1
Supporting Information
Appendix S1: Supplementary methods and results for degree-day model
Methods
5
We combined weather station data with a model of fly development at different temperatures to
better estimate the thermal environment experienced by Drosophila melanogaster across
latitudes (Cooper et al. 2010; Nilsson-Ortman et al. 2012). The modeling approach estimates the
number of generations as well as the mean and variance in temperature experienced by each
population through time, while accounting for the thermal dependence of development. Much
10
work on D. melanogaster indicates that we must consider the thermal dependence of
development and the propensity for reproductive diapause during winter when estimating the
thermal environment experienced by flies (Schmidt and Conde 2006; Schmidt and Paaby 2008;
Schmidt et al. 2005). Our method achieves this, in addition to confirming our simple expectation
of how mean and variance in temperature changes with latitude.
15
To model the development of flies, we first established the number of temperature degree
hours required to proceed from egg to adult using published developmental rate data from eight
populations of D. melanogaster developed at four constant temperatures (18, 22, 25 and 29°C;
Worthen 1996). Using these data, we estimate that flies require 2470 degree-hours to reach
adulthood. Because flies initiate diapause at 12°C, we set this temperature as a minimum
20
threshold below which development does not occur (Emerson et al. 2009). We downloaded daily
minimum and maximum temperatures for 2006-2011 from the weather stations nearest our
collection sites (Stations 437054, 129557, and 317994 for VT, IN, and NC, respectively)
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
2
(Williams et al. 2006). To extract predicted hourly temperatures from these data, we fit a
25
sinusoidal function to the maximum and minimum temperatures for each day (Campbell and
Norman 1998). To estimate the parameters of interest, degree hours were summed until 2470
degree-hours were reached, at which time a new generation began. We then calculated a mean
temperature for each generation, and a grand mean temperature across all generations. The
standard deviation of the grand mean provided an estimate of variance in temperature across
30
generations. While others have estimated within-generation thermal variance (Nilsson-Ortman et
al. 2012), we did not estimate this component. In contrast to seasonal variation, behavior is more
likely to buffer thermal variation within generations, complicating estimates of thermal
variability over small temporal scales.
35
Results
Our model qualitatively agrees with summary statistics of raw weather data (Table S1). As
expected, mean temperatures decrease and thermal variances increase with latitude (Table S1).
However, quantitative differences between summary statistics and estimates from our model
support the biological relevance of this approach. The model estimates of mean temperatures are
40
higher, and estimates of thermal variance are lower, than the raw mean and variance of daily
minima and maxima from 2006-2008. This stems from the model accounting for the thermal
dependence of development; flies are not active and generations are not proceeding during the
coolest and most variable winter months. Further, because VT flies experience cooler
temperatures than the other populations, fewer generations progress in VT, and periods of
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
3
45
reproductive quiescence increase relative to IN and NC flies. Together, our model estimates
support the simple predictions based on latitude alone – that selection on membrane
specialization in response to cooler mean temperature and on membrane generalization (i.e.,
plasticity) in response to thermal heterogeneity should be highest in VT.
50
Literature Cited
Campbell, J.S. & Norman, J.M. (1998) An Introduction to Environmental Biophysics. Springer,
New York.
Cooper, B.S., Czarnoleski, M. & Angilletta, M.J. (2010) Acclimation of thermal physiology in
55
natural populations of Drosophila melanogaster: a test of an optimality model. Journal of
Evolutionary Biology, 23, 2346-2355.
Emerson, K.J., Bradshaw, W.E. & Holzapfel, C.M. (2009) Complications of complexity:
integrating environmental, genetic and hormonal control of insect diapause. Trends in
Genetics, 25, 217-225.
60
Nilsson-Ortman, V., Stoks, R., De Block, M. & Johansson, F. (2012) Generalists and specialists
along a latitudinal transect: patterns of thermal adaptation in six species of damselflies.
Ecology, 93, 1340-1352.
Schmidt, P.S. & Conde, D.R. (2006) Environmental heterogeneity and the maintenance of
genetic variation for reproductive diapause in Drosophila melanogaster. Evolution, 60,
65
1602-1611.
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
4
Schmidt, P.S. & Paaby, A.B. (2008) Reproductive diapause and life-history clines in North
American populations of Drosophila melanogaster. Evolution, 62, 1204-1215.
70
Schmidt, P.S., Paaby, A.B. & Heschel, M.S. (2005) Genetic variance for diapause expression
and associated life histories in Drosophila melanogaster. Evolution, 59, 2616-2625.
Williams, C.N., Jr., Menne, M.J., Vose, R.S. & Easterling, D.R. (2006) United States Historical
Climatology Network Daily Temperature, Precipitation, and Snow Data. ORNL/CDIAC118, NDP-070. Available on-line (http://cdiac.ornl.gov/epubs/ndp/ushcn/usa.html) from
75
the Carbon Dioxide Information Analysis Center, Oak Ridge National Laboratory, U.S.
Department of Energy, Oak Ridge, Tennessee.
Worthen, W.B. (1996) Latitudinal variation in developmental time and mass in Drosophila
melanogaster. Evolution, 50, 2523-2529.
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
5
80
Table S1. Summary of weather data and the estimated mean temperature and thermal variance experienced by D. melanogaster across
latitudes accounting for thermal effects on development.
Population
East Calais, VT, USA
Latitude/
Mean temperature
Estimated mean
Thermal
Estimated thermal Estimated number
Station ID1
(°C)2
temperature (°C)3
variation (°C) 4
variation (°C)5
of generations6
44° 25' N/
7.66
16.48
12.50
6.16
55
11.55
19.51
12.12
5.87
88
15.77
21.71
11.12
5.64
123
437054
Bloomington, IN, USA
39° 59' N/
129557
Raleigh, NC, USA
35° 30' N/
317994
1
The latitude and station identification numbers of the weather stations used in our analyses.
The mean temperature from 2006-2011 calculated using daily minimum and maximum temperatures.
3
The grand mean of the estimated mean temperatures for each generation using a degree-hour model (Cooper et al. 2010).
4
The standard deviation temperatures between 2006-2011 calculated using daily minimum and maximum temperatures.
5
The standard deviation of the grand mean temperature, our estimate for thermal variation across generations, using a degree-hour
model (Cooper et al. 2010).
6
The estimated number of generations between 2006-2011 from a degree-hour model (Cooper et al. 2010). Differences represent
variation among populations in the length of reproductive quiescence, and in the number of generations during the active season.
2
85
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
6
90
Table S2. Populations do not differ in GPL plasticity in response to adult thermal shifts (see Table 2 for P-values), but the magnitude
of adult responses for all populations depends on the direction of the thermal shift. Plasticity in PE/PC is greater when adults are
shifted to 26C, relative to 16C (tpaired = 5.2139, df = 2, P = 0.035), while plasticity in the degree of lipid saturation is greater when
adults are shifted to 16C, relative to 26 C (tpaired = 11.7169, df = 2, P = 0.007).
Treatment
Adult shift to 16C
Adult Trait
∆PE/PC
∆Saturation
Adult shift to 26C
∆PE/PC
∆Saturation
Population
Mean
S.E.
East Calais, VT, USA
0.123
0.029
Bloomington, IN, USA
0.101
0.033
Raleigh, NC, USA
0.115
0.027
East Calais, VT, USA
0.012
0.003
Bloomington, IN, USA
0.015
0.003
Raleigh, NC, USA
0.016
0.004
East Calais, VT, USA
0.195
0.039
Bloomington, IN, USA
0.144
0.050
Raleigh, NC, USA
0.202
0.028
East Calais, VT, USA
0.002
0.003
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
7
Bloomington, IN, USA
0.003
0.002
Raleigh, NC, USA
0.007
0.003
95
100
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth
8
Figure S1. Developmental reaction norms for lipid saturation. All populations increase lipid
105
unsaturation (i.e., increased polyunsaturation relative to monounsaturation) when developed at
cool temperatures as predicted by models of homeoviscous adaptation, but in contrast to PE/PC
plasticity, genotypes from VT do not have increased plasticity of this trait. A) The proportion of
the membrane composed of polyunsaturated GPLs increased during development at 16C, and
thus B) the proportion of the membrane composed of monounsaturated GPLs decreased during
110
development at 16C. Data are means +/- s.e. from all genetic lines within each population.
Thermal adaptation of cellular membranes in natural populations of Drosophila melanogaster
Brandon S. Cooper, Loubna A. Hammad, Kristi L. Montooth