Pioneers in chloroplast & plant molecular
biology
Ruth Sager
(Harvard)
Discovered
chloroplast genetics
(Chlamydomonas)
and DNA
Lawrence Bogorad
(Harvard)
Chloroplast genes in
corn; demonstrated
light regulation
Jean-David Rochaix
(Geneva)
Chloroplast gene
function; nuclear
control
Chloroplast Biology
I. Structure
1.
2.
3.
4.
double-membrane envelope
stroma: large soluble interior
thylakoid membrane system
intrathylakoid space or lumen
From Gunning
& Steer
Isolated Spinach chloroplast
thylakoid membrane
stroma
envelope
From Hoober
High magnification of thylakoid membranes
From Hoober
Stroma of
Avena (rye)
plastid at high
magnification
to show
ribosomes
S – aggregates of
a glucosidase used
in defense against
fungi, substrate is
in vacuole
(avenacoside),
product is a toxic
saponin
II. Functions
1. many important biochemical (anabolic)
pathways, e.g.,
photosynthesis*
starch synthesis
fatty acid synthesis*
amino acids synthesis
pigment synthesis*
nucleotide synthesis
nucleic acids* and protein synthesis
sulfur and nitrogen assimilation
2. own genetic system*
* Indicates that pathway involves a chloroplast
encoded gene in at least some organisms
Many of the biosynthetic
pathways are regulated,
to peak during the light
period of L-D cycle.
Chlorophyll a and b
synthesis during a 24
hour light-dark cycle
(Chlamydomonas).
A similar result was
obtained for carotenoids
and other chloroplast lipids.
(Janero and Barnett, 1982)
III. Reproduction
• all plant and eukaryotic algal
cells have plastids
• chloroplasts form by division;
semi-autonomous
• Involves proteins (Fts) similar
to those that mediate cell
division in bacteria
Cyanidioschyzon chloroplast
dividing
From Miyagishima et al.
IV. Development
There are several forms of plastids:
1. Proplastids - precursor of all plastids, found in
meristems
2. Etioplasts - form in shoots of dark-grown
plants, distinctive internal structure
3. Chloroplasts - in all green tissues
4. Amyloplasts - prominent in roots, store starch,
colorless
5. Chromoplasts - in mature fruit, lots of
carotenoids, little chlorophyll
Plastid development in dark-grown barley
Oldest
Mature
Etioplast
Youngest
Proplastid
Amyloplasts from Glycine (soybean) root-cap
(peripheral cell)
S - starch grains
Also have some thylakoid membranes (star) close-by.
From Gunning & Steer
From U. Wisconsin Botany Dept.
Young chromoplast from developing tomato fruit
Stars mark lycopene crystals; many plastoglobuli
From Gunning and Steer
Plastid development is plastic & mostly under
nuclear control.
Shoots:
light
proplastids
etioplasts
chloroplasts
chromoplasts
Roots:
proplastids
amyloplasts
V. Chloroplast Genetics
1. Inheritance is typically uniparental, usually maternal.
Multiple mechanisms involved, not well understood:
- in Chlamydomonas (next slide), the paternal (-) cpDNA
is destroyed, and the maternal (+) cpDNA is
preferentially replicated
- in some land plants, the paternal plastids are excluded
during fertilization or absent from the sperm cell
2. Essentially all plastids have DNA, usually the same DNA
throughout the organism (homoplasmy).
3. The DNA sequence does not change during differentiation.
There are exceptions to the last 2 statements.
Chlamydomonas life cycle has sexual and asexual reproduction.
For sex, there are 2 mating types, mt+ and mt-,
mt- cpDNA
destroyed
Digestion of cpDNA
of the mt- parent in a
young zygote of
Chlamydomonas
revealed by
fluorescence staining
of DNA.
0 Minutes
10 minutes
Nishimura, Yoshiki et al. (1999) Proc. Natl. Acad. Sci. USA 96, 12577-12582
From Mauseth, 1998
Acetabularia – green alga, fossils known,
many species likely extinct
nucleus
A single giant cell (5 cm), 1 nucleus, ~1
x 106 chloroplasts
~30 % of chloroplasts don’t have DNA!
Chloroplast DNA (cpDNA)
General features:
1. double-stranded, circular molecule
2. no histones, but have bound proteins (e.g.,
Hu), organized into nucleoids
3. G-C content typically less than nuclear DNA
4. multiple copies (~30-100) per plastid (i.e., all
cp genes are multi-copy)
5. can be 10-20% of the total DNA in leaves
“relaxed”
cpDNA
molecule
from
lettuce
From Kolodner & Tewari