Author: mjohnson216

Readers of AA are very familiar with the dramatic differences in limb length among the anole ecomorphs, but we don’t yet know which genomic regions are involved in the evolution of anole limb length.  Carlos Infante, currently a postdoc in Doug Menke’s lab at the University of Georgia, presented a talk on his work to identify enhancers (short regions of DNA where proteins bind to enhance the transcription of a gene) that are associated with anole limb development.

Carlos first described a series of previous studies that did not find differences in the proteins expressed in the limbs of different anole species, suggesting that the differences in limb length are likely controlled by differences in gene regulation. However, examining a series of enhancer regions that were identified from previous work in mice also did not reveal differences in sequence variation that were correlated with limb length.

So, Carlos and his collaborators are using tools from the field of functional genomics to address this issue, using ChIP-Seq (a method that analyzes interactions between DNA and proteins) to identify active enhancers and promotors in embryonic Anolis carolinensis tissue using antibodies against Pitx1 (a transcription factor involved in hindlimb development) and H3K27ac (an acetylated histone mark).  By comparing results from these two datasets, they could identify enhancers that are expressed in forelimbs, hindlimbs, trunk tissues, or tubercles. Their plan for future work involves using the list they’ve generated of enhancers expressed in both forelimbs and hindlimbs to identify the regulatory regions that control the development of limb morphologies among Anolis species.

Much has been written about how differences in relative limb length allow anoles to successfully occupy distinct portions of the arboreal habitat. Most research has focused on large-scale patterns of diversity, which are presumably the result of natural selection. But limb length could also be more finely tuned using an alternative process, whereby an individual’s morphology responds to environmental cues experienced throughout development. This process is known as phenotypic plasticity. We already know that anole limbs exhibit plasticity in the lab (e.g., Losos et al. 2000, 2001 – A. sagrei; Kolbe and Losos 2005 – A. carolinensis), with lizards raised on narrow perches having shorter limbs than those raised on broad perches.  But what about lizards in their natural environments; could plasticity in limb length allow lizards to become more specialized to their particular microhabitat in the wild?  This was the question that Trinity University (San Antonio) undergraduate Alisa Dill asked in summer 2010, working with fellow undergraduate Andrew Battles, regular AA contributor Thom Sanger, and me.

We studied green anole lizards in three plots in Palmetto State Park, in southeastern Texas (Figure 1).

Figure 1 from Dill et al. In press. Study plots in Palmetto State Park. The star on the small
map of Texas (bottom left) indicates the location of the park.

The three plots differed dramatically in perch availability – one was in a dense dwarf palmetto stand in a closed forest, another was trees and bushes surrounding an open field, and the third was in a light forest with many small vines and twigs. We found that female lizards in the third plot (with the narrowest perches) had significantly shorter hindlimbs than females in the other two plots, although males did not differ in limb length among the plots.  Juveniles also did not differ in limb length across the three plots, consistent with the idea that the female’s developmental experience on varying perches may influence their adult limb length.

Why this difference between the sexes?  (This difference is also consistent with results from the laboratory studies mentioned above.)  We also observed where males and females were found in their habitat and how they locomoted through it to determine if differences in behavior between the sexes could affect the way the developing lizards interact with their environment. We found that females performed fewer locomotor behaviors, spending more time on particular perches; thus, perhaps the perches had a stronger influence on female limb length than on males, who used many more perches as they move through their environment.

There is much more work to be done to further test the hypothesis of plasticity in a natural environment. After all, the previous lab experiments were performed by growing lizards on either a wooden 2×4 or a small dowel, a much simpler environment than a forest with many perch options. It would be ideal to perform a “common-garden” experiment where juvenile lizards are transplanted between environments with different perch diameters. We also don’t yet understand the developmental mechanisms that cause limb plasticity in anoles, but more information about those mechanisms will help determine why the sexes respond differently to differences in their microhabitat.

You can read more about our study in our recent publication in the Journal of Zoology, available online now in Early View.

References:

Dill, A.K., T.J. Sanger, A.C. Battles and M.A. Johnson. In press. Sexually dimorphisms in habitat-specific morphology and behavior in the green anole lizard. Journal of Zoology.

Kolbe, J.J. & Losos, J.B. (2005). Hind-limb length plasticity in Anolis carolinensis. J. Herpetol. 39, 674–678.

Losos, J.B., Creer, D.A., Glossip, D., Goellner, R., Hampton, A., Roberts, G., Haskell, N., Taylor, T. & Ettling, J. (2000). Evolutionary implications of phenotypic plasticity in the hindlimb of the lizard Anolis sagrei. Evolution 54,301–305.

Losos, J.B., Schoener, T.W., Warheit, K.I. & Creer, D. (2001). Experimental studies of adaptive differentiation in Bahamian Anolis lizards. Genetica 112-113, 399–415.

There are few topics more exciting than anole reproduction, but there’s still much we have to learn about the neuroendocrine mechanisms that allow these creatures to do the deed.  We know that sex steroid hormones facilitate reproductive behaviors across a diversity of animals, and anoles are no exception.  In particular, an enzyme called aromatase regulates both male and female sexual behaviors by synthesizing estradiol from testosterone.  In a recent study using green anoles (Anolis carolinensis), Rachel Cohen and Juli Wade of Michigan State University examined whether lizard sex and season influenced the expression of aromatase in areas of the brain that are known to influence vertebrate reproductive behavior (the preoptic area (POA), the amygdala, and the ventromedial hypothalamus (VMH)).

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