop yields are likely to be limited by the presence
lack of) particular physiological responses to specific
proximity cues.
The design of crop ideotypes is based on identification
38
of factors that limit commercial yield in a given
cultural/environmental scenario and consists of deciding
what morphological or physiological traits should be added
or dropped from the crop plant in order to make it more
efficient at converting resources into commercial yield.
Until recently, the only way available for translating
ideotypes into real plants was by repeated hybridization and
selection. But with the arrival of molecular biology
techniques, the opportunities for engineering plants with
very specific physiological traits is increasing enormously.
Therefore, knowledge on the mechanistic details of plant
responses to their neighborhood environment is likely to be
now more important than in the past. Based on the limited
information we have today, it would seem that manipulation
of photomorphogenic behavior to induce or suppress
developmental responses to signals of neighbors' proximity
may be rewarding in commercial crops.
V. SUMMARY
Under natural conditions, abiotic factors and the
activity of vegetation determine dramatic variations in the
flux of photosynthetically active radiation received at the
ground surface. Light opportunities in this heterogeneous
environment are perceived by terrestrial plants through a
great variety of specialized mechanisms. At the seed level,
aspects of the thermal regime of the soil and light signals
perceived by phytochrome are used to detect the onset of
periods of high light availability. In the particular case
of seeds of arable weeds, it seems that daily fluctuations
of soil temperature and pulses of light operating through
the very-low-fluence mode of phytochrome action, are very
important to inform seeds about the occurrence of soil
disturbances that usually prelude periods of abundant light.
At the green plant level, photoreceptors play a fundamental
role driving architectural plasticity and morphological
responses to variations in the light environment. One or
more specific photoreceptors for B light are essential for
phototropic movements of leaves and stems, which, in many
species, allow fine adjustment of leaf display to rapid
changes in the direction of incoming light. The role of
phytochrome in the detection of light gradients during
phototropism under natural conditions is not completely
39
clear at present. Precise control of stem elongation rate
may be essential for plants that forage for light in plant
communities formed by individuals of similar height. A
number recent of experiments have indicated that phytochrome
can detect FR-rich radiation scattered by neighboring plants
and trigger changes in stem elongation rate before the plant
is subjected to a reduction in light availability. In
addition to this R:FR-mediated response, fluence rate
signals, perceived by phytochrome(s) and B / UV-A sensors
may influence stem elongation in even canopies. In the case
of multi-story canopies, glasshouse and field experiments
with photomorphogenic mutants indicated that, at least in
the short term, elongation responses to leaf shading depend
to a large extent on processes in which a stable pool of
phytochrome is directly or indirectly involved. Evidence has
also been obtained over the last few years for the
involvement of phytochrome in the control branching and
lateral spreading in clonal plants.
The acquisition of information on present and future
light conditions appears to be a major priority for plants
of early successional communities, where dramatic changes of
plant cover (and hence light availability) occur over
relatively short periods of time due to disturbances and
plant growth. Mechanistic models of plant competition
usually fail to acknowledge that each plant in the community
is affected by neighboring individuals not only because they
modify the availability of energy and materials necessary
for growth, but also because they affect aspects of the
environment that are specifically used by the plant to
obtain information about neighbors' activities and to make
developmental "decisions". Responses to proximity signals
may, in turn, have a dramatic impact on the resourceharvesting capacity of the plant, and influence competitive
ability. A better understanding of the mechanisms whereby
plants acquire and "interpret" environmental information
should provide new insights into the factors that determine
differences between genotypes in the ability to compete for
light. From a practical standpoint, this knowledge would be
an useful aid to the design of more productive crop
genotypes.
40
Acknowledgements. Many of the ideas expressed in this
chapter are the product of discussions with Pedro Aphalo,
Jorge Casal, Claudio Ghersa, Steve Radosevich, Rodolfo
Sanchez, and Ana Scopel. I thank the Consejo Nacional de
Investigaciones Cientificas y Tecnicas (CONICET, Argentina)
and the University of Buenos Aires for supporting the
research presented here. I am also indebted to CONICET and
the Dept. of Forest Science for financing my stay at Oregon
State University. Les Fuchigami, Mary Lynn Roush, and Joe
Zaerr gave useful suggestions on an earlier draft of the
text and Bracher Gretchen Bracher prepared the figures.
41
UV
BA
FR
PAR
L1.1
0
Z
<
E) ..--..
<
cc E
eC c 4
z in
0HC
0E
I
o_ Z.
--I
<
E
c
c
m
m
CC
I-
0
w
-cl-
E
c
o
co
co
m
o_
E
c
c)
CO
N
CC
LL
(/)
I
200 300 400 500 600 700 800 900
WAVELENGTH (nm)
PHYTOCHROME
A MAX Pr = 660 (R)
A MAX Ph = 730 (FR)
B/UV-A P
UV -B P
FIG. 2.1. Spectral distribution of daylight and wavelengths
absorbed by known plant photoreceptors. The shaded region is
the visible portion of the spectrum. Abbreviations: B, blue;
B/UV-A P, blue/ultraviolet-A absorbing photoreceptor(s); FR,
far-red; PAR, photosynthetically active radiation; Pfr, farred absorbing form of phytochrome; Pr, red absorbing form of
phytochrome; R, red; UV-B P, ultraviolet-B absorbing
photoreceptor. The spectrum was scanned near noon on a clear
summer day at Corvallis, OR, USA.
42
100
T
80
SL
T
FR
4
0
1
z0 60
z
cc
40-
w
20-
00
10-1
100
101
104
103
102
106
106
SUNLIGHT (400-800 nm) FLUENCE (.1 mol m-2)
0
0.0001 0.001
0.01
0.1
1
10
100
FULL SUNLIGHT EQUIVALENT (seconds)
FIG. 2.2. Effects of sunlight pulses on the germination of
buried seeds of Datura ferox under field conditions. Seeds
were unearthed near midday using light-tight equipment,
exposed to sunlight and buried again at -7 cm. The open
symbol indicates germination of seeds irradiated with FR
after a saturating exposure to sunlight. After Scopel et al.
1991.
43
7-
1--
5
-
I
0.3
0.5
1.0
LEAF AREA INDEX
I
2
15
<
>
- 0<
1.5 2.0
E5
licz
a_
FIG. 2.3. Elongation rates of the first internode of Datura
ferox seedlings growing in even canopies of different
densities under natural radiation. Also shown is the effect
of density on PAR interception by individual seedlings.
After Ballare et al. 1990.
44
9
8
E7
E
I10 6
zw
J
5
12;
4-
0
z
i
cc
w
Z
3_
2
1
0
GREEN FENCE
II
III
0 BLEACHED FENCE
I
I
10 12 14 16 18 20 22 24
DAYS OF TREATMENT
FIG. 2.4. Effect of the proximity of a green grass canopy on
the elongation of the first internode of Sinapis alba
seedlings growing under full sunlight. Control seedlings
were grown in front of canopies bleached with an application
of paraquat. After Ballare et al. 1987b.
45
1.0
SECOND INTERNODE
MIDDAY
0.8
_
---.
LAI
it4 IfA. ,14
0.00
0/2
.1
I
1
1
I
1
0.6
eI
I
I
o
I
I
I
ig
0.4
0.2
FR
R
a_
0
600
700
800
WAVELENGTH (nm)
FIG. 2.5. Effects of neighboring seedlings on the spectral
distribution of the radiation scattered within the stems of
Datura ferox seedlings in an even canopy of LAI = 0.72.
AbbrevitifiTiiis: FR, far-red; R, red. After Ballare et al.
1989.
46
z
0
17-
uj
0
Z -E
I CC Id
-
Z a.
E
0
3
INTEGRATING
CYLINDER
FR
R
B
1
-i_f_21
wea.rliZ
es,'"
111"
are.
FIBER OPTIC
2_
PROBE
A
.4.AAt
4.
:1
tA A
as.
S.
.
0
0.05 0.1
0.5
1
2
5
LEAF AREA INDEX
FIG. 2.6. Effects of increasing density in even canopies of
dicotyledonous seedlings on light interception by leaves
(top) and the light environment of the stems. The
integrating cylinder collects side light received by the
stem surface; the fiber optic probe collects light scattered
within the stem tissue. Abbreviations: B, blue; FR, far-red;
R, red. After Ballare et al. 1991c.
47
6
E
w 4
cc
z
0
z0
_1
w
2
0
ISOLATED
WITH
NEIGHBORS
FIG. 2.7. Elongation responses of Datura ferox first
internodes when seedlings were placed in the center of an
even canopy of LAI -0.9 under natural radiation. During the
experiment the internodes were surrounded by annular
cuvettes containing distilled water (clear filter) or a
CuSO4 solution that absorbed FR radiation and maintained the
R:FR ratio at ca. 1.1 (FR-absorbing filter). Adapted from
Ballare et al. 1990.
48
R:FR
the
reduced
that
filters
main
of
field.
the
in
of
2
had
and
the
above
cm
1990.
et
After
Novoplansky
pot's
rim
al.
transmission.
only
directions.
compass
PAR
similar
extended
and
gray
filters
different
the
of
in
each
frequency
grown
relative
the
plants
Green
developing
stems
their
with
bars
of
The
length
oleracea
Portulaca
of
shoots
represents
seedlings
orientation
azimuthal
the
on
of
side
light
green
of
effect
The
2.8.
FIG. ratio
49
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64
CHAPTER 3
PHOTOMORPHOGENIC EFFECTS OF UV-B RADIATION ON HYPOCOTYL
ELONGATION IN WILD-TYPE AND STABLE-PHYTOCHROME-DEFICIENT
MUTANT SEEDLINGS OF CUCUMBER
Carlos L. Ballare*, Paul W. Barnes and Richard E. Kendrick
Depts of Forest Science and Crop and Soil Science, Oregon
State Univ., Peavy Hall 154, Corvallis, OR 97331-5705,
U.S.A. (C. L. B.); Dept of Biology, Southwest Texas State
Univ., San Marcos, TX 78666-4616, USA (P. W. B.); Dept of
Plant Physiological Research, Wageningen Agricultural Univ.,
Generaal Foulkesweg 72, NL-6703 BW Wageningen, The
Netherlands (R. E. K.).
Physiologia Plantarum 83:652-658 (1991).
65
ABSTRACT
Hypocotyl elongation responses to ultrviolet-B (UV-B)
radiation were investigated in glasshouse studies using de-
etiolated seedlings of a long-hypocotyl mutant (lh) of
cucumber (Cucumis sativus L.) deficient in stable
phytochrome, its near isogenic wild type (WT), and a
commercial cucumber hybrid (cv. Burpless). A single 6- or 8h exposure to UV-B applied against a background of white
light inhibited hypocotyl elongation rate by ca 50 % in lh
and WT seedlings. This effect was not accompanied by a
reduction in cotyledon area expansion or dry matter
accumulation. Plants recovered rapidly from inhibition and
it was possible to stimulate hypocotyl elongation in plants
exposed to UV-B by application of gibberellic acid. In all
genotypes inhibition of elongation was mainly a consequence
of UV-B perceived by the cotyledons; covering the apex and
hypocotyl with a filter that excluded UV-B failed to prevent
inhibition. These results indicate that reduced elongation
does not result from assimilate limitation or direct damage
to the apical meristem or elongating cells, and strongly
suggest that it is a true photomorphogenic response to UV-B.
The fact that UV-B fluences used were very low in relation
to total visible light, and the similarity in the responses
of lh and wild-type plants, are consistent with the
hypothesis that UV-B acts through a specific photoreceptor.
It is argued that, given the weak correlation between UV-B
and visible-light levels in most natural conditions, the UVB receptor may play an important sensory function providing
information to the plant that cannot be derived from light
signals perceived by phytochrome or blue/UV-A sensors.
Key words - Cucumber, Cucumis sativus, photomorphogenesis,
phytochrome mutants, hypocotyl elongation, UV-B radiation.
66
INTRODUCTION
Photomorphogenesis in higher plants involves the action
of at least three photoreceptor systems: phytochromes, a
variety of receptors for blue / ultraviolet-A (UV-A)
radiation, and an as yet little- studied UV-B receptor (e.g.
Mohr 1986). The existence of the latter photoreceptor has
been inferred from studies on light-mediated synthesis of
anthocyanins and flavonoids (Wellmann 1971, 1976, 1983,
Yatsuhashi et al. 1982, Mohr and Drumm-Herrel 1983, Beggs et
al. 1986). Besides influencing pigment synthesis, UV-B
radiation is known to induce a variety of changes in plant
morphology, including inhibition of elongation growth
(Steinmetz and Wellmann 1986, Tevini and Teramura 1989,
Barnes et al. 1990, and references therein). Barnes et al.
(1988) and Ryel et al. (1990) suggested that differential
morphological responses to UV-B may be important in
determining competitive relations between species in
communities subjected to increased levels of UV-B.
The mechanisms underlying the effects of UV-B on plant
morphology are poorly understood and, for a number of
reasons, the involvement of a specific receptor in the
responses is far from established. In many studies where UVB was applied against a background of white light, altered
plant morphology was accompanied by a reduction in
photosynthesis and total biomass production. Thus, it is
difficult to determine whether reduced elongation of stem
and leaves is due to specific photomorphogenic effects of
UV-B or is simply a consequence of a general reduction in
growth capacity resulting from photosynthetic inhibition
(Tevini and Teramura 1989). UV-B inhibition of elongation
may also result from direct cellular damage at the level of
meristems and elongating organs. In fact, the action
spectrum obtained by Steinmetz and Wellmann (1986) for the
inhibition of hypocotyl elongation in etiolated seedlings of
67
Lepidium sativum resembles those related to DNA damage in
other systems (see Caldwell 1971). Even when damaging
processes are ruled out as a cause of inhibition, the
possible involvement of other photoreceptors must be
considered. Thus, the interpretation of experiments with
monochromatic UV-B sources is complicated by the fact that
the protein moiety of phytochrome also absorbs in the UV-B
region, and part of the energy can be transferred to the
chromophore and can be effective in photoconversion (Pratt
and Butler 1970). There have been suggestions that some
morphological responses to UV-B might be a consequence of
direct absorption of UV-B by growth regulators (e.g. auxins,
see Curry et al. 1956, Tevini and Teramura 1989).
Recent field and glasshouse experiments with grasses
have indicated subtle effects of UV-B on shoot (leaf)
elongation (10 to 15 % reduction in total height) that were
not accompanied by similar changes in total dry matter
(Barnes et al. 1988, 1990) or photosynthesis (Beyschlag et
al. 1988). It is very unlikely that this effect of UV-B was
mediated by phytochrome, because the fluence rates of UV-B
used were very small in relation to the total phytochromeabsorbable radiation that plants received under natural
conditions. However, phytochrome might still have played a
role, either by influencing the synthesis of UV-B screening
pigments, or as a result of interactions with the putative
UV-B receptor system. Several types of interactions between
UV-B and light treatments that affect phytochrome are well
documented in the case of anthocyanin and flavonoid
synthesis in various plant systems (e.g. Wellmann 1971, Mohr
and Drumm-Herrel 1983, Beggs and Wellmann 1985, Arakawa
1988) .
We have investigated the roles played by UV-B damage
and phytochrome in the inhibition by UV-B of hypocotyl
68
elongation by means of physiological experiments with deetiolated seedlings of the lh mutant of cucumber and its
near isogenic wild type (WT). The lh mutant has been
proposed to be deficient in the stable type phytochrome on
the basis of spectrophotometric (Adamse et al. 1988, Peters
et al. 1991), immunochemical (Lopez et al. 1991, E. LopezJuez and A. Nagatani, personal communication), and
physiological (Adamse et al. 1987, 1988, Lopez-Juez et al.
1990, Ballare et al. 1991) evidence.
Abbreviations - lh, long-hypocotyl mutant; UV-A,
Ultraviolet-A (320-400 nm); UV-B, Ultraviolet-B (280-320
nm); WT, wild-type.
MATERIALS AND METHODS
Seed sources and growth conditions
Seeds of the long-hypocotyl (lh) mutant and near
isogenic wild type (WT) of Cucumis sativus L. (Koornneef and
van der Knaap 1983, Adamse et al. 1987) were multiplied in
the departments of Genetics (Wageningen Agricultural Univ.,
The Netherlands) and Ecology (Facultad de Agronomia, Univ.
de Buenos Aires, Argentina). Seeds of the commercial hybrid
cv. Burpless used in some experiments were obtained from
Fredonia Seeds Co., Fredonia, NY, USA.
All the experiments were carried out during the autumn
and winter months under glasshouse conditions at Corvallis,
OR, USA. Seeds were germinated on moist filter paper under
continuous white light provided by fluorescent tubes at
25°C. Two days after germination (3.5 days after sowing)
seedlings were transferred to the glasshouse, planted in 10cm pots filled with standard 1:1 (v/v) perlite (Supreme
Perlite Co., Portland, OR, USA) / promix (Promix Premier
Brands Inc., New Rochelle, NY, USA) mixture, and grown for
69
an additional 4 days under natural radiation supplemented
with light from sodium vapor lamps (see below). Air
temperature fluctuated daily between 18 and 26°C
(experiments in Figs 3.2 to 3.4), or between 17 and 33°C.
(experiments in Fig. 3.5); photoperiod was 14 h.
Solutions of gibberellic acid (GA3)
(Sigma) were
applied to seedlings by gently painting the apex, both sides
of the cotyledons and the hypocotyl with a brush. The
concentration of GA3 used (1 mM) was at least 10 times
greater than the concentration required to saturate
elongation responses under our conditions.
Light sources and filters
During de-etiolation in the glasshouse and during the
experimental period plants were grown under natural
radiation supplemented with light from high pressure sodium
vapor lamps (Lucalox LU1000, General Electric Co.,
Cleveland, OH, USA). Maximum instantaneous
photosynthetically active radiation (PAR) at plant level,
measured with a cosine-corrected sensor (Quantum type,
Q4888, Li-Cor, Lincoln, NE, USA), typically varied between
150 and 480 umol m2 s4 depending on sky conditions. PAR
levels > 1,200 umol m2 s4 were recorded only occasionally.
UV-B was supplied by panels of 8 fluorescent tubes (UV-B
313, Q-Panel, Cleveland, OH, USA) enclosed by either 0.13
mm -thick clear cellulose acetate film (UV-B treatment) or
0.13-mm-thick clear polyester (optically equivalent to
Mylar, Hillcor Plastics, Santa Fe Springs, CA, USA; control
treatment) as described by Barnes et al. (1990). The UV-B
lamps were turned on and off 3 h (4 h in some experiments)
before and after solar noon, respectively. Filters were preaged for 6 h before the experiments and replaced after 18-24
h of use.
70
Spectral irradiance measurements were taken with a
double-monochromator spectroradiometer (Optotronic Model
742, Orlando, FL, USA). Details about instrument calibration
and wavelength-accuracy tests are given by Barnes et al.
(1990). Representative scans of the light sources are shown
in Fig. 3.1. The estimated effective UV-B fluence at plant
level in the UV-B treatment was 9.6 kJ m2 d4 (6 h) or 12.7
kJ m2 d4 (8-h exposures). These figures were calculated by
using the generalized plant action spectrum of Caldwell
(1971) normalized to unity at 300 nm, and are given to
facilitate comparison with other studies. Effective UV-B
fluences calculated in this way are comparable to those that
naturally exist in tropical locations.
In studies on the site of perception, different plant
organs were covered with UV-B-excluding clear polyester
during irradiations. Two plastic 'leaves' were placed over
the adaxial surface of the cotyledons and were held in place
by means of small wire arms. The filters did not project
shadows beyond the edge of the cotyledons and left the apex
of the seedling uncovered. The apex and hypocotyl were
covered by means of an inverted folder made of clear
polyester in which two small windows were open to exclude
the cotyledons.
Growth measurements and statistics
Hypocotyl length (defined as the distance between the
connection with the seed and the base of the apex) was
measured to the nearest 0.5 mm with a ruler. Leaf area was
measured with a LI-3100 area meter (Li-Cor, Lincoln, NE,
USA). Dry weights (shoot + root) were obtained after tissue
was dried at 70°C for 48 h. Nondestructive estimations of
initial cotyledon areas and dry weights were obtained by
relating these attributes to measurements of cotyledon
71
length and width. The regressions models were fitted using
WT and lh seedlings of the same age and grown under the same
conditions as the experimental plants. Values of the
coefficients of determination ranged from 0.85 to 0.99.
Individual plants were assigned at random to each of the
light treatments and the type of filter (clear polyester or
cellulose acetate) used in each of the light banks was rerandomized at every filter change. Each experiment was
repeated at least twice with similar results; data from
different experimental days were pooled for analysis. Mean
comparisons were made with Student's t-tests using SAS
algorithms (SAS Institute Inc. 1987); in factorial
experiments (e.g. Fig. 3.5) only probabilities associated
with pre-planned comparisons are reported. Appropriate
transformations were used to obtain variance homogeneity
whenever necessary.
RESULTS
Six day-old, fully de-etiolated WT and lh seedlings
were exposed to UV-B during the central 6 h of the
photoperiod. The increment in hypocotyl length was evaluated
24 h after the beginning of the UV-B treatment. Hypocotyl
elongation was much more rapid in the stable-phytochromedeficient lh mutant than in WT seedlings. UV-B strongly
inhibited hypocotyl elongation and the relative effect was
similar in both genotypes (inhibition was 53 and 47 % in lh
and WT seedlings, respectively, Fig. 3.2A).
Photosynthetic activity is frequently lowered by UV-B
treatments, particularly in plants receiving low PAR levels
(Warner and Caldwell 1983, Mirecki and Teramura 1984), and
cucumber has been reported to be very sensitive in this
regard (e.g. Takeuchi et al. 1989, Tevini and Teramura
1989). Therefore, the observed inhibition of elongation
might be simply another consequence of a drastic drop in
72
photosynthetic assimilation. Plants in our experiments were
growing very rapidly (relative growth rates 25-30 % dry
weight per day). However, under our conditions, we did not
find any effect of 6-h exposures to UV-B on cotyledon area
expansion or dry matter accumulation (Fig. 3.2B,C). It is
important to note here that differences between means on the
order of 50 % (that is an effect comparable to that found in
the case of hypocotyl elongation, Fig. 3.2A) would have been
rated as highly significant in these experiments, in which
standard errors of means were ca 12 % in the case of dry
weight increments.
In order to study the effects of UV-B over time we
conducted another series of experiments. Plants were given
an 8-h exposure to UV-B on the sixth day after germination
(7.5 days after sowing) and elongation rates during and
after the UV-B treatment were estimated from length
measurements. These experiments showed that: 1) the
immediate effect of UV-B on elongation (i.e. inhibition
during the UV-B treatment) was very similar in WT and lh
seedlings, and 2) the recovery from UV-B inhibition was
rapid. This was particularly true in the case of WT
seedlings, where the difference between control and UV-B
treated plants disappeared overnight. The lh mutant
recovered the day after UV-B exposure (Fig. 3.3).
In another group of experiments with a similar
irradiation schedule, plants were coated with saturating
GA3
(1 mM) at the beginning of the UV-B treatment. Control
plants received distilled water only. Applications of GA3
increased hypocotyl elongation by two-fold in both control
and UV-B treated plants (Fig. 3.4). Thus, the relative
inhibition was not affected by GA3 treatments and, in the
case of the WT seedlings, recovery after UV-B was delayed by
73
GA3 (data not shown).
In order to determine the sites of UV-B action we
selectively covered the adaxial surface of the cotyledons or
the apex and the hypocotyl with clear polyester film. The
filter excluded UV-B with minimal effects on UV-A and PAR
levels received by the tissue. These experiments were
carried out with WT and lh seedlings and with the commercial
hybrid cv. Burpless. In all cases the inhibitory effect of
UV-B on hypocotyl elongation was abolished by covering the
cotyledons (Fig. 3.5). Inhibition was not prevented by
covering the apex and hypocotyl in WT and cv. Burpless
seedlings, but was reduced to some extent in seedlings of
the lh mutant. When plants were not exposed to UV-B (i.e.
lamps enclosed by clear polyester film), elongation rates of
covered and uncovered plants were not significantly
different (0.18 < P < 0.84).
DISCUSSION
UV-B radiation strongly inhibited hypocotyl elongation
in the three genotypes of cucumber investigated (Figs 2-5).
Several lines of evidence suggest that this effect of UV-B
is not a consequence of nonspecific metabolic damage.
It is clear that the strong inhibitory effect of UV-B
on hypocotyl elongation is not accompanied by similar
effects on dry matter accumulation or cotyledon area
development (Fig. 3.2), even when PAR levels were relatively
low. Although we have observed some growth inhibition by UVB in longer term experiments (i.e., 1 week or more of
treatment), that effect seems to lag well behind the rapid
elongation response. Our results with cucumber hypocotyls
parallel previous observations on leaf elongation responses
to UV-B. In Rumex patientia leaves, UV-B radiation appears
74
and Caldwell (1978)
to reduce cell division rate (Dickson
that is not
and causes a rapid inhibition of elongation
(Sisson and
correlated with photosynthetic responses
(1988, 1990)
Caldwell 1977). Similarly, Barnes et al.
elongation in
reported that UV-B reduces sheath and blade
affecting total
field- and glasshouse-grown grasses without
shoot biomass.
elongation
Our experiments also showed that hypocotyl
increased
rates of UV-B treated plants can be substantially
weight to
by GA3 applications (Fig. 3.4). This adds extra
is not merely a
the conclusion that inhibition of elongation
to UV-B
consequence of assimilate limitation. Some responses
with
in other systems have been found to be correlated
cited in Tevini and
changes in GA3 content (Rau et al. 1988,
Teramura 1989). It is interesting to note that even at
saturating doses, GA3 failed to negate the inhibitory effect
inhibition is not
of UV-B (Fig. 3.4), suggesting that the
mediated by effects on GA3 synthesis or activity.
Time course experiments (Fig. 3.3) showed that recovery
that, if any
from UV-B inhibition is rapid, suggesting
without having
cellular damage exists, it is easily repaired
long-lasting consequences on elongation growth. The
differences between WT and lh seedlings in the kinetics of
recovery might explain observations showing a slightly
higher inhibition in the lh mutant in long-term experiments
(e.g. field
where plants were not exposed to continuous UV-B
experiments by Ballare et al. 1991).
the entire
In most previous studies UV-B was applied to
experiments
shoot; a significant observation in the present
extension is exerted
was that UV-B inhibition of hypocotyl
mainly through the cotyledons (Fig. 3.5). Therefore, direct
75
action on the dividing or elongating cells cannot explain
the UV-B effect.
Our experiments have eliminated the most common
explanations for the inhibition of elongation by UV-B that
include some component of damage (i.e. reduced
photosynthetic assimilation and damage to meristems and
elongating regions). Although it would be difficult to
reject all of them, it should be noted that our data are
totally consistent with a model in which hypocotyl
elongation is controlled by UV-B acting through a receptor
located in the cotyledons. This is analogous to the
inhibition of hypocotyl elongation by red light in cucumber
(Black and Shuttleworth 1974), a response that appears to be
mediated by the stable type of phytochrome (Adamse et al.
1987). However, it is clear from the relatively low fluences
of UV-B used in our experiments (Fig. 3.1), and the
similarity in the responses of WT and lh seedlings, that UVB is not acting through phytochrome. A specific UV-B
receptor is therefore implicated.
Phytochrome and the UV-B receptor(s) are known to
interact in several plant systems. These interactions are
well documented in the case of the photoregulation of
flavonoid and anthocyanin synthesis. Thus, in etiolated corn
seedlings, red-light pretreatments, probably acting through
phytochrome, enhance the response to UV-B (Beggs and
Wellmann 1985). In apple fruits (Arakawa 1988) and in
etiolated wheat seedlings (Mohr and Drumm-Herrel 1983) the
effect of UV-B on anthocyanin production is greatly enhanced
by subsequent red-light treatment. In most cases it would
seem that Pfr is necessary for the expression of the
response to UV-B, although the precise mode of coaction
between the two photoreceptor systems is still unclear
(Beggs et al. 1986). In our experiments the responses to UV-
76
B of the phytochrome-deficient lh mutant were very similar
to those of the near isogenic WT. We therefore conclude that
the effect of UV-B on hypocotyl elongation in de-etiolated
cucumbers is independent of the presence of stable
phytochrome. Steinmetz and Wellmann (1986) reported that the
inhibitory effect of monochromatic-UV-B plus red light
pulses on hypocotyl elongation in etiolated Lepidium sativum
was not affected by subsequent modifications in phytochrome
status.
Morphological effects of UV-B have most commonly been
interpreted as by-products of nonspecific damage caused by
UV-B radiation. As pointed out by Wellmann (1983) some of
these effects may represent active responses that confer
some adaptive advantage or protection to the plant (e.g.
reduced exposure to UV-B). As a result of very effective
Rayleigh (molecular) scattering through the atmosphere, a
large proportion (usually >50 %) of global UV-B is received
at the Earth's surface as diffuse radiation (Caldwell 1971,
Grant 1991). Strong scattering, and absorption in the ozone
layer, contribute to determine that the incoming irradiance
and spectral photon distribution are much more dramatically
affected by solar elevation in the UV-B region than in the
visible part of the electromagnetic spectrum (Caldwell
1971). An important implication of this, which is often
overlooked, is that it might be impossible for the plant to
retrieve information about how much UV-B is being received
by a particular organ at a given point in time from light
signals sensed by other photoreceptors (i.e. phytochrome and
cryptochrome). Thus, if morphological responses to UV-B are
adaptive to plants, they may require a specific receptor to
sense the UV-B environment.
77
Acknowledgements. We are most grateful to Dr. Maarten
Koornneef and members of the Genetics Dept (Wageningen
Agricultural Univ.) for provision of original seed batches
and to Marjorie Storm and Dave Olszyk for their help in
setting up the experiments in the facilities of the U. S.
Environmental Protection Agency (Corvallis). C.L.B. was
supported by the Consejo Nacional de Investigaciones
Cientificas y Tecnicas (Republica Argentina) and thanks Dr
Steve Radosevich for his hospitality at OSU.
78
80
mW m-2 n
'1
UV-B
Control
40
296
306
316
11111111111111111111111111111111111111111
300
350
400
450
500
550
600
650
700
750
800
Wavelength (nm)
FIG. 3.1. Spectral photon distributions measured at plant
level under control (polyester-filtered) and UV-B
(cellulose-acetate filtered) lamp systems. Measurements were
taken under completely overcast sky. Spectral irradiances
for the UV region (inset) are given in energy units to
facilitate comparison with previous works.
79
A
PoNM:01
1.6
Control
ED uv -a
0.6
1.1,7110007
0
B
- control
C
1ZO
in, -a
Pr4".0.2406
PoT-0.9437
0
10
C
Pr,T.O.WM
- contra
CM uv-e
P1,T.0.6352
6
4
2
0
/h-mutant
Wild-Type
FIG. 3.2. Effect of 6-h exposures to UV-B on hypocotyl
elongation, cotyledon area expansion and dry matter
accumulation in de-etiolated lh and WT cucumber seedlings.
Initial values of leaf area and dry weight were estimated
from measurements of leaf width and length. Final
measurements were taken 24 h after the beginning of the UV-B
treatment. Significance levels are indicated for each
difference; n=16.
80
A: Wad-Type
Coatiel
ENS uv-s en Day I
0.2
PoT-0.7995
Pr)T-0.4996
PoT43.0027
day 1
night 1
day 2
Time Period
FIG. 3.3. Recovery from UV-B inhibition of hypocotyl
elongation in de-etiolated lh and WT cucumber seedlings. Day
1 refers to the 8-h period during which plants were exposed
to UV-B in the middle of a 14-h photoperiod, day 2 is the
equivalent period on the next day, night 1 is the
intervening 16-h (10 h night) period. Significance levels
are indicated for each difference; n=12. Note difference
between panels in y-axis scale.
81
0.4
A:Wild -Type
I= control
CM .uv-e
Por-00001
0.3
0.2
0.1
0
Water
GA3
FIG. 3.4. Effects of GA3 on hypocotyl elongation growth in
de-etiolated lh and WT cucumber seedlings exposed to UV-B.
Eight-hour exposures were used; length increments were
measured at the end of the UV-B treatment. Significance
levels are for the effect of GA3; n=14. Note difference
between panels in y-axis scale.
82
0.5
0.2
0.1
0
2
1
0
0.3
0.2
0.1
0
none
cotyledons
hypocotylapex
Organ Covered
FIG. 3.5. Effects of covering different parts of the
seedling with a UV-B-excluding polyester filter on hypocotyl
elongation in de-etiolated cucumbers exposed to UV-B for 6
h. Length increments were measured 24 h after the beginning
of the UV-B treatment. Significance levels are indicated for
each difference; n=14-20. Note differences among panels in
y-axis scale.
83
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Kendrick, and M. Koornneef M. 1988. Photophysiology and
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Adamse, P., P. A. P. M. Jaspers, R. E. Kendrick, and M.
Koornneef. 1987. Photomorphogenic responses of a long
hypocotyl mutant of Cucumis sativus L. Journal of Plant
Physiology 127:481-491.
Arakawa, 0. 1988. Photoregulation of anthocyanin synthesis
in apple fruit under UV-B and red light. Plant and Cell
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Ballare, C.L., J. J. Casal, and R. E. Kendrick. 1991.
Responses of light-grown wild-type and long-hypocotyl
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shadelight. Photochemistry and Photobiology 54:819-826.
Barnes, P.W., P. W. Jordan, W. G. Gold, S. D. Flint, and M.
M. Caldwell. 1988. Competition, morphology and canopy
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Barnes, P. W., S. D. Flint, and M. M. Caldwell. 1990.
Morphological responses of crop and weed species of
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Beggs, C.J., and E. Wellmann. 1985. Analysis of lightcontrolled anthocyanin formation in coleoptiles of Zea
mays L.: The role of UV-B, blue and far-red light.
Photochemistry and Photobiology 41:481-486.
Beggs, C. J., E. Wellmann, and H. Grisebach. 1986.
Photocontrol of flavonoid biosynthesis. Pages 467-499
in R. E. Kendrick and G. H. M. Kronenberg, editors.
Photomorphogenesis in plants. Martinus Nijhoff,
Dordrecht. The Netherlands.
Beyschlag, W., P. W. Barnes, S. D. Flint, and M. M.
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Caldwell. 1988. Enhanced UV-B irradiation has no effect
on photosynthetic characteristics of wheat (Triticum
aestivum L.) and wild oat (Avena fatua L.) under
greenhouse and field conditions. Photosynthetica
22:516-525.
Black, M., and J. E. Shuttleworth. 1974. The role of the
cotyledons in the photocontrol of hypocotyl extension
in Cucumis sativus L. Planta 117:57-66.
Caldwell, M. M. 1971. Solar UV radiation and the growth and
development of higher plants. Pages 131-177 in A. C.
Giese, editor. Photophysiology, Vol. 6. Academic Press,
New York, NY, USA.
Curry, G. M., K. V. Thimann, and P. M. Ray. 1956. The base
curvature response of Avena seedlings to the
ultraviolet. Physiologia Plantarum 9:429-440.
Dickson, J. G., and M. M. Caldwell. 1978. Leaf development
of Rumex Datientia L. (Polygonaceae) exposed to UV
irradiation (280-320 nm). American Journal of Botany
65:857-863.
Grant, R. H. 1991. Ultraviolet and photosynthetically active
bands: plane surface irradiances at corn canopy base.
Agronomy Journal 83:391-396.
Koornneef, M., and B. J. van der Knaap. 1983. Another long
hypocotyl mutant at the lh locus. Cucurbita Genetics
Cooperative Reports 6:13-18.
Lopez, E., A. Nagatani, R. E. Kendrick, M. Koornneef, J.
Wesselius, and M. Furuya. 1991. The lh mutant of
cucumber lacks a type II phytochrome protein and shows
an extreme shade-avoidance reaction. Page 7 in Book of
Abstracts, Proceedings of the Annual Meeting and 31d
Symposium of the Japanese Society of Plant Physiology.
Okayama. Japan.
LOpez-Juez, E., W. F. Buurmeijer, G. H. Heeringa, R. E.
Kendrick, and J. C. Wesselius. 1990. Response of light-
85
grown wild-type and long-hypocotyl mutant cucumber
plants to end-of-day far-red light. Photochemistry and
Photobiology 52:143-150.
Mirecki, R. M., and A. H. Teramura. 1984. Effects of
Ultraviolet-B irradiance on soybean V. The dependence
of plant sensitivity on the photosynthetic photon flux
density during and after leaf expansion. Plant
Physiology 74:475-480.
Mohr, H. 1986. Coaction between pigment systems. Pages 547564 in R. E. Kendrick and G. H. M. Kronenberg,
editors. Photomorphogenesis in plants. Martinus
Nijhoff, Dordrecht, The Netherlands.
Mohr, H., and H. Drumm-Herrel. 1983. Coaction between
phytochrome and blue/UV light in anthocyanin synthesis
in seedlings. Physiologia Plantarum 58:408-414.
Peters, J. L., R. E. Kendrick, and H. Mohr. 1991.
Phytochrome content and hypocotyl growth of longhypocotyl mutant and wild type cucumber seedlings
during de-etiolation. Journal of Plant Physiology
137:291-296.
Pratt, L., and W. Butler. 1970. Phytochrome conversion by
ultraviolet light. Photochemistry and Photobiology
11:503-509.
Ryel, R. J., P. W. Barnes, W. Beyschlag, M. M. Caldwell, and
S. D. Flint. 1990. Plant competition for light analyzed
with a multispecies canopy model I. Model development
and influence of enhanced UV-B conditions on
photosynthesis in mixed wheat and wild oat canopies.
Oecologia (Berlin) 82:304-310.
SAS Institute Inc. 1987. SAS/STAT Guide for Personal
Computers. SAS Institute Inc., Cary, NC, USA.
Sisson, W. B., and M. M. Caldwell. 1976. Photosynthesis,
dark respiration, and growth of Rumex patientia L.
exposed to ultraviolet irradiance (288 to 315
nanometers) simulating a reduced atmospheric ozone
86
column. Plant Physiology 58:563-568.
Steinmetz, V., and E. Wellmann. 1986. The role of solar UV-B
in growth regulation of cress (Lepidium sativum L.)
seedlings. Photochemistry and Photobiology 43:189-193.
Takeuchi, Y., M. Akizuki, H. Shimizu, N. Kondo, and K.
Shugahara. 1989. Effect of UV-B (290-320 nm)
irradiation on growth and metabolism of cucumber
cotyledons. Physiologia Plantarum 76:425-430.
Tevini, M., and A. H. Teramura. 1989. UV-B effects on
terrestrial plants. Photochemistry and Photobiology
50:479-487.
Warner, C.W., and M. M. Caldwell. 1983. Influence of photon
flux density in the 400-700 nm waveband on inhibition
of photosynthesis by UV-B (280-320 nm) irradiation in
soybean leaves: separation of indirect and immediate
effects. Photochemistry and Photobiology 38:341-346.
Wellmann, E. 1971. Phytochrome-mediated flavone glycoside
synthesis in cell suspension cultures of Petroselinum
hortense after preirradiation with ultraviolet light.
Planta 101:283-286.
Wellmann, E. 1976. Specific ultraviolet effects in plant
photomorphogenesis. Photochemistry and Photobiology
24:659-660.
Wellmann, E. 1983. UV radiation in photomorphogenesis. Pages
745-756 in W. Shropshire Jr. and H. Mohr, editors.
Encyclopedia of Plant Physiology, New Series, Vol. 16B.
Springer Verlag, New York, NY, USA.
Yatsuhashi, H., T. Hashimoto, and S. Shimizu. 1982.
Ultraviolet action spectrum for anthocyanin formation
in broom sorghum first internodes. Plant Physiology
70:735-741.
87
CHAPTER 4
PHYTOCHROME-MEDIATED PHOTOTROPISM IN DE-ETIOLATED SEEDLINGS:
OCCURRENCE AND ECOLOGICAL SIGNIFICANCE
Carlos L. Ballare., Ana L. Scopel, Steven R. Radosevich, and
Richard E. Kendrick
Depts of Forest Science and Crop and Soil Science, Oregon
State Univ., Peavy Hall 154, Corvallis, OR 97331-5705, USA
(C. L. B., A. L. S., S. R. R), Dept de Ecologia, Facultad de
Agronomia, Univ. of Buenos Aires, (1417) Buenos Aires,
Argentina (C. L. B., A. L. S.) and Dept of Plant
Physiological Research, Wageningen Agricultural Univ.,
Generaal Foulkesweg 72, NL-6703, The Netherlands (R. E. K.)
Plant Physiology, in press (1992).
88
ABSTRACT
Phototropic responses to broadband far-red (FR)
radiation were investigated in fully de-etiolated seedlings
of a long-hypocotyl mutant (lh) of cucumber (Cucumis sativus
L.), which is deficient in phytochrome-B, and its nearisogenic wild type (WT). Continuous unilateral FR light
provided against a background of white light induced
negative curvatures (i.e. bending away from the FR light
source) in hypocotyls of WT seedlings. This response was
fluence-rate dependent and was absent in the lh mutant, even
at very high fluence rates of FR. The phototropic effect of
FR light on WT seedlings was triggered in the hypocotyls and
occurred over a range of fluence rates in which FR was very
effective in promoting hypocotyl elongation. FR light had no
effect on elongation of lh-mutant hypocotyls. Seedlings
grown in the field showed negative phototropic responses to
the proximity of neighboring plants that absorbed blue (B)
and red light and back-reflected FR radiation. The bending
response was significantly larger in WT than in lh
seedlings. Responses of WT and lh seedlings to lateral B
light were very similar; however, elimination of the lateral
B light gradients created by the proximity of plant
neighbors abolished the negative curvature only in the case
of lh seedlings. More than 40% of the total hypocotyl
curvature induced in WT seedlings by the presence of
neighboring plants was present after equilibrating the
fluence rates of B light received by opposite sides of the
hypocotyl. These results suggest that: (i) phytochrome
functions as a phototropic sensor in de-etiolated plants,
and (ii) in patchy canopy environments, young seedlings
actively project new leaves into light gaps via stem bending
responses elicited by the B-absorbing photoreceptor(s) and
phytochrome.
89
INTRODUCTION
Phototropism occurs when a lateral light stimulus
induces a difference in extension rate between the two sides
of an elongating organ. The difference in extension rate is
a response to the internal light gradient created by light
scattering and absorption between the "illuminated" and
"shaded" flanks of the phototropic organ (Reviews by Firn
[1986], Briggs and Baskin [1988], and lino [1990]). Action
spectra for phototropism in a variety of plant systems show
peaks of quantum effectiveness between 320 and 500 nm
(References in Dennison [1979] and lino [1990]), indicating
that a B/UV-A photoreceptor plays a key role in the
detection of internal light gradients. The roles of
phytochrome in phototropism are not completely clear.
Studies with etiolated oats (Briggs 1963, Dennison 1979) and
etiolated dicotyledonous seedlings (Shropshire and Mohr
1970) suggested that phytochrome does not function as a
detector of radial light gradients, and it has been proposed
that its main function in phototropism is to modulate the
response to B light mediated by another receptor (Woitzik
and Mohr 1988). However, recent work with dark-adapted maize
(lino et al. 1984, Kunzelmann and Schafer 1985) and totally
etiolated pea seedlings (Parker et al. 1989) indicted that
the radial Pfr gradient formed upon exposing the shoots to
short pulses of lateral light can explain observed curvature
responses. The picture is more complex in the case of deetiolated seedlings, because detailed information on
spectral sensitivity is scant. lino (1990) suggested that
the R:FR gradient that exists across green organs exposed to
unilateral polychromatic light (Seyfried and Fukshansky
1983, Vogelmann 1986) should generate a radial gradient of
Pfr/P. This radial Pfr/P gradient might, in turn, lead to a
gradient of growth inhibition and eventually to phototropic
bending. Rough action spectra obtained by Atkins (1936)
using broadband light sources for de-etiolated seedlings of
90
two dicotyledonous species do, in fact, show a small peak in
the R region. Shuttleworth and Black (1977) obtained
positive curvatures in green cucumber seedlings using
broadband R light; the response was not observed in
sunflower seedlings, where only one fluence rate was tested.
Winning (1938) reported a phototropic effect of R light in
sunflower seedlings de-etiolated under reduced sunlight; the
same treatment was ineffective in the case of Sinapis alba.
Recent studies with cucumber indicated that the bending of
hypocotyls toward direct sunlight under glasshouse
conditions is mainly a response to the B component;
experimental reversal of the natural B light gradient
between opposite sides of the seedlings led to reversal of
the direction of curvature, and the effect of this treatment
was the same in seedlings of wild type (WT) and the
phytochrome-B-deficient (lh) mutant (Ballare et al. 1991b).
In parallel with the uncertainty on the role of
phytochrome in the detection of radial light gradients,
there is little information on the influence of the spectral
distribution of natural radiation on phototropic movements
of light-grown plants (Morgan and Smith 1981). Remarkably
few studies on phototropism have been conducted in natural
light environments, and the gaps in our understanding of the
ecological aspects of phototropic responses have been
discussed recently by Firn (1988) and Iino (1990). In this
study we set out to test the role of phytochrome in the
detection of radial light gradients in de-etiolated
seedlings exposed to continuous unilateral illumination. We
also tested phototropic responses in plant canopies and
evaluated the possibility that changes in the spectral
composition of scattered radiation, brought about by nearby
plants, may elicit phototropic responses in young seedlings.
Our approach is based on physiological experiments with
seedlings of the lh mutant of cucumber and its near isogenic
91
WT. The lh mutant lacks a phytochrome-B polypeptide (LOpezJuez et al. 1992), shows reduced levels of
spectrophotometrically-detectable phytochrome after deetiolation (Adamse et al. 1988, Peters et al. 1991), and
lacks (or exhibits severely reduced) elongation responses to
end-of-day (Adamse et al. 1988, Lopez-Juez et al. 1990) and
day-time (Ballare et al 1991b) R:FR treatments.
Abbreviations: B, blue; UV-A, ultraviolet-A (320-400 nm);
FR, far-red; R, red; P, total phytochrome; WT, wild type;
lh, long-hypocotyl mutant; WL, white light.
MATERIALS AND METHODS
Plant material and growth conditions
Seeds of the long-hypocotyl (lh) mutant and the near
isogenic wild-type (WT) of Cucumis sativus L. (Adamse et al.
1987) were obtained from the Department of Genetics
(Wageningen Agricultural Univ., The Netherlands). For the
field experiments, cucumber seeds were germinated under
white light (WL) as described previously (Banat6 et al.
1991a). Two-day-old seedlings were transferred to a
glasshouse, planted in 10-cm pots containing a standard
soil-less mixture and grown for an additional 4 d under
natural radiation supplemented with light from high-pressure
sodium vapor lamps (Banat-6 et al. 1991a). The pots were
then transferred to the field and distributed among light
treatments. Maize seeds (Zea mays L. cv Early Jubilee) were
sown directly in the field in recently cultivated soil on 15
May, 1991. Six plots (1.0 x 0.5 m; east-west orientated)
were planted at a rate of ca. 4000 seeds m2 in order to
produce dense, homogeneous canopies. The experiments were
carried out between early July and early September, 1991;
during this period maize plants were periodically clipped to
maintain canopy height at 50 + 5 cm. All field experiments
92
were carried out at the Vegetable Research Farm, Department
of Horticulture, Oregon State University, near Corvallis,
OR. For the controlled-environment experiments, cucumber
seeds were sown in 2.5-cm pots at a depth of 0.5 cm. Pots
were incubated in the same growth room where the phototropic
experiments were carried out. The cotyledons began to emerge
from the soil 7 d after sowing; after two more days of
growth, seedlings were selected for uniformity and allocated
to the different light treatments. The WL (Fig. 4.1) in the
growth room was provided by a mixture of 110-W F96T12/CW/HO
fluorescent tubes (Sylvania) and 300-W incandescent lamps
(Photoperiod = 12 h d4; PPFD [plant level] = 160 gmol
s4;
day/night temperature = 22/16°C).
Light treatments and experimental procedures
were
edge
plot
soil
In the field experiments, potted cucumber seedlings
placed ca. 8 cm to the south of the directly sunlit
of dense maize canopies or at the center of a 10 x 10 m
where all weedy vegetation was removed periodically by
cultivation. In the initial experiments only WT
seedlings were used. The pots were fitted inside 10 cm
(i.d.) x 30-cm high clear-polyester open cylinders in order
to prevent movement of the seedlings by wind. In experiments
designed to test the role of phytochrome in natural
phototropism, pots were fitted inside 10 cm (i.d.) x 30-cm
(WT seedlings) or 40-cm high (lh seedlings) acetate
cylinders that were prepared as follows. The hemi-cylinder
that faced south (i.e. the one that received direct
sunlight) was made of a sheet of Roscolux #21 (Golden Amber)
acetate (Rosco Labs., Port Chester, NY), that absorbed most
of the B/UV-A radiation with minimal effects on R-FR
wavelengths, or a sheet of Roscolux #34 (Flesh Pink)
acetate, that reduced PPFD to a similar extent due to strong
absorption in the green region (Transmission of the pink
93
film in the B and R-FR regions was 50 % and >80 %,
respectively). This southern wall of the cylinder filtered
all direct sunlight received by the seedlings. The hemicylinder that faced north (i.e. the one that received
diffuse skylight or light scattered by the nearby canopy)
was made of Roscolux #34. Seedlings were placed in front of
the maize canopies or at the center of a clear plot as
described above.
In the controlled-environment experiments, continuous
unilateral FR light (Fig. 4.1) was provided from the side
against a WL background (from above) by a bank of
incandescent lamps. The light beam was filtered trough 3 cm
of water and one sheet of each of the following Roscolux
filters: #83 (Medium Blue), #12 (Straw), and #42 (Deep
Salmon). The resulting beam was a rectangle 60 cm wide and
ca. 13 cm high which struck the entire shoot of the
seedlings. Different fluence rates were obtained by
inserting neutral density filters constructed of 1.1-mm mesh
plastic netting between the light sources and the seedlings.
A mock source covered with black plastic was used as a
control. In preliminary experiments we found that seedlings
tended to bend away from the simulated and FR light sources
even if the lights were off. This was remedied by placing,
on the opposite side of the seedlings, a 30-cm high Babsorbing curtain (Roscolux #12) at a distance of 23 cm. In
the experiments where FR light was applied bi-laterally, the
seedlings were positioned midway between the light source
and a mirror; FR fluence rates were calculated by doubling
the measured output of the lateral light source. In the
experiments to determine the site of perception, carried out
with WT seedlings, a small, "U"-shaped aluminum piece (1 x
0.3 [lateral wings) x 2.5 [height] cm), was used to prevent
lateral FR from reaching the hypocotyl. The shield was
positioned at a distance of 5 mm from the hypocotyl.
94
Applications of GA3 (100 gM) were carried out as described
previously (Ballare et al. 1991a).
Phototropic responses were determined by measuring the
angle between the vertical vector and the upper 1.5-cm
section of the hypocotyl on the plane parallel to the
lateral light beam (controlled-environment experiments) or
on the north-south plane (field experiments). The angle of
curvature was measured 48 h after the beginning of
treatments in the field experiments and at different
intervals (see figures and figure legends) in the controlled
environment. Hypocotyl length (defined as the distance
between the rim of the pot and the base of the apex) was
measured to the nearest 0.5 mm with a ruler. All light
measurements were obtained by using a cosine-corrected
receiver and a calibrated LI-COR 1800 spectroradiometer (LiCor, Lincoln, NE).
Statistics
Individual plants (=pots) were assigned at random to
each of the light treatments (i.e. light source or canopy x
filter combination). Each experiment was repeated at least
three times with similar results; data from different
experimental days were pooled for analysis. Results are
presented as means ± SE, except where indicated otherwise.
RESULTS
Cucumber seedlings placed on the southern (sunlit) side
of a green, dense maize crop bent toward the south (i.e.
away from the neighboring crop) within 2 d of the beginning
of the treatment. The hypocotyls of their isolated
counterparts were almost vertical (Fig. 4.2). This neighborinduced negative tropism could have been mediated by a Babsorbing photoreceptor or by phytochrome, since the
95
proximity of green plants can change the horizontal
component of radiation absorbed by these two photoreceptors
due to absorption and reflection of sunlight (Ballare et al.
1987, 1991c, Kasperbauer et al. 1984, Smith et al. 1990).
Induction of hypocotyl curvature by lateral B light
gradients is a well documented phenomenon in de-etiolated
dicotyledonous seedlings. A series of controlled-environment
experiments were carried out to test whether a phytochromemediated phototropism can be demonstrated in de-etiolated
cucumbers. Seedlings de-etiolated for 2 d were given
additional FR light from one side against a background of WL
provided from above. In WT seedlings, continuous unilateral
FR induced a negative phototropic bending that was
statistically detectable after 2 h of treatment; curvature
was about 90 % of the maximal after 4 h. No curvature was
detected in the lh mutant (Fig. 4.3). In WT seedlings,
increasing the fluence rate of unilateral FR light between
17 and 122 gmol m2 s4 had a log-linear effect on hypocotyl
curvature (Fig. 4.4). When WT seedlings were irradiated bilaterally, variation of FR fluence rate over such a range
led to a ca. 2 fold increase of hypocotyl elongation rate
(Fig. 4.5). In seedlings of the lh mutant, additional FR
light failed to promote elongation; however, applications of
GA3 were clearly effective in FR-treated and control
seedlings (Fig. 4.5). This effect of GA3 corroborates
results obtained under a variety of other lighting
conditions (Adamse 1988, Ballare et al 1991a,b) showing that
the potential for longitudinal growth promotion exists in lh
seedlings.
In our experiments the seedlings were positioned in
front of the FR light source with the long axis of their
cotyledons perpendicular to the lateral light beam. This
96
positioning was done in order to minimize differences in
light environment between cotyledons, which might have
indirectly induced hypocotyl curvature (the so called
"simulated phototropism" [Shuttleworth and Black 1977]).
Under these conditions, interposing an opaque barrier
between the FR source and the hypocotyl, without affecting
the light environment of the cotyledons, drastically reduced
the negative phototropic response (Fig. 4.6).
Spectral scans of the light received by cucumber
seedlings in the field showed that plants grown close to the
sunlit edge of a maize canopy received less B and R light
from the northern side and much more FR than isolated
seedlings (Table 4.1). In order to assess the relative
contribution of each of these changes to the bending
response showed in Fig. 4.2, we: (i) compared the curvature
responses to green neighbors of WT and lh seedlings, and
(ii) tested the effect of eliminating the steep south-tonorth B-light gradient created by the canopy by placing a Babsorbing filter on the southern side of the seedlings. To
compensate for the reduction in PPFD caused by this
treatment, control seedlings were shaded by a greenabsorbing acetate (see Materials and Methods). The effects
of the acetate filters and plant neighbors on the light
received by the plant from the northern and southern sides
are shown in Fig. 4.7. Seedlings grown without neighbors
(i.e. isolated seedlings) experienced a natural south-tonorth gradient of B light (Fig. 4.7A), and their hypocotyls
were slightly curved toward the south (Fig. 4.8, A and B).
The B-absorbing filter placed on the southern side reversed
the direction of the natural B-light gradient (Fig. 4.7B)
and caused isolated seedlings to bend toward the north (Fig.
4.8, A and B). The response to the B barrier was of the same
magnitude in isolated WT and lh seedlings. The presence of a
green canopy on the northern side exaggerated the south-to-
97
north B-light gradient and, in addition, created a steep
R:FR gradient due to absorption of R light and backreflection of FR radiation (Fig. 4.7C; cf. Fig. 4.7A and
Table 4.1). Plants grown in front of a green canopy were
markedly curved toward the south (i.e. away from the canopy)
(Fig. 4.8, C and D; see also Fig. 4.2). The bending response
to the proximity of the canopy was significantly larger in
WT than in lh seedlings (Fig. 4.8, C and D). Eliminating the
B-light gradient induced by the canopy by means of a Babsorbing filter (Fig. 4.7D) abolished the negative tropic
response in lh seedlings (Fig. 4.8C). In contrast, a
residual curvature of ca. 11 degrees (i.e. about 46 % of the
maximum response) remained in WT plants (Fig. 4.8D)
DISCUSSION
Our results strongly suggest that phytochrome functions
as a detector of light gradients established across the
hypocotyls of green cucumber seedlings and mediates
phototropic responses to long-wavelength (> 600 nm)
radiation. Unilateral FR radiation administrated against a
background of WL provided from above induced a negative
phototropic response in WT seedlings (Fig. 4.3). This
response was: (i) fluence rate dependent (Fig. 4.4), (ii)
not an indirect consequence of differential illumination of
the cotyledons (Fig. 4.6; cf. Shuttleworth and Black 1977),
and (iii) not observed in the phytochrome-B-deficient lh
mutant (Fig. 4.3), even at very high fluence rates of FR
light (Fig. 4.4). Comparison of these results with previous
work is difficult because other authors used different
species or experimental protocols. Atkins (1936) obtained
rough action spectra for the induction of phototropic
responses to continuous irradiation using broadband light
sources. The spectra for de-etiolated dicotyledonous
seedlings (Lepidium sativum and Celosia cristata) showed a
small (positive) peak in the R region in addition to the
98
peak in the B/UV-A. No data points were reported between ca.
710 and 850 nm (L. sativum) or 650 and 850 nm (C. cristata).
No such R-peak was observed for etiolated oat coleoptiles,
where the ineffectiveness of radiation above 500 nm has been
documented extensively (e.g. Briggs 1963, Dennison 1979).
Shropshire and Mohr (1970) stated that continuous R or FR
light did not induce hypocotyl curvature in etiolated
seedlings of two dicotyledonous species: Fagopyrum
esculentum and Sinapis alba. Continuous R light was also
ineffective in de-etiolated S. alba, but elicited curvature
in de-etiolated sunflowers (BUnning 1938). Shuttleworth and
Black (1977) found that continuous unilateral R light
induced positive curvature in hypocotyls of de-etiolated
cucumbers. However, the authors were suspicious of the
spectral purity of the R light source (i.e. B light
contamination) and did not discuss that result further. More
recently, Parker et al. (1989) interpreted the phototropic
response of totally etiolated pea epicotyls to short B light
pulses on the basis of phytochrome action. Based on previous
results with dark-adapted maize mesocotyls (lino et al.
1984, Kunzelmann and Schafer 1985), they proposed that
epicotyl curvature in their experimental conditions was
induced by a Pfr gradient established across the epicotyls
after illumination with unilateral B light. A related
mechanism may account for the negative phototropism induced
by continuous FR light in our experiments. Although fluence
rate gradients for 660 and 730 nm have been measured in
green hypocotyls of other species (Vogelmann 1986),
extrapolation to the present experiments is difficult
because our seedlings received omnidirectional WL
simultaneously with the unilateral FR treatment. The
important point is that, in our experiments, curvature was
induced over a range of FR fluence rates (Fig. 4.4) where a
2-fold change of fluence rate (i.e. equivalent to the
fluence-rate differential that could be expected between
99
opposite flanks of a 2-mm diameter, green hypocotyl
[Vogelmann 1986]) should produce a sizable (ca. 0.1 mm 114)
difference in elongation rate (Fig. 4.5). Simple geometrical
relations indicate that such a difference in elongation
between the "illuminated" and "shaded" flanks would yield a
bending rate of about 2.9 degrees h4, which is roughly
similar to the bending rates measured during the initial 2 h
in our experiments (Fig. 4.3).
The question at this point is whether a phytochromemediated phototropic mechanism plays a role in the curvature
responses of young seedlings to the proximity of green
neighbors (Fig. 4.2). The observation that hypocotyl-bending
responses were larger in WT than in lh seedlings (Fig. 4.8,
C and D, controls) is consistent with such a possibility,
but certainly can not be taken as definite evidence. In
fact, it is well known from a number of other studies that
an important action of phytochrome in phototropism is to
alter the pattern of response to unilateral B light mediated
by a specific photoreceptor (e.g. Woitzik and Mohr 1988).
Therefore, the observed differences between WT and lh
seedlings might simply reflect an effect of phytochrome-B on
the curvature response to the steep south-to-north B-light
gradient (Table 4.1 and Fig. 4.7C) created by the presence
of a green canopy. However, when that B-light gradient was
experimentally eliminated by placing a B-absorbing filter on
the southern side of the plants (Fig. 4.7D), a significant
southward curvature was still observed in the hypocotyls of
WT plants (Fig. 4.8D). Moreover, when the B-absorbing
barrier was placed on the southern side of isolated plants,
the northward curvature responses of WT and lh seedlings
were similar (Fig. 4.8, A and B). These and previous data
(Ballare et al. 1991b) suggest that the lack of phytochromeB does not affect the (steady state) curvature induced by
100
lateral B light in de-etiolated cucumbers under natural
light fluences. If we therefore assume that the actions of
phytochrome and the B-absorbing photoreceptor are roughly
additive in this system, we can calculate that about 46% of
the total bending induced by the proximity of a green canopy
is due to radial R:FR gradients perceived by phytochrome.
The observation that the hypocotyl is the main site of
perception of lateral FR (Fig. 4.6) is consistent with
results obtained in other dicotyledonous seedlings
irradiated with B light or WL (Franssen and Bruinsma 1981,
Hart and MacDonald 1981, Schwartz and Koller 1980,
Shuttleworth and Black 1977) and with the hypothesis that it
is the R:FR gradient established across the hypocotyl that
elicits the bending response. This observation is very
important with respect to the ecological significance of
stem phototropism. In open plant stands, the presence of
neighboring individuals can alter the light environment of
vertically-oriented stems well before there is any mutual
shading at leaf level (Ballare et al. 1987, 1990, 1991c, and
Fig. 4.7, this paper). These alterations influence the rate
of internode elongation and may convey critical information
to plants that are competing with neighbors for colonizing
the aerial environment (Ballare et al. 1990). According to
our data (Fig. 4.7), the radial light gradients perceived by
the stems are related to the spatial distribution of
neighbors around the plant. Tropic responses to these
gradients would improve the efficiency with which a seedling
collects light in a patchy light environment, as the
probability of new leaves being actively projected into
light gaps would increase. It has been reported recently
that seedlings of Portulaca oleracea, a plant with a
plagiotropic shoot system, tend to avoid growing toward
plant neighbors or plastic objects that back-scatter
radiation with a low R:FR ratio. Part of the response in
101
this case appeared to be because main stems were unlikely to
become recumbent toward natural or simulated neighbors
(Novoplansky et al. 1990). Our results indicate that, in
light-grown cucumbers, phototropic mechanisms controlled by
the B-absorbing photoreceptor(s) and by phytochrome are
responsible for the negative bending responses elicited by
the proximity of neighboring plants.
Acknowledgements. We are most grateful to Dr. Maarten
Koornneef and members of the Department of Genetics
(Wageningen Agricultural Univ.) for provision of the
cucumber seeds. We also wish to thank Randy Hopson (Veg.
Res. Farm, OSU) and Lloyd Graham (Forest Res. Lab., OSU) for
practical assistance, and Gretchen Bracher for preparing the
figures. C.L.B. and A.L.S. were supported by the Consejo
Nacional de Investigaciones Cientificas y Tecnicas
(Argentina) and the Dept. of Forest Science (OSU).
102
TABLE 4.1. Effects of the proximity of green plants on the
light flux measured parallel to the ground.
Measurement
Photon Irradiance
(Amol m4 s4)
B
Receiver pointed
toward the south
137.5
Receiver pointed
toward the north:
(a) no neighbors
R
R: FRt
FR
223.7
238.3
1.00
36.3
42.8
72.6
0.63
8.0
21.6
128.1
0.16
(b) in front of a
green canopy
*Measurements were taken near midday under clear sky on
28 July 1991. The surface of the light receiver was
normal to the ground and received direct solar
radiation when pointed toward the south. The receiver
was positioned 8 cm to the south of the maize canopy to
obtain the last row of data. B=400-500 nm, R=600-700
nm, FR=700-800 nm.
t(654- 664/726 -736) quantum flux ratio.
103
1.0
11
cm_
WL
FR
/
ee
III
2
....-
I
0
0
C
I
I
I
CIS
I
"1:3
I
2 0.5c0
I
I
t0
t0
I
I
I
I
I
ELI
I
I
I
0.
u)
r
400
I
I
500
600
el
1
700
800
Wavelength (nm)
FIG. 4.1. Spectral photon distributions of the light sources
used in the controlled-environment experiments. The quantum
integral between 300 and 400 nm in the FR beam was < 0.01 %
of the output between 700 and 800 nm.
104
Isolated
Canopy
FIG. 4.2. Effect of the proximity of a green canopy on the
orientation of the hypocotyl of WT seedlings at solar noon.
Seedlings were grown for 2 d at the center of a clear plot
(isolated) or 8 cm to the southern (sunlit) side of a dense
maize crop (canopy). Data are means of 4 independent
experiments (n=12-16).
105
4
20
-4,3'
a)
e
-2-
a)
-4-
92
-6-
48
8_
-0
=
WT
O Ih
WT + FR
lh + FR
m
0 -10-
i'-- ... ...
-12-
-14-16
0
4
5
6
7
II
Time (h)
FIG. 4.3. Time courses of hypocotyl bending in WT and lh
seedlings growing under WL with or without exposure to
unilateral FR light (86.5 gmol m2 s-1). Time = 0 indicates
the onset of FR irradiation; negative values on the y-axis
indicate bending away from the FR source; II indicates the
end of the second day of treatment (i.e. two 8-h exposures
to unilateral FR light). Data are means of 4 independent
experiments (n=15-28); for clarity of presentation, SE (all
<1 degree) were omitted in the upper curves.
106
4
2-
-
0
1;
4-,
-2-
-8 -4= -6-
o wr
-10-12-
0 lh
WT + FR
Ih+ FR
-14-16
0
2
3
4
5
In FR fluence rate (Imo! m-2 s-1)
FIG. 4.4. Effects of increasing the fluence rate of
unilateral FR light on hypocotyl curvature in WT and lh
seedlings. Curvatures were measured after 5 h of continuous
FR light; negative values on the y-axis indicate bending
away from the FR source. Data are means of 4 independent
experiments (n=68-80, controls, or 8-29, intermediate
fluences, or 29-37, highest fluence).
107
1.4
+ GA3
'T
1.27e--'
1.0-
Control
E
E
1
0.8
a)
WI'
Iris
oc 0.6ga)
O lh
WT + FR
lh + FR
0.4'
El
0.2-,
0..-1,,---r--.T--r---6
2
3
4
In FR fluence rate (gmol
5
m-2 s-1)
FIG. 4.5. Effects of increasing the fluence rate of bilateral FR light on hypocotyl elongation in WT and lh
seedlings (the latter with or without application of GA3).
Elongation rates were estimated from length measurements
taken at the beginning of the experiment and after 5 h of
continuous FR irradiation. The solid line parallel to the
abscissa denotes the range of FR fluence rates used in the
curvature experiments. Data are means of 3 independent
experiments (n=13-28).
108
Control
FR
Treatment
FIG. 4.6. Effect of shading the hypocotyl during irradiation
with unilateral FR light (121.7 Amol m-2 sl) on hypocotyl
curvature in WT seedlings. Curvatures were measured after 5
h of continuous FR irradiation; negative values on the yaxis indicate bending away from the FR source. Data are
means of 9 independent experiments (n=28-37).
109
.... / ....
I11
E
E. --, -.---.. .....-.--.I 1 II
i ....i if %.,
I %.1
-es
-?.--
cas
0
400
,
500
600
700
II
0
800400
500
600
700
800
3
Cu
e 3
.r.
C canopy (control)
D canopy (B barrier)
0
15
.c
ca. 2-
e
t
a)
0.
(1)
1
0
400
,_- .... ...... .
I
,
I
500
600
700
0 '--:----si" a i
800 400
500
I
I
600
700
800
Wavelength (nm)
FIG. 4.7. Effects of the proximity of a green maize canopy
and B-absorbing acetate filters on the spectral photon
distribution of the light flux measured parallel to the
ground. Measurements were taken near midday under clear sky
on 28 July 1991. The surface of the light sensor was normal
to the ground and pointed toward the southern (solid line)
or northern (dashed line) hemisphere. When pointed toward
the south, the sensor received either direct sunlight
filtered by a Roscolux #34 acetate (control) or direct
sunlight filtered by a B-absorbing acetate #23 (B-barrier).
When pointed toward the north, the sensor was always
shielded by the acetate #34 and received either diffuse
skylight (isolated) or light back-scattered by an 8-cm
distant maize canopy (canopy).
110
12
_c
0
A /h/isolated
B WT/isolated
8
4
0
=0
4
8
12
T/canopy
Control
B barrier
Control
B barrier
FIG. 4.8. Effects of the proximity of a green maize canopy
and B-absorbing acetate filters on the orientation of the
hypocotyl of WT and lh seedlings at solar noon. Seedlings
were grown for 2 d at the center of a clear plot (isolated)
or 8 cm away from the southern edge of a dense maize crop
(canopy). Data are means of 11 independent experiments
(n=19-26, isolated, or 42-56, canopy).
111
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115
CHAPTER 5
FORAGING FOR LIGHT IN HORIZONTALLY-PATCHY ENVIRONMENTS BY
WILD-TYPE AND PHYTOCHROME-B-DEFICIENT CUCUMBER PLANTS
116
ABSTRACT
In this paper we address the following questions: (i)
Do plants that colonize horizontally-patchy environments
actively project leaf area into light gaps and avoid poorly
illuminated sites? (ii) What are the roles of the
photoreceptor phytochrome in gap colonization? Our approach
was based on dynamic studies of gap colonization by cucumber
(Cucumis sativus L.) plants and comparisons of morphological
development between wild-type (WT) and lh-mutants. The lh
mutant of cucumber is deficient in phytochrome B and, when
de-etiolated, seedlings show reduced or no response to
alterations of red:far-red (R:FR) ratio, one of the
environmental factors that plants use to monitor the
proximity of other plants. WT plants grown at the edges of a
maize canopy in the field projected outside the canopy
nearly all the nodes produced over a period of four weeks.
In contrast, about 50 % of the nodes and more than 80 % of
the apical buds of lh plants were positioned within the
maize crop. Controlled-environment experiments showed that
seedlings of both genotypes are able to detect horizontal
light / shade mosaics and actively project leaves toward
light gaps via phototropic (stem-bending) responses. These
responses are presumably mediated by a specific blue-lightabsorbing photoreceptor. A mechanism independent of
phytochrome B also appeared to control the angle of display
of individual leaves with respect to the direction of
illumination. Our controlled-environment experiments showed
that, when compared over periods of several weeks, WT plants
are far more efficient than lh mutants at invading and
maintaining their shoots in light gaps. The reasons for this
difference were multiple and appeared to result from several
actions of phytochrome B on WT plants that received full
light with high R:FR ratio. These actions included: (i)
Influencing the gravitropic response of stems, so that WT
shoots grown under full light were diagravitropic, whereas
117
lh shoots were orthogravitropic. The diagravitropic habit,
coupled with an efficient phototropic system, appears to
help WT shoots to escape from the shaded sectors and invade
light gaps. (ii) Delaying tendril production and reducing
stem-bending responses to mechanical stresses exerted by
tendrils. These two actions combine with the ones listed
above to determine that the direction of shoot growth in WT
plants is mainly dictated by the spatial distribution of
light gaps, and not by the distribution of potential points
of support for tendril attachment. (iii) Switching plants
from the "shade-avoiding" (default) developmental program to
the one that results in a phenotype with short internodes
and reduced apical dominance. These two actions would favor
concentration of nodes and leaf area in light gaps.
118
INTRODUCTION
The light environment in plant canopies is
heterogeneous in space and time. Plants are considered to
"forage" for light if they tend to adjust the spatial
distribution of their light-absorbing surfaces to track the
spatial distribution of the light resource (Grime 1979,
Slade and Hutchings 1987, Hutchings and Mogie 1990).
Although the ultimate manifestation of foraging for light in
space is concentration of plant modules or biomass in highlight patches, such a pattern of asymmetric growth may
originate for various reasons, i.e.: (i) simple differences
in plant growth rate between low- and high-light patches,
(ii) differences in growth rate preceded by specific
morphological responses to the local light climate, and
(iii) active projection of modules or leaf area into light
gaps and concomitant avoidance of poorly illuminated sites.
The first case is probably trivial. If the plant has equal
access to patches of different quality, one would expect to
find most of the biomass concentrated under high-light
conditions after a sufficiently long period of growth. The
only necessary condition for such asymmetry to occur is that
individual modules of the plant have some degree of growth
independence. The second case can be exemplified with the
observations of Pefialosa (1983) on the morphological
development of the vine (Ipomoea phillomega) in tropical
rain-forests of Mexico. This liana "travels" through the
forest floor by means of creeping stolons with long
internodes and rudimentary leaves. The transformation of
stolons into twinning shoots with short internodes and large
leaves appears to be favored by the changes of light
environment that the stolons experience when they reach
sunny patches. These morphological changes can be
interpreted as a form of foraging for light, because they
increase the light-harvesting capacity of the shoot when it
reaches a gap in the forest canopy (for a more complete
119
discussion see Penalosa 1983, Hutchings and Slade 1988, and
Hutchings and Mogie 1990). The third case (selective
projection of leaf area toward light gaps) is a clear
example of true foraging for light, and probably the most
efficient mode of space occupation in patchy light
environments. These considerations suggest that static
descriptions of plant architecture can be interpreted in
several ways. Therefore, they are not sufficient to decide
whether or not a selective pattern of space invasion is the
principal mechanism that causes a non-random distribution of
plant biomass between high- and low-light patches.
For selective invasion to occur, plants need sensors
capable of monitoring environmental changes associated with
variations in light conditions. A considerable body of
evidence derived from experiments in controlled-environments
(reviewed by Smith 1986) and conditions of natural radiation
(reviewed by Ballare et al. 1992b) indicates that projection
of leaves into the upper strata of the canopy is a process
modulated by at least two specific light sensors:
phytochrome and a blue- (B-) absorbing photoreceptor. In
plant canopies formed by plants of similar height (e.g. crop
stands), the light signals that trigger changes in stem
elongation rate via these photoreceptors may be perceived by
an individual plant well before its leaves become shaded by
neighbors (Ballare et al. 1987; Ballare et al. 1991c).
Therefore, the acceleration of stem elongation elicited by
high population densities in even-aged monocultures (Ballare
et al. 1988; Ballare et al. 1990) can be regarded as a true
process of foraging for light in the vertical dimension.
Light signals perceived by phytochrome also have been
implicated in mechanisms that influence the direction of
growth on the horizontal plane in patchy canopy
environments. Novoplansky et al.
(1990) observed that the
120
shoots of Portulaca oleracea, a rapidly-growing recumbent
herb, showed a consistent tendency to avoid developing
toward neighboring individuals when colonizing patchy
canopies. They also found that main stems of E., oleracea
seedlings did not normally become recumbent toward plastic
objects which light absorption spectrum was similar to that
of green leaves.
Recently, it has been demonstrated that the proximity
of green neighbors elicits a rapid phototropic response
(stem-bending) in young seedlings of cucumber grown in the
field. This response is mediated by a B-absorbing
photoreceptor and phytochrome (Ballare et al., 1992a). The
phototropic response might be the initial step of a
mechanism leading to neighbor avoidance and selective
invasion of high-light patches in heterogeneous canopies.
The experiments reported in this paper were designed to gain
mechanistic insight into the process of foraging for light
in horizontally-patchy canopies and to test the involvement
of the photoreceptor phytochrome B (phyB) (See Smith and
Whitelam 1990, and Quail 1991, for a description of the
phytochrome family). To this end, we compared the
morphological development of wild-type and phyB-deficient
mutant seedlings of cucumber in experimental mosaics of
light and shade and studied the influence of phytochrome on
morphological processes that defined the dynamics of gap
colonization.
MATERIALS AND METHODS
Plant material
Seeds of the long-hypocotyl (lh) mutant and the near
isogenic wild-type (WT) of Cucumis sativus L. (Adamse et al.
1987, Kendrick and Nagatani 1991) were obtained from the
Department of Genetics (Wageningen Agricultural University,
The Netherlands). The lh mutant lacks a PHYB polypeptide
121
(Lopez Juez et al. 1992) and shows reduced levels of
spectrophotometrically-detectable phytochrome after deetiolation (Adamse et al. 1988; Peters et al. 1991). phyBdeficient lh plants grown under white light of high red (R)
far-red (FR) ratio have a phenotype that resembles that of
WT plants grown under deep vegetational shade, where the low
R:FR ratios maintain low levels of the active form of
phytochrome (Pfr) at photoequilibrium. Thus, compared with
the WT, lh plants have much longer hypocotyls and
:
internodes, increased apical dominance and tendril formation
(e.g. Lopez Juez et al. 1990), and higher shoot:root ratios
(Casal J. J. and Ballare C. L., unpublished data). Mutant lh
seedlings lack (or exhibit severely reduced) elongation
responses to end-of-day (Adamse
al. 1990) and day-time (Ballare
treatments. The phyB deficiency
elongation responses to natural
et al., 1988, Lopez Juez et
et al. 1991b, 1992a) R:FR
severely reduces the
shadelight and appears to
eliminate elongation responses to B light mediated by a Babsorbing photoreceptor (Ballare et al. 1991b). Light-grown
mutant seedlings retain phototropic (hypocotyl-bending)
responses to B light, but lack phytochrome-mediated tropic
responses to FR radiation (Ballare et al. 1992a).
Experimental procedures
(1) Field experiment. The experiment was carried out in the
fields of the Vegetable Research Farm, Department of
Horticulture, Oregon State University, near Corvallis, OR.
Maize seeds (Zea mays L. cv Early Jubilee) were sown in rows
at a density of 100 plants m2 in a 3 x 9 m plot laid out in
recently cultivated soil on 15 May, 1991. A 2-m wide zone
was delimited around the crop and kept free of weeds. Maize
leaves that extended beyond the perimeter of the plot were
clipped periodically to produce a distinct canopy / openground border. Weeds that emerged within the limits of the
122
maize crop were not controlled. The most common species were
Amaranthus retroflexus and Solanum nigrus.
Cucumber seeds were germinated in a growth chamber
under white light provided by fluorescent tubes. Two-day old
seedlings were transferred to a glasshouse, planted in 10-cm
pots containing a standard potting mixture and grown for an
additional 4 days under natural radiation supplemented with
light from sodium vapor lamps (see Ballare et al. 1991a).
The pots were then transferred to the field, where the
seedlings were grown for another 3 days before being
transplanted into holes dug at the edges of the maize crop.
Differences in initial height between WT and lh seedlings
were minimized by adjusting planting depth. Seedlings were
well watered and fertilized with a standard 12:12:12 NPK
mixture. By the time of transplanting (July 1991), the maize
crop was 50-cm tall, and reduced midday photosyntheticallyactive radiation (PAR) and R:FR ratio at ground level
(measured with a LiCor 1800 spectroradiometer, Lincoln, NE)
by 85 % and 59 %, respectively. One month after
transplanting, the number of new nodes produced by WT and lh
seedlings, their location in relation to the maize canopy,
and the height of the main-stem apex were recorded.
(2) Controlled-environment experiments. These experiments
were carried out in a 2.5 x 2.5 x 2.5-m walk-in growth room
with controlled environment. Light was provided by a mixture
of 110-W F96T12/CW/HO fluorescent tubes (Sylvania) and 300-W
incandescent lamps (Photoperiod = 14 h day''; photosynthetic
photon flux density at plant level = 160 gmol m'a s'; R:FR
ratio = 2.24; day/night temperature = 22/16°C). Cucumber
seeds were sown in 1-L pots at a depth of ca. 0.5 cm. The
cotyledons began to emerge from soil 7 days after sowing;
after seven more days, four metal rods (1-m long) were
123
driven into each pot in order to provide potential points of
attachment for the cucumber tendrils. The morphological
development of seedlings was followed over periods of
different duration in conditions of uniform illumination or
in artificial light / shade-support mosaics. In this latter
case, areas of reduced irradiance were created in the growth
room by means of opaque screens that were suspended 80 cm
above the pot rim level. Stray light in the shaded areas was
reduced by covering the walls of the chamber with black
plastic sheets. A large number of rods were placed under the
artificial canopies (shaded sectors) in order to simulate
the presence of stems or culms that might serve as sources
of support for climbing plants like cucumber. The pots
containing the seedlings were placed at the light / shadesupport border. Variations of this general setup were used
in individual experiments as explained below. In each case,
cucumber plants (=pots) were assigned at random to the
different treatments and were considered as replicates for
statistical analysis. Results are given as means ±SE except
when otherwise is indicated.
RESULTS
Morphology of WT and lh seedlings under uniform illumination
Wild type and lh-mutant seedlings of cucumber were
grown in a controlled environment and their morphological
development was followed over a period of one month focusing
on attributes that, a priori, were considered to be of
consequence in processes of space colonization. The
generation of main-stem nodes occurred at a similar rate in
both genotypes (Fig. 5.1A) but, as observed in other studies
(e.g. L6pez-Juez et al. 1990), internode elongation was much
faster in lh plants. This was reflected in large differences
between genotypes in total plant height (Fig. 5.1B). In WT
seedlings the first axillary branches were produced at the
second main-stem node, about three weeks after emergence
124
(Fig. 5.1C). The onset of branching was comparatively
retarded in lh seedlings and, more remarkably, all the
laterals produced remained undeveloped during the period of
observations (Fig. 5.1D). Undeveloped lateral stems
consisted of a short internode (length < 0.5 mm), a leaf
primordium, and a very small apex. Lateral branches produced
by WT seedlings elongated rapidly (Fig. 5.1D) and were
almost horizontal (inclination with respect to vertical =
79.2 ± 3.5 degrees; n = 12; 39 days).
Mutant lh seedlings produced the first tendrils on the
third main-stem node (Fig. 5.1E) and, by the third week of
observations each plant had, on the average, 2 tendrils
coiled around the vertical supports established at the
perimeter of the pots (Fig. 5.1F). In comparison, WT plants
started to produce tendrils later and at higher node numbers
(Fig. 5.1E), which resulted in delayed attachment to the
supporting rods (Fig. 5.1F).
None of the main-stems became recumbent during the
period comprised by this set of observations, but we noticed
that WT shoots tended to become horizontal as plants entered
the reproductive phase (data not shown). Differences between
WT and lh-mutant in the gravitropic response of their stems
were therefore investigated using mature plants. The 12-th
main-stem internode was fixed onto a horizontal plane and
the inclination of successive internodes measured 48 h
later. The substrate was covered with aluminum foil to
create a nearly-isotropic light environment in the vicinity
of the shoot tips. No potential sources of support for
tendrils were provided in order to avoid possible effects of
tendril attachment on stem orientation (see below). Shoots
of mature WT plants maintained low deflection angles; in
contrast, lh-mutant internodes rapidly tended to resume a
vertical stance (Fig. 5.2). A similar diagravitropic re-
125
orientation of WT shoots was observed 96 h after placing
mature internodes in a vertical, upside-down position (not
shown). In summary, these observations showed that the lack
of phyB results in increased stem elongation, reduced branch
development, stimulated tendril production, and altered stem
gravitropism in mature plants.
Morphological development of WT and lh seedlings in a patchy
canopy environment
Wild type and lh seedlings were grown in the field for
3 days and transplanted to the edges of a maize crop. After
four weeks of growth, the number and spatial location of all
new nodes generated by the seedlings were recorded. WT
plants produced 8.9 (± 0.8) nodes compared with 6.9 (± 0.4)
generated by lh plants. Eighty percent of the total
population of nodes produced by WT plants were placed
outside the maize canopy; in contrast, about one half of the
nodes produced by lh plants were found within the crop (Fig.
5.3A). All WT plants had approximately horizontal stems
after one month, and their apical buds were placed close to
the ground surface (Fig. 5.3B). The direction of horizontal
spreading was clearly non-random, since more than 80 % of WT
plants had their apical bud outside the canopy (Fig. 5.3C).
In contrast, only 5 % of the lh seedlings developed a
diagravitropic stance. The main-stem apices of lh plants
were at an average height of 24 cm and, most significantly,
nearly all of them were positioned within the maize crop
(Fig. 5.3C). This experiment revealed that the lack of a
single photoreceptor (phyB) has a striking impact on the way
in which cucumber seedlings position their growth points
relative to the distribution of the light resource in a
patchy canopy.
126
Morphological development of WT and lh seedlings in
artificial canopies
Artificial canopies were created in a growth room using
neutral screens supported at a height of 80 cm by means of
numerous vertical rods. This experimental setup mimicked the
situation in dense natural canopies, where shading by
overhanging leaves is normally associated with the presence
of large numbers of (vertically-oriented) stems, culms or
petioles that provide potential points of attachment for
tendrils of climbing plants. WT and lh seedlings grown in
individual pots were placed at the edges of these artificial
canopies and the changes in plant morphology and shoot
orientation were followed over time.
As they emerged from the soil, seedlings of both
genotypes bent along the hypocotyls and projected their
cotyledons toward the full-light sector (Table 5.1).
Seedlings of the lh mutant initially invaded the artificial
gaps more rapidly than the WTs (Fig. 5.4B). In fact,
although the angles of hypocotyl curvature were very similar
in both genotypes under our lighting conditions, hypocotyl
elongation was much faster in lh than in WT seedlings
(Table
5.1).
Wild type and lh seedlings produced main-stem nodes at
a similar rate (Fig. 5.4A). By the third week, most lh
seedlings had produced a tendril on the second node (Fig.
5.4C). When tendrils were first visible, they appeared as
tightly-packed coils at the axil of the youngest unfolding
leaf. As tendrils uncoiled, their tips were projected within
a few hours ca. 7 cm ahead of the apical bud. Tendrils at
that stage elongated very rapidly (2.4 + 0.3 mm h4; n = 10)
and nutated actively (not shown). The maximal length of
functional tendrils recorded under our conditions was about
127
25 cm. At that point tendrils usually developed several free
coils or buckles at the tip and ceased to elongate. By the
fourth week, two thirds of the lh plants had at least one
tendril coiled around the supporting rods or around stems of
neighboring
produced by
tendrils of
there was a
plants (Fig. 5.4C). No tendrils had been
WT seedlings. After the attachment of the first
lh plants to the outer row of vertical rods,
very rapid, backward movement of the terminal
buds from the full-light to the shade-support sectors (Fig.
5.4B). In turn, as the apical meristems moved closer to the
full-light / shade-support border, new tendrils contacted
and coiled around more internal supports. At that point the
direction of growth appeared to be primarily dictated by the
spatial arrangement of potential supporting points. More
than one half of the apical buds were in the shaded areas
after 6 weeks of growth. WT plants projected all new growth
into the full-light areas throughout the experiment (Fig.
5.4B). Their internodes were short (< 2cm) and fewer than 10
% of the plants produced tendrils after 6 weeks (Fig. 5.4C).
Measurements of leaf orientation, taken on the fifth
week, showed that, in both genotypes, the laminas were
inclined toward the full-light area (Fig. 5.5). The angle of
leaf display showed remarkably little variability among
replicate leaves, and was completely independent of the
spatial orientation of the main stem. That constancy
appeared to be a consequence of adjustment in the angle
between the stem and the petiole, not between the petiole
and laminae (not shown). Main stems of WT plants were
consistently inclined toward the artificial gaps. There was
much more variation in stem angle among lh plants, where the
average angle was negative, indicating preferential
inclination toward the shade-support sector after five weeks
(Fig. 5.5). Thus, the lack of phyB had marked consequences
128
on the distribution of nodes through the light / shadesupport mosaic, but did not affect orientation of individual
leaves.
The effects of tendrils on gap colonization
We noticed that, after several weeks of growth, WT
plants of the above experiment started to produce tendrils
at the axil of every new leaf and some of the tendrils
eventually became attached to the supports (not shown). Two
additional experiments were carried out to investigate how
tendril production influences stem motility and foraging for
light in a patchy environment.
In the first experiment we tested whether or not the
ability to project stems into light gaps is limited by the
activity of thigmotropic tendrils. WT and lh plants were
grown to the 11-th node stage in the controlled environment.
The ninth internode of six WT and six lh plants was affixed
with its long axis oriented vertically in an artificial
light gap, 2 cm from the full-light / shade border. A row of
vertical rods was installed along that border, but no
sources of support were provided in the shaded sectors. The
angular orientation of the internodes produced above the
clamped stem section was followed over time (Fig. 5.6). WT
plants projected all new nodes to the full-light sector and
the shoots tended to become horizontal with time (Fig. 5.6).
This was so in spite of the fact that WT plants at that
stage produced a tendril in every new node, and some of the
older tendrils reached and coiled around the supporting
rods. Shoot of the lh mutant tended to grow upwardly and, by
the end of the experiment, the total population of nodes
produced by the six plants was almost equally distributed
between the full-light and shade sectors (Fig. 5.6). In some
cases failure of early tendrils to attach to the supporting
rods resulted in partial "lodging" of the shoot toward the
129
shade or full-light areas (see second, fifth, and sixth
plant in Fig. 5.6). However, the stems rapidly returned to a
nearly vertical stance. Elongation rates of tendrils
produced by WT and lh plants were similar (See Fig. 5.7A),
and the maximum length reached by the tendrils before
becoming non-functional was about 25 cm in both genotypes.
In a second experiment we tested whether the internodes
modify their direction of elongation when their distal-node
tendril becomes attached to supports. Plants were grown in
the controlled environment up to the 12-th node stage. The
tenth internode was marked, laid horizontally on a flat
surface and fixed with masking tape. The tendril produced at
the distal node of internode number 12 was placed in contact
with a small plastic rod and allowed to coil
thigmotropically (See Fig. 5.7, upper diagram). Changes in
tendril morphology and angular orientation of the internodes
were followed over a period of 12 hours. The tendril section
between the point of contact with the support and the node
elongated very little during the experimental period (Fig.
5.7A). In addition, free coils or buckles were formed along
the tendrils (Fig. 5.7B). Internodes of lh mutant plants
bent along their axis toward the side of tendril attachment
(Fig. 5.7C). This bending could not be detected in WT
internodes. The basis of this difference between WT and lh
internodes is not clear at present. Bending of lh internodes
was most likely initiated by the pulling exerted by the
coiling tendril. In fact, when tendrils were attached to
mobile supports no bending was observed (data not shown).
However, it is clear that during the process of bending in
lh there is a concomitant realignment of stem growth. This
is demonstrated by the observation that the orientation of
"curved" lh internodes changes very little upon severing the
attached tendril (Fig. 5.8). Irrespective of the mechanism,
this experiment demonstrated that tendril attachment can
130
actively alter the direction of growth of lh stems.
DISCUSSION
This study showed that cucumber plants tend to
selectively invade light gaps when growing in a patchy light
environment, and revealed that the photoreceptor phyB has a
dramatic and multiple influence on the efficiency of gap
colonization. Our results have important implications for
the understanding of mechanisms of foraging for light and
the role of information acquired by photoreceptors in
shaping plant architecture.
Mechanisms of foraging for light
Wild type and phyB-deficient lh-mutant seedlings of
cucumber actively projected their main-stem apical buds
toward artificial light gaps set up in the growth room (Fig.
5.4B). The hypocotyls of both genotypes bent toward the
light gaps soon after seedling emergence (Table 5.1). This
phototropic response has been characterized in several plant
systems (reviewed by lino 1990). In dicots, phototropism is
triggered in the hypocotyls themselves, although
differential illumination of the cotyledons may cause
hypocotyl curvatures under some circumstances (Shuttleworth
and Black 1977). Two photoreceptor systems are involved in
the detection of lateral light gradients established across
elongating organs: a B-light photoreceptor and phytochrome
(lino 1990; Ballar6 et al. 1992a, and references therein).
In the artificial full-light / shade gradient, we used
neutral screens to modify total fluence rate without
changing the R:FR ratio. Therefore, stem bending responses
observed in this experiment (Table 5.1) can be attributed
solely to the fluence rate gradient established between the
illuminated and shaded flanks of the hypocotyl, which was
most likely perceived by the B-absorbing photoreceptor.
Previous studies (Ballar6 et al. 1991b, 1992a) showed that
131
WT and lh seedlings have similar phototropic responses to B
light under conditions of continuous illumination. In
natural canopies, the changes in R and FR radiation imposed
by neighboring individuals can trigger similar phototropic
responses via phytochrome (Ballare et al. 1992a). We have
previously demonstrated that hypocotyls of cucumber
seedlings grown in the field tend to bend away from
neighboring plants that absorb R light and back-reflect FR
radiation, even if the lateral B-light gradient created by
the presence of neighbors is experimentally eliminated. This
bending response elicited by lateral R:FR gradients is
absent in the phyB-deficient lh mutant (Ballare et al.
1992a). The lack of phytochrome-mediated phototropism might
be therefore one of the reasons why the lh mutant was less
efficient than the WT at positioning nodes outside the maize
canopy in the field experiment (Fig. 5.3).
The observation that shoots of WT and lh mature plants
have different responses to gravity (orthogravitropic in lh,
diagravitropic in WT; Figs. 5.2 and 5.6) is broadly
consistent with previous findings in other species showing
that treatments that lower the Pfr level in plant tissue
(e.g. high plantation densities or supplemental FR light)
tend to produce more vertically-oriented shoots or leaves
(Casal et al. 1990; Aphalo et al. 1991). We suggest that
this regulation of gravitropism by Pfr levels plays an
important role in the adjustment of plant morphology to
patchy situations. Thus, orthogravitropic stems would confer
an advantage to plants that compete for light in a crowded
patch, but diagravitropic stems are likely to be far more
effective at invading the gaps that the shoots eventually
find when moving through the canopy.
Phototropic and gravitropic responses would establish
the initial direction of spreading in relation to the light
132
/ shade mosaic. However, an important determinant of the
ability to colonize light gaps is the extent to which the
modules that eventually reach a high-light environment are
able to stay within the gap and take advantage of the
increased PAR (Slade and Hutchings 1987, Hutchings and
Mogie, 1990). Simple models of optimal plant architecture in
heterogeneous media (Sutherland and Stillman 1988) predict
that the production of short internodes and profuse
branching in "good" sites would improve the overall
efficiency of resource acquisition by the plant. This is the
morphological pattern that we observed in WT cucumbers grown
under full light in the controlled environment (Fig. 5.1, B,
C, and D). In contrast, lh plants grown under unaltered
white light produced extremely long hypocotyls and
internodes and their axillary branches remained undeveloped
(Fig. 5.1 B, C, and D). Acceleration of stem elongation and
reduction of branching are typical responses of WT cucumbers
to natural (Banat6 et al. 1991b) and simulated (Lopez-Juez
et al. 1990) shadelight.
This and previous studies (Lopez-Juez et al. 1990) with
the lh mutant have revealed that tendril production in
cucumbers is under phytochrome control. WT plants respond
with increased tendril production when exposed to light
conditions that simulate aspects of vegetational shade (e.g.
end-of-day FR pulses, Lopez-Juez et al. 1990). WT plants in
our controlled-environment experiments (high R:FR ratio)
produced the first tendril at node number 6, between one and
one-and-a-half months after emergence (Figs. 5.1 and 5.4).
In contrast, lh-mutant seedlings started to tendril at node
number 2 (Figs. 5.1 and 5.4), and plants became attached to
the rods installed in the shaded sectors soon after
emergence (Fig. 5.4C). By that time, stems of lh plants were
inclined toward the gaps (Table 5.1, and Fig. 5.4B), but
attachment of successive tendrils to the rods was
133
accompanied by a re-direction of main-stem growth. Since lh
internodes elongated exceptionally fast (Fig. 5.1) and
curved in response to the "pulling" exerted by attached
tendrils (Fig. 5.7 and 5.8), the apical buds were rapidly
projected back into the shaded areas (Fig. 5.4B).
These dynamic studies suggest that differences between
WT and phyB-deficient lh plants in regard to the spatial
placement of nodes in natural (Fig. 5.3) and artificial
(Fig. 5.4) full-light / shade-support mosaics are a function
of differences in (i) internode elongation rate, (ii) rate
of tendril production, (iii) gravitropic responses of stems
and, probably, (iv) stem bending responses to mechanical
stimuli. In the lh mutant, the orthogravitropic habit would
limit exploration of the surroundings, tendrils would limit
motility and cause stems to bend toward points of
attachment, and, under these conditions, fast internode
elongation would entail the risk of rapid penetration into
shaded environments, where further tendrils would find
abundant sources of support. We therefore conclude that phyB
is responsible for detecting the occurrence of gaps in the
canopy and switching the plant from the "shade-avoiding"
(default!) developmental program to the one that results in
the short-internoded, more branched, and diagravitropic
phenotype. In addition to these functions, phyB is the
sensor of a phototropic system that would help WT plants to
actively avoid crowded patches in natural canopies (Ballare
et al. 1992a).
The above-mentioned response factors would determine
the three-dimensional distribution of stems and nodes in the
canopy space as a function of the spatial distribution of
light gaps and potential points of attachment. However, in
cucumbers, each individual leaf retains the capacity to
orientate itself in relation to the local light climate,
134
independently of the orientation of the supporting
internodes (Fig. 5.5). The similarities observed between WT
and lh in regard to leaf display are consistent with the
information derived from physiological experiments showing
that the B-light region of the spectrum, perceived by a
photoreceptor different from phytochrome, plays a central
role determining the direction of leaf orientation (e.g.
Koller 1990).
Role of information acquired by photoreceptors in shaping
plant architecture
The idea that plants "select" the direction of radial
expansion on the basis of mechanisms that detect
environmental heterogeneity has been suggested and discussed
(e.g. Schellner et al. 1982, Billow-Olsen et al. 1984,
Eriksson 1986, Schmid 1986, Novoplansky et al. 1990, Bazzaz
1991). Yet, observed canopy asymmetries induced by the
proximity of neighbors are commonly interpreted as products
of different levels of resource availability experienced by
different parts of an uniformly-expanding genet. Moreover,
models of foraging plants (Sutherland and Stillman 1988,
1990, de Kroon and Schieving 1991) do not consider the
possibility that plants acquire information on the spatial
distribution of resources and "choose" to project their
stems toward the most prospective sectors, although they
often incorporate morphological responses of plant modules
to the local light environment. The data used as the basis
for discussion of foraging processes commonly consist of
maps showing that the spatial distribution of leaves, buds,
or some other plant parts is non-random, and that modules
tend to concentrate in "good" sites. With few exceptions
(Billow -Olsen et al. 1984), observations on morphology and
biomass distribution are made many weeks or months after the
beginning of the experiments, which provides little
information on mechanisms and makes it difficult to
135
distinguish among simple growth responses to local stresses,
local morphological adjustment, or foraging involving
selection of the direction of spreading (see Eriksson 1986).
This study incorporated two key elements: time courses
of morphological development and comparison between
genotypes that differ only in a morphogenic photoreceptor.
Cucumber plants grown in a heterogeneous light environment
were shown to selectively project their main stems toward
the gaps (Table 5.1, and Fig. 5.4B). The ability to persist
within the invaded high-light patches was found to be
severely impaired in the phyB-deficient lh-mutants (Fig.
5.4B). Apart from the mechanisms that determine the overall
placement of stem nodes in the light / shade mosaic, a
precise regulation of leaf display in relation to the
direction of illumination was found to be operative in both
genotypes (Fig. 5.5). According to these data, we suggest
that the main effect of local differences of PAR
availability on the spatial distribution of plant growth
would be to exaggerate asymmetries that are triggered by
photomorphogenic systems that acquire information about the
radiation environment.
CONCLUSIONS
Our experiments showed that the photoreceptor phyB
substantially enhances the capacity of cucumber plants to
forage for light in horizontally-patchy canopy environments.
From these and previous experiments it is suggested that
this enhancement results from a combination of several
regulatory actions of phyB on plant growth and development.
The critical actions identified by the studies with cucumber
are:
1) Establishment of a phototropic system that responds to
lateral R:FR gradients created by asymmetric distribution of
136
neighbors around the plant (Ballare et al. 1992a).
2) Delay of tendril production under high light / high R:FR
conditions, and reduction of stem bending responses to
lateral forces created by attaching tendrils. These elements
would combine to determine that the direction of growth is
primarily dictated by the spatial distribution of light gaps
and not by the availability of support (which is likely to
be more abundant in the vicinity of other plants)
3) Inhibition of stem elongation and promotion of axillary
branching under high-light / high-R:FR conditions, which
would favor occupation of high-light gaps.
4) Modulation of mature stem gravitropic response by the
level of phyB Pfr, so that light conditions that establish
relatively low Pfr levels (vegetational shadelight) result
in orthogravitropic shoots and those that establish high Pfr
concentrations result in diagravitropic stems. The
diagravitropic habit, coupled with an efficient phototropic
system (driven in part by phyB), would help plants to
rapidly colonize light gaps and escape from shaded areas.
Acknowledgements. I am most grateful to Dr. Maarten
Koornneef and members of the Department of Genetics
(Wageningen Agricultural Univ.) for provision of the
cucumber seeds. I also wish to thank Randy Hopson (Veg. Res.
Farm, OSU) and Lloyd Graham (Forest Res. Lab., OSU) for
practical assistance, and Gretchen Bracher for preparing the
figures. Financial support from the Consejo Nacional de
Investigaciones Cientificas y Tecnicas (Argentina) and the
Dept. of Forest Science (OSU) is gratefully acknowledged.
137
TABLE 5.1. Stem length and angle of inclination toward the
artificial light gaps in WT and lh-mutant seedlings.*
Time after
emergence
genotype
(days)
4
17
length
(cm)
angle
(degrees)t
WT
2.6+0.1
35.6+1.9
lh
7.4+0.6
38.9+1.8
WT
4.3+0.1
30.7+1.9
lh
22.5+0.1
27.0+3.4
*Data are means ±SE (n=14, WT or n=12, lh).
"'Vertical = 0 degrees.
138
12
10_
S', -0
8.
6_
-E0, 0
2
4-
ci5 za-C
6.
E
z E.°
2-
-
2
0' 8
0 8
C
6-
6-
2
C Tin
4-
-
4
E
co
2-
(2)
x<
(3)
40
30
20
10
(days)
Emergence
after
Time
controlled
the
in
illumination
under
conditions
of
uniform
plants
lh
and
WT
of
development
Morphological
5.1.
FIG.
18
or
were
nodes
larger
than
the
size
of
the
symbols).
Main-stem
(whenever
+SE
indicate
bars
vertical
plants;
(1h)
replicate
(WT)
14
of
mean
the
is
point
datum
Each
environment.
one.
number
node
as
node
cotyledonary
the
from
counted
branch.
oldest
second
the
on
determined
were
lengths
Branch
E
and
C
respectively.
tendrils,
or
branches
of
axillary
appearance
the
indicate
the
number
of
nodes
produced
panels before
in
brackets
between
given
figures
The
139
WT
48 h
lh
FIG. 5.2. Gravitropic responses of mature WT and lh plants
under conditions of full light in the controlled
environment. The 12-th internode was attached to a
horizontal substrate (indicated with an X), and the angular
orientation of three successive internodes was measured 48 h
later. The segments delimited by circles indicate the
average angles of the internodes; the range is indicated by
the arcs (n=7).
140
4
2
0
2
4
6
8
30
B
0-
80
w c;;C
C
60-
c7)
Ta cl-
c 0 40-
-E.
a"
c 200
WT
lh
FIG. 5.3. Spatial distribution of nodes produced by WT and
lh plants after four weeks of growth at the border of a
maize canopy in the field. The upper panel indicates the
average number of nodes positioned inside or outside the
canopy by WT and lh plants and includes nodes of the main
stem (except the cotyledonary node) and axillary branches if
present. Thin bars represent +SE (n=20).
141
8- A
o, p WT
lh
12
10
8
6
4
2
0
-2
-4
-6
-8
10
Time after Emergence (days)
FIG. 5.4. Internode production, spatial distribution of main
stems and tendril dynamics in WT and lh plants grown for 6
weeks at the border of simulated canopies in a controlled
environment. Main-stem nodes were numbered from the
cotyledonary node as node number one. Main-stem projection
is the distance between the apical bud and the base of the
plant measured on a horizontal plane and perpendicularly to
the shade-support / full-light border (see diagram in the
upper-right corner). Positive values indicate projection of
apical buds toward the artificial gaps. Each datum point is
the average of 12 (WT) or 16 (lh) plants +SE. Plants were
considered to be "attached" when they had at least one
tendril coiled around the rods placed in the shade-support
sector.
142
Shade-support
Full-light
FIG. 5.5. Angular orientations of leaves and main-stem
internodes of WT and lh plants grown for 5 weeks at the
border of artificial canopies in the controlled environment.
Angles were measured on a vertical plane perpendicular to
the shade-support / full-light border and correspond to the
third youngest leaf of each plant and the internode below
its point of insertion; arcs indicate +SE (n=13). The angles
and lengths of petioles Were not recorded; the dashed lines
were drawn to clarify presentation assuming that the angle
between lamina and petiole was 110 degrees.
143
is
N
ca
0-
U)
ca
0-
co
is
0-
0
1
2
3
4
5
6
7
Time (days)
FIG. 5.6. Time courses of artificial gap invasion by six WT
(0) and six lh () shoots in the controlled environment. The
gray-tone and white sectors in the figure indicate the
shaded and full-light areas in the growth room,
respectively, and the dashed line denotes the row of
vertical rods established at the border of the gaps. The
ninth internode of each shoot was oriented vertically and
clamped 2 cm toward the gaps on day -1. The segments
delimited by circles indicate the angular orientation of the
successive internodes (from 10 to 14) produced by the
plants. Angles were measured on a vertical plane
perpendicular to the shade / full-light border.
144
Free coils
=t
Bending angle = a
A distance
= (rfo - n o)
A
B
lh
Zr) 1.0-
WT t
,c)
E
2
.
0)
0" 0.5a)
22
u.
0
c 12 C
T
m
v0
1
^
u
0
F
<
m
c
c0
6
3
15
m
0
-3
WT
Ih
WT
Ih
FIG. 5.7. Effects of tendril attachment to the supports on
tendril morphology and internode orientation. The upper
diagram shows a top view of the experimental setup and the
meaning of the different variables measured. The duration of
the experiments, from the beginning of tendril coiling
(dashed lines), to the final set of measurements (solid
lines) was 12 h. Tendril elongation rates given in panel A
were calculated by measuring the length of the tendril
segment between the node and the point of contact with the
support. The lines parallel to the abscissa indicate
elongation rates of tendrils before attachment to the
supports. Positive bending angles (panel B) and negative
Adistances (panel C)
indicate that the internodes curved
toward the support after tendril attachment. Each value is
the average of 22 (bending angles) or 15 observations +SE.
145
100
FIG. 5.8. Changes of internode curvature after severing the
attached tendril in lh plants. The setup sketched in Fig.
5.7 was used. Twelve hours after tendril attachment, the
internode bending angle was measured (a=+12.3 ±2.4; n=21)
and the curvature remaining after cutting the tendrils was
measured again at the indicated times.
146
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Ballare, C. L., A. L. Scopel, S. R. Radosevich, and R. E.
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150
CHAPTER 6
PHOTOMORPHOGENIC PROCESSES IN THE AGRICULTURAL ENVIRONMENT
Carlos L. Bellaire*, Ana L. Scopel, Rodolfo R. Sanchez and
Steven R. Radosevich
Depts of Forest Science and Crop and Soil Science, Oregon
State Univ., Peavy Hall 154, Corvallis, OR 97331-5705, USA
(C. L. B., A. L. S., S. R. R), Dept de Ecologia, Facultad de
Agronomla, Univ. of Buenos Aires, (1417) Buenos Aires,
Argentina (C. L. B., A. L. S., R. A. S.)
Photochemistry and Photobiolgy, in press (1992).
151
ABSTRACT'S
Studies under natural light conditions are revealing
how photomorphogenic mechanisms influence processes of
significance for agriculture. In this paper we review recent
advances in three areas: photocontrol of weed seed
germination, acclimation to solar UV-B radiation, and plant
interactions in canopies.
In the area of the photocontrol of germination there is
increasing interest for understanding the role of
phytochrome in the mechanisms whereby soil tillages trigger
weed seed germination. It has been demonstrated recently
that seeds of some weed species acquire the ability to
respond to very low photon fluences during burial. Field
studies showed that germination of sensitized seeds can be
triggered by millisecond-exposures to sunlight, and there
are indications that short light pulses of this nature may
be important light signals of soil disturbance.
In the area of UV-B acclimation, there is increasing
evidence from action spectroscopy indicating that a specific
UV-B photoreceptor participates in the control of flavonoid
accumulation in the leaf epidermis. Recent studies provided
experimental support for the notion that UV-B-induced
flavonoids play an important role protecting the
photosynthetic system against damage by UV-B radiation. Some
effects of UV-B on plant morphology have also been
interpreted as photomorphogenic responses mediated by an UVB receptor.
In the area of plant interactions in canopies, progress
has been made studying the roles of phytochrome and blue /
UV-A sensors under natural conditions. It has been shown
that, by perceiving light signals through these two
photoreceptors, plants obtain information about the
152
characteristics of the surrounding vegetation well before
they experience reductions in the availability of resources
as a consequence of neighbors' activities.
There appears to be a large potential for improving
existing cropping systems by manipulating light environment
or by deliberately changing the photomorphogenic behavior of
crop plants. However, important gaps still exist in our
understanding of how the expression and agronomic
significance of photomorphogenic responses are influenced by
other factors of the plant's environment.
tThis Abstract has been rewritten after the paper was
accepted by Photochemistry and Photobiology and is more
detailed than the published version.
153
INTRODUCTION
In this paper we review some recent advances in our
understanding of how photomorphogenic mechanisms operate
under natural conditions and influence processes that are
important in the agricultural environment. Many significant
developments have occurred over the last few years in such
disparate areas as grassland ecology and transgenic plant
technology. Our strategy is to focus on areas where a
substantial proportion of the information has been derived
from experiments carried out under conditions of natural
radiation, and where recent developments (i) imply a
qualitative change in the way an important agronomic issue
is addressed, and (ii), are likely to influence applied
agricultural research in the near future. The three areas we
discuss are:
1) photocontrol of weed seed germination in arable
land,
2) plant responses to solar UV-B radiation, and
3) photomorphogenic signals in processes of plant
competition in crop canopies.
Abbreviations: B, blue; FR, far-red; LAI, leaf area index;
LD, long day; LF, low fluence; P, total phytochrome; PAR,
photosynthetically active radiation (400-700 nm); Pfr, FRabsorbing form of phytochrome; PhyA, phytochrome-A gene;
PHYA and PHYB, phytochrome-A and -B polypeptides; R, red;
VLF, very low fluence.
PHOTOCONTROL OF WEED SEED GERMINATION IN ARABLE LAND
Soil cultivation in arable fields is typically followed
by pulses of weed seedling emergence (e.g. Chancellor 1964;
Banat-6 et al. 1988a). Moreover, seeds of some weed species
or ecotypes appear to require periodic disturbance of the
soil to lose primary dormancy and germinate (e.g. Soriano et
al. 1970). Sauer and Struik (1964) and Wesson and Wareing
154
(1969) first established a connection between light signals
and germination flushes in disturbed soil. Since then, work
in different laboratories has focused on such aspects as the
penetration of light through soil (reviewed by Tester and
Morris 1987), variations in light requirements for
germination during burial (see references in Karssen 1982),
and interactions between light and other environmental
factors (e.g. Egley 1989). Most experimentation in the last
two areas has been carried out on unearthed seed samples
that were subjected to standard germination tests under
controlled conditions. Only a small number of light
treatments were usually compared in the experiments (e.g.
darkness vs. one or two broad-band sources at one fluence).
Yet, an important generalization that can be derived from
such studies is that the light responses of weed seeds may
change dramatically over relatively short periods of time
(i.e. weeks). In a number of cases (Taylorson 1972; Baskin
and Baskin 1980, 1985), and for reasons that are not clear
even at the simplest physiological level, the light
requirement for germination appears to fluctuate in a
seasonal fashion during burial.
Most laboratory studies on phytochrome control of
germination have been carried out with seeds that displayed
red (R)+-far-red (FR) reversible photoresponses (i.e. low
fluence [LF] responses). In recent years, effects of light
on germination that are not reversible by FR have received
attention. Using suitable pretreatments (e.g. chilling,
giberellic acid, ethanol), the sensitivity of seeds of some
species to Pfr can be increased several orders of magnitude
(VanDerWoude and Toole 1980; VanDerWoude 1985; Cone et al.
1985; Rethy et al. 1987; Taylorson and Dinola 1989).
Germination of sensitized seeds can be triggered by light
treatments that establish < 0.01 % Pfr. Because a small
155
number of light quanta are required to obtain such low
levels of Pfr, this photoresponse has been termed very-lowfluence (VLF) response. Although VLF responses can be
readily demonstrated in the laboratory, the ecological
significance of the very high light sensitivity may be not
immediately obvious (Smith and Whitelam 1990).
Bouwmeester and Karssen (1989) showed that the
expression of the dormancy status of seeds retrieved from
soil can be substantially influenced by pretreatments (e.g.
drying of the seed sample) and incubation conditions during
the germination tests. Therefore, little of the information
derived from standard tests under laboratory conditions can
be directly used to establish the nature of the light
signals that trigger weed seed germination in the field.
Indeed, we still know relatively little about how important
light signals are in relation to other environmental
variables that are also affected by tillage (see also
Woolley and Stoller 1978). The frequently held view that
light is the ultimate signal for germination induction in
light-requiring seeds should not be accepted without
scrutiny. In Datura ferox, where a light requirement for
germination has been well established, formation of Pfr must
be followed by a period of alternating temperatures in order
to obtain full expression of the germination response. Pfr
can be stored for several hours or days until the seeds are
exposed to temperature fluctuations (Casal et al. 1991). In
turn, tillages can profoundly affect the thermal environment
sensed by seeds by affecting the amount of residues left on
the ground surface and the vertical distribution of the seed
population in the soil profile.
Some recent studies have specifically focused on the
interphase between photomorphogenesis and seed ecology in
arable land. In a series of laboratory experiments, Scopel
156
et al. (1991) studied the germination responses of seeds of
D. ferox to R and FR light pulses that established different
calculated percentages of Pfr. Seeds that had been kept in
dry storage in the laboratory only germinated when the Pfr
level established by the light treatments was above 2 % and
the germination-%Pfr response function resembled those
obtained for typical LF responses. Similar responses are
usually found in freshly collected seeds of this species
(Casal et al. 1991). In contrast, most of the seed
population retrieved from soil could be induced to germinate
by pulses that provided < 0.01 % Pfr, and the response curve
was clearly biphasic. This increase in light sensitivity was
interpreted as a natural transition from the LF response to
the VLF response. A period of cold temperature did not
appear to be required for the increase in light sensitivity
in D. ferox. In a related field experiment, sunlight pulses
were given to seed samples that were incubated in the soil
at 7 cm depth after exposures to sunlight. The goal was to
mimic the effect of soil cultivation. The major findings
were: (1) exposure to light was essential to trigger
germination in the soil, (2) most of the seeds could be
induced to germinate by light pulses equivalent to 10 ms of
full sunlight, which would establish < 0.01 %Pfr
(temperature fluctuations were not limiting in this
experiment), and (3) less than 10 % of the effect of
saturating exposures to sunlight could be reversed by
subsequent FR. These results show that VLF responses can be
demonstrated in the field. Other observations of high light
sensitivity in exhumed seeds have been reported (Taylorson
1972; Baskin and Baskin 1979), which suggests that
germination responses to VLFs are probably very frequent
among seeds that have spent some time in the soil.
Scopel et al. (1991) have suggested that the natural
switch to the VLF state may be essential for detection by
157
seeds of very short exposures to sunlight while the soil is
being disturbed through tillage. How important these short
pulses are in relation to other light signals is not clear
at present. Scopel et al. (1991) hypothesized that such
factors as desiccation, predation, and photoinhibition may
reduce the contribution of seeds left at the soil surface to
the flush of germination. It should be noted, however, that
seeds buried a few mm might be protected from some of these
hazards and could integrate a weak light signal (Tester and
Morris 1987; Mandoli et al. 1990) over several photoperiods
after tillage, which might yield enough Pfr to trigger
germination in a population of sensitized seeds. In
principle, this possibility is not unreasonable since, in
Datura ferox at least, promotion of seed germination by
light is mediated by a stable type of phytochrome (Casal et
al. 1991; see also Lercari and Lipucci di Paola 1991). The
interesting observations of Hartmann and Nezadal (1990)
indirectly support the suggestion that light pulses received
during the actual episode of disturbance are key signals for
germination. They found that weed populations declined
dramatically in experimental plots where all soil disturbing
operations were conducted during the night. The effect of
artificial lighting during the night tillage was not tested.
Given the increased concern about massive herbicide inputs
and the need to find alternative methods of weed control
(e.g. Frisbie and Smith 1990), the issue has substantial
practical interest and warrants further attention.
PLANT RESPONSES TO SOLAR UV-B
Concern about the impact of high levels of solar UV-B
radiation on natural and cultivated plant communities has
increased after recent indications of a global reduction of
the stratospheric ozone layer (Kerr 1988, 1991; Solomon
1990) and a trend of increasing surface UV-B irradiances in
some regions (Lubin and Frederick 1989; Blumthaler and
158
Ambach 1990). Tevini and Teramura (1989) and Caldwell et al.
(1989) have reviewed most of the current literature on
ecophysiological effects of UV-B on higher plants. We will
briefly discuss recent work on pigmentation and
morphological responses to UV-B and its agricultural
significance. Light induction of photoreactivating enzymes
will not be covered here (See Langer and Wellmann 1990; Pang
and Hays 1991).
Light promotes the accumulation of anthocyanins and
flavonoids in many plant systems. Visible light is sometimes
effective (acting through phytochrome and blue / UV-A
receptors) but, in many cases, exposure to UV radiation is
required for flavonoid induction (See Beggs et al. 1986a,b).
Action spectra for UV-induced synthesis of anthocyanins and
flavonoids are characterized by maximum quantum
effectiveness in the UV-B region (i.e. 290 - 300 nm;
reviewed by Wellmann 1983; Beggs et al. 1986b; Hashimoto
1990). In contrast, action spectra for damaging effects of
UV radiation in plant systems, such as inhibition of
photosynthesis in intact leaves (Caldwell et al. 1986),
inhibition of photosystem II activity in thylakoid
suspensions (Bornman et al. 1984) and inhibition of R- or
R+UV-B-induced anthocyanin synthesis (Wellmann et al. 1984;
Hashimoto et al. 1991) show greater photon effectiveness at
shorter wavelengths. This observation has lent to the
suggestion that the effect of UV-B on flavonoid synthesis is
mediated by a specific photoreceptor (Beggs et al. 1986b;
Hashimoto 1990). Apparently complex interactions between UVB and light absorbed by other photoreceptors have been
reported and Beggs et al. (1986b) concluded that in most
systems it would seem that the presence of Pfr is at least
necessary for full expression to the response to UV-B.
Several enzymes of the flavonoid synthetic pathway are
159
induced by light and in some cases it has been possible to
relate the increases in enzyme activity with increases in
the rates of transcription of the relevant genes (Refs. in
Beggs et al. 1986b and Hahlbrock and Sheel 1989). Rapid
effects of light on the activity of phenylalanine ammonialyase (PAL), an enzyme of the general phenylpropanoid
pathway, mediated by isomerization of the PAL inhibitor
trans-cinnamic acid has also been reported (Tevini and
Teramura 1989). Work with mustard (Sinapis alba) cotyledons
has indicated that mechanisms regulated by phytochrome
beyond enzyme induction also influence the rate of
accumulation of some products of the flavonoid pathway
(Brodenfeldt and Mohr 1988). Considerable progress has been
made over the last few years in elucidating the molecular
mechanisms that mediate the effects of light and other
environmental factors on the expression of genes encoding
enzymes of the flavonoid pathway (See Hahlbrock and Sheel
1989; Fritze et al. 1991 and references therein).
Flavonoids absorb effectively in the UV, show little
absorption in the visible region and accumulate in the
vacuoles of epidermal cells. In parsley (Petroselinum
crispum) leaves the whole sequence of light-triggered
biosynthetic events leading to flavonoid production have
been localized in the epidermis (Schmelzer et al. 1988).
These observations are consistent with the view that
flavonoids play an important role as natural UV filters,
protecting plant tissues from deleterious effects of UV-B
radiation (Caldwell et al. 1983). Under natural conditions
the fluxes of UV-B and visible radiation are poorly
correlated (Caldwell 1971). Therefore, the involvement of a
specific UV-B photoreceptor in the control of flavonoid
accumulation might be adaptive, as this sensor would provide
information on the UV-B environment that plants can not
derive from light signals sensed by other photomorphogenic
160
pigments. Indirect evidence supporting an important role of
flavonoids in acclimation to high UV-B levels has been
obtained by examining the natural variation in leaf
epidermal transmittance along geographic gradients that are
associated with large changes in solar UV-B irradiance.
Robberecht et al. (1980) found that in plants from lowlatitude alpine environments (high UV-B), UV epidermal
transmittance was always very low (<2 %). In contrast, at
high and medium latitudes (lower UV-B) there was a larger
variability in epidermal transmittance among species. Plants
exposed to experimentally enhanced UV-B levels under field
conditions often respond with increased flavonoid
accumulation and decreased UV-epidermal transmittance (Flint
et al. 1985). More direct tests of the protective role of
flavonoids have recently been reported. Tevini et al. (1991)
used variable chlorophyll fluorescence to measure the
effects of UV-B on photosynthetic activity in rye (Secale
cereale) primary leaves that had accumulated different
amounts of UV-B absorbing compounds in the epidermis. They
found that short white-light + UV-B pretreatments, which
stimulated flavonoid accumulation and decreased UV
transmittance of the epidermis, reduced the negative impact
of short-wave UV-B treatments on the estimated quantum yield
and photosynthetic capacity. A similar approach had been
previously used by Beggs et al. (1986a) to evaluate the role
of flavonoid pigments in protecting parsley roots against
UV-B damage. The UV-B fluences required to produce a
significant accumulation of flavonoids in the rye system
were roughly similar to those received at the Earth's
surface during a clear summer day at mid latitudes. However,
as the authors acknowledge, a full assessment of the
ecological significance of their findings requires more
study. In part this is because the spectral distributions of
artificial and solar UV-B are not identical, which makes it
necessary to use some type of "weighing" function for
161
comparisons (see Caldwell et al. 1986). In addition, other
factors may influence the degree of protection achieved by
leaves under field conditions. For instance, increases in
leaf thickness in response to high levels of
photosynthetically active radiation (PAR) (Warner and
Caldwell 1983) and UV-B (Cen and Bornman 1990; Bornman and
Vogelmann 1991) may afford some additional protection
against UV-B.
Besides influencing pigment synthesis, UV-B may induce
a number of changes in plant morphology. Attempts to unravel
the underlying mechanisms are typically complicated by the
fact that, particularly in controlled-environment
experiments with low PAR, morphological responses to UV-B
are usually accompanied by reductions in photosynthesis and
total growth (Tevini and Teramura 1989). However, a number
of effects of UV-B on morphology are difficult to interpret
as mere by-products of photosynthetic inhibition. Sisson and
Caldwell (1976) found that UV-B inhibits leaf elongation in
Rumex patientia, and the effect is not correlated with
changes in photosynthesis. Recent field and glasshouse
experiments have shown that UV-B reduces shoot (leaf)
lengthening in grasses without affecting total growth
(Barnes et al. 1988, 1990) or photosynthesis (Beyschlag et
al. 1988). UV-B also inhibits axis elongation in etiolated
seedlings (Steinmetz and Wellmann 1986). In de-etiolated
cucumbers (Cucumis sativus) growing in a glasshouse, 8-h
exposures to supplemental UV-B inhibit hypocotyl elongation
up to 50 % without affecting growth rate or cotyledon area
expansion (Ballare et al. 1991a). Morphological effects of
UV-B may be the consequence of disturbing effects of UV-B on
cellular functions other than photosynthesis (e.g. damage to
meristems or elongating cells). In fact, the action spectrum
for the inhibition of hypocotyl elongation in Lepidium
sativus resembles those obtained for DNA damage (Steinmetz
162
and Wellmann 1986). In cucumbers the inhibitory effect of
UV-B on elongation is triggered mainly in the cotyledons;
therefore, direct damage to hypocotyl or apical cells is
obviously not involved in the response (Ballare et al.
1991a). Barnes et al. (1990) have shown that moderate levels
of UV-B applied to glasshouse-growing plants stimulate
tillering in some grass species. These lines of evidence
suggest that some responses to UV-B radiation should be
regarded as true photomorphogenic effects. Recent field
studies (Barnes et al. 1988) and simulations using a multispecies canopy model (Ryel et al. 1990) suggest that
differential morphological responses to UV-B are important
in determining the balance of competition between species in
cultivated plant associations and should be taken into
account when trying to predict the consequences of
stratospheric ozone depletion at the plant community level.
PHOTOMORPHOGENIC SIGNALS IN PROCESSES OF PLANT COMPETITION
The idea that neighboring plants in a cultivated plant
association compete with each other for access to
environmental resources is familiar. Yet, the factors that
determine variations in competitive success among plant
individuals are often hotly discussed among ecologists and
agronomists. Recent studies highlight the potential
significance of information acquired by plant photoreceptors
in the processes of competition for light. We will review
some of the major findings and discuss them in relation to
their potential significance for crop improvement.
R:FR signals and phytochrome
Spectroradiometric studies of the natural light
environment (e.g. Kasperbauer 1971; Holmes and Smith 1977)
and physiological experiments with light-grown plants (e.g.
Morgan and Smith 1978) have demonstrated that a number of
typical morphological reactions to shadelight (increased
163
stem elongation, reduced leaf to stem dry-weight ratio) can
be triggered by phytochrome in response to the low R:FR
ratios that prevail under leaf canopies (See reviews by
Smith 1982, 1986). Results of field- and glasshouseexperiments with light-grown seedlings of the lh mutant of
cucumber, which is deficient in a PHYB polypeptide (LopezJuez et al. 1992), are fully consistent with the notion that
this phytochrome species plays a fundamental role in
elongation responses to shadelight (Ballare et al. 1991b).
It has been shown recently that, in even canopies (i.e.
canopies formed by plants of similar height), several
morphological responses to the proximity of other plant
individuals, such as reduced tillering rate in grasses
(Casal et al. 1986) and increased stem elongation rate in
dicotyledonous seedlings (Ballare et al. 1987, 1988b), can
be detected well before the onset of severe mutual shading
among neighbors. This observation led Ballare et al. (1987)
to propose that the FR radiation reflected by nearby leaves
may work as an early signal of oncoming competition during
canopy development.
Plant tissues absorb very little in the FR region, and
most of the photons in this waveband exit plant organs as
scattered radiation. Kasperbauer et al. (1984) found that
the R:FR ratio measured with cosine-corrected light
receivers at the outer strata of soybean canopies varied
between the east and west side of north-south oriented rows.
The differences were attributed to variations in the
contribution of FR reflected by adjacent plants. Using
cuvettes containing purified phytochrome, Smith et al.
(1990) found that the Pfr/P ratio at photoequilibrium
decreases progressively as the cuvettes are moved closer to
sunlit plant fences. Measurements obtained in even canopies
of dicotyledonous seedlings of very low density (leaf area
164
index [LAI] < 1), where mutual shading among neighbors is
negligible, indicated that the proximity of plant
individuals causes a marked increase in the amount of FR
received at the surface of (vertically oriented) stems
(Ballare et al. 1987, 1991c). Fiber optic studies showed
that this increase in FR receipt is paralleled by a
detectable increase in the amount of FR scattered inside the
stems, a concomitant drop of the R:FR ratio, and a reduction
in the estimated phytochrome photoequilibrium which are
quantitatively related with the LAI of the population
(Ballare et al. 1989, 1991c).
Several experiments in which mirrors and plant hedges
of different colors were used to provide additional FR to
plants growing in the field demonstrated that stem
elongation can be promoted by spectral changes that mimic
the proximity of other plants, even if the seedlings are
growing under full sunlight (e.g. Ballare et al. 1987,
1989). Comparable results have been obtained in studies of
other morphological responses (e.g. Casal et al. 1987;
Novoplansky et al. 1990; Davies and Simmons 1991). Ballare
et al. (1990) provided direct evidence for the involvement
of R:FR ratio by showing that, in seedling canopies of low
LAI, the typical stem elongation responses to increased
population density can be reduced or abolished by covering
individual stem sections with FR-absorbing filters.
Phototropic responses elicited by non-shading neighbors and
mediated by phytochrome have also been demonstrated (Ballare
et al. 1992).
Fluence-rate signals
Recent work under natural conditions of radiation has
shown that, in even plant canopies, there are at least two
more photomorphogenic factors that influence stem elongation
apart from the change in R:FR ratio. First, when the canopy
165
begins to close, there is a rapid drop in blue (B) light
irradiance at the stem level. Simulation experiments have
shown that local depressions of B light trigger an
acceleration of stem elongation rate in plants growing under
sunlight. This effect is not mediated by phytochrome or
photosynthetic pigments and, most likely, involves the
action of a specific B light photoreceptor (Ballare et al.
1991c). Second, the drop in R and FR irradiances that occurs
under similar conditions may stimulate elongation
independently of changes in R:FR ratio. An inverse
relationship between R+FR irradiance received by the stem
and elongation rate has been documented in sunlight-grown
mustard and Datura ferox seedlings (Ballare et al. 1991c).
Responses to R:FR ratio and irradiance might be mediated by
different phytochrome species. Transgenic tobacco plants
that overexpress an oat phyA gene, and maintain abnormally
high levels of oat PHYA polypeptide when grown in the light,
are dwarf and respond to additional FR with further
reduction in stem elongation (McCormac et al. 1991).
An important conclusion derived from spectroradiometer
studies in even canopies of dicotyledonous seedlings is
that, at low LAIs, increases in plant density can bring
about significant spectral and fluence-rate changes at the
level of the stem with minimal effects on the light climate
of the leaves (Ballare et al. 1989, 1991c; see Mancinelli
(1991] for a discussion of the influence of plant geometry
on light perception). In these conditions, therefore, there
is a clear separation between the flux of photomorphogenic
information received by the plant and the effects of
neighbors on the flux of photosynthetic light energy.
Collectively, these results provide a mechanistic
explanation for the early morphological responses to canopy
density that are observed in even-aged populations of
seedlings.
166
Implications for crop improvement
Going back to the starting point of this discussion,
the role of photoreceptors in plant competition, these
studies provide clear evidence for the notion that plant
growth and morphology in crop canopies is affected by
neighboring individuals not only because they directly
affect the availability of resources that are necessary for
growth but also because they modify aspects of the light
environment that are used by plants to make developmental
"decisions". Thus, in even plant stands, the R:FR ratio and
fluence-rate perceived by the stems convey information on
the proximity of neighbors that each individual plant uses
to make decisions about morphology and assimilate allocation
as the canopy develops. Since morphological changes elicited
by these signals (e.g. increased stem elongation rate) may
have a major impact on the resource-harvesting capacity of
the plant (e.g. Ballard et al. 1988b), it may be anticipated
that variations in the ability to gather and interpret
photomorphogenic information would dramatically affect
competitive ability in fast-growing plant stands.
The concept is important in attempts to understand
weed-crop competition or to explain yield-density responses
of field crops. Regarding possible strategies for crop
improvement, it is valid to ask: Can we engineer better crop
plants by altering specific aspects of their
photomorphogenic behavior? (See also Smith 1991). We will
discuss some potentially rewarding research avenues in the
remaining of this section.
Stem elongation responses. Factors that confer
competitive ability to the individual crop plant are not
necessarily "desirable" agronomic traits (See discussions by
Donald [1968] and Marshall [1991]). As discussed earlier in
this paper, the initial response of seedlings of shade-
167
intolerant species to an increase in canopy LAI is a rapid
acceleration of stem growth that is triggered by a variety
of light signals. It is tempting to speculate, therefore,
that genotypes showing reduced stem elongation responses to
photomorphogenic signals of plant proximity may save
assimilates that could be used to grow more leaves, roots,
or reproductive structures. However, two important points
should be considered. First, the increase in stem length may
be proportionally larger than the increase in stem biomass,
and second, the increase of stem biomass may imply more than
a simple redistribution of a fixed pool of photosynthate. In
a series of glasshouse experiments with amaranths
(Amaranthus quitensis), Ballare et al. (1991d) showed that
canopies accumulate much more dry matter in the internodes
when they receive the full sunlight spectrum than when they
are grown under FR-absorbing filters. However, screening the
FR did not promote the growth of leaves or roots in these
experimental crops. In other words, the extra-carbon used to
sustain higher rates of stem growth under full-spectrum
conditions resulted from an increase in canopy net
photosynthesis. In turn, the increase in canopy productivity
appeared to be related to the stem elongation responses,
because filtering the FR had no effect on biomass
accumulation when plants were grown as isolated individuals.
There are at least two possible explanations for these
results. First, an increase in internode elongation
(triggered by low R:FR ratios) might alter the pattern of
light penetration in the canopy, resulting in increased
light interception and/or more efficient utilization of the
intercepted light. Second, an increase in stem growth may
stimulate leaf-source photosynthesis through a feedback
mechanism. This latter possibility has been indirectly
supported by experiments with isolated plants showing that,
if the R light irradiance received by the stem is reduced,
elongation and biomass accumulation in the internodes can be
168
substantially increased without affecting the growth of
other plant parts (i.e. total plant biomass increases as a
result of a localized reduction of R:FR ratio) (Ballare et
al. 1991d). Responses of this type are not likely to be
observed in experiments carried out under low irradiances.
In wheat (Triticum aestivum) crops, the Rhtl and Rht2
alleles are known to reduce plant height and increase grain
yield, but the causal links between these two effects are
still not completely clear (e.g. Borrell et al. 1991).
Moreover, in some species the assimilate invested in stems
contributes a significant proportion of the carbohydrate
used to fill harvestable organs towards the end of the crop
cycle (Incoll and Neales 1970). Testing different varieties
of one such crop, Helianthus tuberosus, Soja and Haunold
(1991) found that tuber yield was significantly and
positively correlated with mid-season stem dry weight; midseason leaf area and maximum leaf photosynthesis were not
good predictors of crop yield. Low assimilate-storage
capacity, whether in stems or in tubers, appeared to be an
important limitation to crop yield. It is not clear to what
extent the storage capacity of the stems is influenced by
the amount of carbon invested in structural stem material,
which, in turn, is likely to be modulated by
photomorphogenic factors. To conclude, the evidence reviewed
in this section indicates that the estimation of the
opportunity cost of the assimilate invested in stem
elongation would require careful experimentation under field
conditions.
It should be noted that photomorphogenic elongation
responses may affect crop productivity independently of
their effects on carbon allocation. Thus, a high sensitivity
to light signals of plant proximity will have detrimental
effects whenever lodging is a factor affecting yield, but
might be desirable in the case of timber trees to favor
169
rapid growth in height and reduced branching.
Branching. It is well established that apical dominance
is influenced by the R:FR ratio of the incident radiation
acting through phytochrome in dicotyledonous plants (e.g.
Tucker and Mansfield 1972) and grasses (Deregibus et al.
1983, Kasperbauer and Karlen 1986). The axillary buds are
one of the sites of light perception: local reductions of
R:FR obtained by increasing FR (Casal et al. 1987) or by
reducing R (Begonia et al. 1988, their "FR" treatment) lead
to reduced branching. Experiments under natural radiation
have demonstrated that in open grass canopies the drop in
R:FR ratio caused by neighboring plants can reduce branching
(tillering) rate even if the production of new branches is
not limited by the availability of photosynthetic light
energy (Casal et al. 1986). The evidence is based on the
following observations: (1) tillering in a number of
temperate grasses was found to be very sensitive to small
depressions in R:FR below sunlight values (Casal et al.
1985, 1987), and (2), in sparse grass populations, the
detrimental effect of neighbors on tillering rate can be
countered by delivering small amounts of R light to the base
of the plants (Deregibus et al. 1985, Casal et al. 1986).
Little is known about the role played by photomorphogenic
signals in the control of branching in dense canopies, where
resource limitations are likely to be severe, and where many
aspects of the light environment sensed by buds and other
plant parts are simultaneously altered. Frequently, the
inhibitory effects of heavy natural- or simulated-leaf
shading (i.e.: low B and R, high FR) on branching and ramet
production are more severe than those produced by comparable
reductions of PAR obtained by means of neutral filters
(Solangaarachchi and Harper 1987, Methy et al. 1990). This
is most likely due to differences between shading treatments
in the R:FR ratio, but the role of direct responses to B and
170
R fluence rate has not been investigated. A better
understanding of the photomorphogenic regulation of
branching would provide the bases for incorporating specific
patterns of response to plant population density. In grasses
and cereals, a high sensitivity to small reductions in R:FR
would be desirable in species or agronomic scenarios where
tillering does not contribute to yield stability and reduces
the harvest index (i.e. high-input agricultural systems;
Donald 1968, 1979, Bugbee and Salisbury 1989), but may be
deleterious in less predictable environments (e.g. Kirby and
Faris 1972).
Developmental timing. One potential mechanism for crop
improvement is to alter the relative lengths of successive
developmental phases (e.g. Rawson 1988). Thus, crop
genotypes may be designed so that their phenology is better
adjusted to match the temporal pattern of resource
availability in a given environment. Mondal et al. (1986a)
reported detailed studies on onion (Allium cepa) development
in different cultural environments. They found that a number
of key developmental events, such as the transition to bulb
scale production and bulb maturity, are substantially
advanced when planting density is increased. Bulbification
in onions is stimulated by long days (LD) and it is well
established that addition of FR during the photoperiod can
accelerate development in this (Lercari 1982, Mondal et al.
1986b) and other (e.g. Deitzer et al. 1979) LD plants. The
effect of FR has the characteristics of a classical high-
irradiance response and, therefore, it is most likely
mediated by Type I phytochrome (Carr-Smith et al. 1989).
Further field experiments by Mondal et al. (1986c) showed
that in onion plants shaded by green leaves (low PAR, high
FR), leaf production ceased before and maturity occurred
earlier than in plants shaded by neutral filters (low PAR,
low FR). The effects of varying PAR alone on bulb
171
development were small. These studies suggest that changes
in the canopy light environment perceived by phytochrome may
be involved in the modulation of the observed developmental
responses of onion plants to population density. It is
interesting to note that in other species where flowering is
promoted by LD, such as barley (Hordeum vulgare) (Kirby
1967), radish (Raphanus sativus) (Weston 1982) and wheat (Yu
et al. 1988), increasing population density typically has
the effect of accelerating reproductive development. In many
LD plants, floral formation is closely associated with rapid
stem elongation ("bolting") (e.g. Metzger 1987). Therefore,
one of the advantages that an individual plant may gain from
accelerating reproductive development in a crowded
population would be to anticipate its neighbors in the race
for colonizing the aerial space. A better understanding of
the action of phytochrome(s) in the photoperiodic induction
of LD plants, and more detailed physiological experiments in
the field, would help to interpret the observed
relationships between plant density and developmental timing
and may provide clues to crop improvement programs.
Flower and fruit abscission and reproductive
allocation. As evidenced by some of the work discussed
earlier in this review, the study of the photoregulation of
assimilate partitioning among plant organs and its relation
to crop productivity has considerable agronomic interest
(see also Kasperbauer et al. 1984, Britz 1990 and references
therein). In seed crops, the allocation of photosynthate to
reproductive structures is obviously a major determinant of
yield. Heindl and Brun (1983) found that the abscission of
reproductive structures in soybean (Glycine max) crops can
be, in certain cases, reduced under field conditions by
lighting the lower canopy strata during flowering and early
pod growth with R or white light. Responses to R and white
light were similar in their experiments (in spite of the
172
fact that the R source provided -3 times less PAR quanta)
and were not observed when the reproductive primordia were
artificially shaded. Subsequent studies (Myers et al. 1987)
showed that R light (compared with FR or darkness) increases
sucrose accumulation by soybean racemes cultured in vitro.
Similar results had been obtained previously by Mor et al.
(1980) working with dark-adapted rose (Rosa hvbrida) shoot
tips. In growth chamber experiments with whole soybean
plants, R light applied to the racemes via fiber optic
guides was found to increase fruit set compared with FR and
control (no lighting) treatments (Myers et al. 1987). These
observations led to the conclusion that reproductive
abscission in the lower soybean canopy and assimilate
accumulation by young fruits are modulated by
photomorphogenic signals perceived by the primordia (Heindl
and Brun 1983, Myers et al. 1987). In other crops it has
been shown that abscission of reproductive structures
induced by shading can be prevented by applications of
inhibitors of ethylene action (Wien and Zhang 1991). This
suggests that the actual level of abscission may be set by
factors other than assimilate supply. Although it is not
easy to visualize an ecological scenario in which such a
response mechanism could have been selected, the results
obtained with soybeans suggest that there is scope for
reducing abscission by manipulating photomorphogenic
responses of reproductive primordia. The impact of such
manipulations on crop yield will depend on (i) the relative
importance of reproductive abscission as a determinant of
seed numbers per unit area, and (ii) whether or not
increases in seed numbers are accompanied by compensatory
reductions in individual seed weight. A number of
observations suggest that soybean yields are limited by
canopy photosynthesis (Egli and Yu 1991 and references
therein), which would imply that the chances of improving
crop yields by reducing reproductive abscission are small.
173
On the other hand, other field studies with soybeans suggest
that leaf photosynthesis can be enhanced by treatments that
increase the number of reproductive sinks (reviewed by
Shibles et al. 1987). These considerations indicate that
carefully designed field experiments will be required in
order to asses the agronomic consequences of interfering
with the photocontrol of flower and fruit abscission.
SUMMARY AND OUTLOOK
Plant photomorphogenesis has evolved in dark rooms and
growth cabinets as a vigorous research area. Studies in
controlled environments provided the template, and indeed
much of the inspiration for more field-based projects. In
turn, recent research in natural light environments has
raised issues and questions that had not received attention
before. Field experiments on the photocontrol of seed
germination drew attention to the dynamics of light
sensitivity and demonstrated that seeds of some weed species
naturally switch to the VLF state after a short period of
burial. This transition might be a crucial step in the
sequence of events that enable light-requiring weed seeds to
detect soil disturbances in arable fields. This hypothesis
is supported by circumstantial evidence, but more direct
experimentation is needed to test the ecological and
management consequences of changes in light sensitivity. In
the area of UV-B photophysiology, it has been clearly shown
that UV-B irradiation may cause alterations in plant
morphology without affecting total growth or photosynthesis.
These morphological responses, that could be mediated by a
specific UV-B photoreceptor, may have important consequences
on competitive interactions among plant species in mixed
canopies. Field experiments also revealed the limitations of
studies of UV-B effects on plant productivity carried out
under low levels of PAR. The hypothesis that a specific UV-B
photoreceptor is involved in the induction of flavonoid
174
accumulation in the epidermal cells by UV radiation is
supported by action spectroscopy, and the protective role
played by these compounds as natural UV-filters has been
clearly demonstrated in controlled-environment experiments.
In the area of photomorphogenesis in plant canopies, recent
studies have shown that, by perceiving R:FR and fluence-rate
signals through phytochrome(s) and a specific B light
receptor, plants obtain information about the proximity of
other plant individuals well before the PAR received by
leaves is reduced by the activities of neighbors. This
demonstration suggest that competitive interactions among
plants in crop canopies may be much more complex than
previously thought. The ability of individual plants to
obtain photomorphogenic information may indeed be crucial
for success in the competition for light in dense crops.
Studies under natural light conditions are also contributing
to clarify the role of photomorphogenic signals in processes
such as organ abscission and assimilate partitioning. The
observation that stem-perceived light signals may stimulate
stem growth without affecting biomass accumulation by other
plant parts suggests that the assumptions that underlie
calculations of the costs associated with photomorphogenic
responses should be tested experimentally under field
conditions. In addition to the well known changes in
morphology, some species respond to increased plant density
with accelerated development. The photomorphogenic bases of
this phenomenon are still not well understood.
These developments are very likely to influence more
applied agricultural research in the near future. The
general conclusion that we draw from this review is that the
acquisition of information through morphogenic
photoreceptors plays a substantial role controlling plant
behavior in agricultural environments. This is usually
overlooked in conventional texts on physiological plant
175
ecology and crop physiology. The increases in crop yield
that occurred over the last decades have been largely based
on increasing the amount of resources available to
"desirable" plants via fertilization, irrigation, and weed
control. The evidence reviewed here suggests that further
improvement can be achieved by manipulating the way in which
plants gather and interpret photomorphogenic information.
One way to this end is the production of genetically
engineered crop plants with specific alterations in
photomorphogenic behavior. Such endeavors would require more
knowledge on the photobiology of plants under natural field
conditions. In fact, most photomorphogenic responses may
have a large impact on fluxes of energy and materials
between the plant and the environment and among plant parts.
Therefore, their occurrence and agronomic impact will be
influenced by other environmental variables that affect the
"economy" of the plant, particularly the availability of
resources (i.e. light and nutrient levels). A particularly
important question posed by some of the examples discussed
in this review is to what extent carbon fixation by the
canopy and assimilate allocation patterns in the field are
influenced by photomorphogenic processes taking place in the
assimilate sinks (e.g. stem internodes, reproductive
primordia). Experiments under natural conditions of
radiation are necessary to solve this question and to
evaluate the potential benefits of modifying the
photophysiology of sink organs.
Acknowledgements. This review was written while C.L.B. and
A.L.S. were on a leave at Oregon State University financed
by the Consejo Nacional de Investigaciones Cientificas y
Tecnicas (Argentina) and the Dept. of Forest Science (Oregon
State University). We thank Barbara Yoder for critical
comments on an early draft of the manuscript.
176
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