Physiological and genetic mechanisms underlying population divergence across
an altitudinal gradient
Proposed Research
Jonathan Warner Atwell
Timothy James Grieves
Dawn Michelle O’Neal
Introduction
• Understanding mechanisms that underlie adaptive divergence is a central goal of evolutionary biology and ecology
• Investment in reproduction vs. survival is a ubiquitous life-history trade-off, and optimal strategies should vary with environment
• Few studies have robustly characterized physiological and genetic substrates of divergent life-history strategies
Local adaptation to seasonal environments
From Bronson 1990
•Breeding season length varies with latitude
• Life-history traits also vary with breeding season & latitude, including testosterone and immune function
•Similar differences are seen across an altitudinal gradient
• How has selection shaped the regulatory physiology of divergent mouse populations across an altitudinal gradient?
• Breeding phenology• Seasonal testosterone & immune function
• What are the genetic mechanisms underlying physiological variation?
• Divergent candidate genes
General Questions
Rocky Mountain Biological Laboratory
Alpine tundra
prairie grasslands
Subalpine forest
(Storz et al. 2004)
Field Capture methods
•Capture in baited sherman traps•Sample February-September
•Morphological measures•Assessment of reproductive condition (fondling)•Retro-orbital blood sample•Ear-tag and release
Breeding season
0
5
10
15
20
25
30
35
jan
jan fe
b
march
march
april
may
may
june ju
ly juy
aug
sept
spet oc
tnov
nov
dec
0
5
10
15
20
25
30
35
jan
jan
feb
march
march
april
may
may
june ju
ly juy
augsept
spet oc
tnovnov
dec
0
5
10
15
20
25
30
35
jan
jan
feb
march
march
april
may
may
june ju
ly juy
augsept
spet oc
tnov
nov
dec
Alpine tundra
Subalpine forest
Prairie grasslands
Measuring seasonal T and Innate Immune Function profiles…
-Capture free-living individuals each month before, during, & after breeding in both populations
-Collect sub-orbital blood samples
-Centrifuge blood, freeze plasma
-Conduct EIA assays to measure [ T ]
-Bacterial Killing Assays (Petri Dishes)
-Heamolytic Complement (EIA Plate)
= anti-testosteroneantibody
Y
Y Y
YY
Y
Y
Y
Y
Y
YY Y Y YYY Y YY YY
Y Y YY YY
= hormone in serum sample
= labeled hormoneof known amount
Y Y YY
ColorColor
Y Y
How do you measure hormones?
• Hypothesis: seasonal variation in T expression
Mountain MicePrairie Mice
T TTT
Immune Function
• Innate– Nonspecific antigen defense mechanisms – Bactericidal assay
• Measure of susceptibility
– Hemolytic complement• Measure of ability to respond
• Acquired/adaptive– Developed in response to specific antigens
• KLH– Antibody production
Alpine tundra
Subalpineforest
Prairie grassland
Hemolytic Complement Activity by Altitude
CH50
Alti
tud
e
0 0.2 0.4 0.6 0.8 1 1.2 1.4 1.6
Prairie Grasslands
Subalpine Forest
Alpine Tundra
anti
-KL
H I
gG
(%
pla
te p
osi
tive
) Alpine Tundra
Prairie Grasslands
0
20
40
60
80
Acquired Immune Response (KLH)
Common garden study…
-Differences in behavior, hormones, immunity could be genetic or plastic
-Collect 20 males and females from each population from early in life (or pregnant females).
-Establish in common captive rearing conditions
-Collect same measures across season, as in free-living studies
-We expect to document genetic change underlying phenotypic divergence.
Microarray study to identify important loci...
-Construct cDNA library
-Sacrifice individuals during peak breeding from each free-living population
-Compare gene expression profiles
-Bayesian statistics to identify genes that differ
-Use molecular database to evaluate roles of genes that differ between populations
Conclusions• Utilizing a wide array of methods and techniques, we will
be able to assess the mechanisms underlying divergence in breeding phenology, hormonal, and immunological traits.
Broader Impacts:
-Train a diverse group of undergraduates, high-schoolers, and senior “citizen scientists,” in field and lab methods.
-Identify crucial physiological and genetic mechanisms that influence the ability of populations to persist in differing environments (e.g. global climate change, habitat destruction, disease epidemiological patterns)
-Dissemination of our results will include public outreach programs (e.g. “A Moment of Science,” Discovery Channel for Kids, National Geographic, Fox News’s “Nutty Professors”).