Author: Renata Pirani

Distribution of sampling localities for individuals included in our breeding experiment. A Male slender anole dewlaps come in two morphs: solid and bicolor (photographs taken by John David Curlis). B Sampling localities in the Canal Zone of central Panama and associated morph frequencies of males in those populations (circle size corresponds to relative sample size from each site). The frequency of solid morph individuals generally declined from the Pacific to the Caribbean versant of Panama. C Sampling localities for individuals included in our Poolseq experiment. All these individuals were collected along a central trail that bisects Soberanía National Park, near the town of Gamboa.

Color us intrigued! In the world of Anolis lizards, the dewlap—a colorful throat fan used for communication—is one of the most iconic features. It’s been flaunted, flashed, and filmed in countless studies of behavior and adaptive radiation. But for all the attention it gets, we still know surprisingly little about how this flashy ornament is inherited at the genetic level.

Enter the slender anole (Anolis apletophallus), a small but charismatic lizard that ups the dewlap drama with a striking polymorphism. Males come in two distinct throat flavors: the “solid” morph, sporting a fully orange dewlap, and the “bicolor” morph, featuring a mostly white dewlap with a splash of orange at the base. What’s behind this variation?

To find out, we set up 99 crosses (yes, 99!) using individuals from populations that were either monomorphic (fixed for one morph) or polymorphic (home to both morphs). The results? A classic Mendelian plot twist. The dewlap polymorphism in this species is likely controlled by a single autosomal locus, with the solid orange morph dominant over the bicolor version. Simpler than we expected—but the story doesn’t end there.

Pooled sequencing revealed a strong candidate locus (single-minded 1, SIM1) that may underly the slender anole dewlap polymorphism. A Manhattan plot illustrating genomic differentiation between the dewlap morphs using pairwise FST values, and B Fisher exact tests. The red lines in B and C represent the Bonferroni correction (p value= 8.9), with points above the line indicating significantly differentiated SNPs across the slender anole genome. C One locus on scaffold 3, the transcription factor SIM1, contained a peak of many highly differentiated SNPs between morphs. The vertical green bars in the inset represent exons and the arrow represents the direction of genome annotation. D Nucleotide diversity (π) of SIM1 for the bicolor (green) and solid (brown) dewlap morphs. Note that nucleotide diversity was often higher in the dominant solid morph. E SNP panel showing segregating alleles in all significant SNPs (n = 175) in the candidate region of SIM1 for each morph plotted as a heatmap. We denote ref/ref in green representing that solid morph individuals were fixed for one allele and alt/alt in purple indicates that bicolor individuals were fixed for an alternative allele. Ref/alt in blue represents more than two alleles segregating at that position for either the solid or bicolor population. F Male slender anole dewlaps come in two morphs: solid and bicolor.

We also dove into the genome using pooled sequencing (Pool-seq) to search for regions associated with the trait. Our outlier analysis spotlighted a single genomic region with a strong signal—and within it, a prime suspect emerged: single-minded 1 (SIM1), a transcription factor with a flair for phenotypic control. Could this be the mastermind behind dewlap color in slender anoles?

The plot thickens, and we’re excited to keep unraveling the genetic threads behind one of the most iconic signals in lizard evolution. Stay tuned!

Link to the paper just published in Heredity.

We recently published a chromosome-scale assembly of the slender anole (Anolis apletophallus) genome, a species that has been studied for decades at the Smithsonian Tropical Research Institute in Panama.

Here is the abstract: The slender anole, Anolis apletophallus, is a small arboreal lizard of the rainforest understory of central and eastern Panama. This species has been the subject of numerous ecological and evolutionary studies over the past 60 years as a result of attributes that make it especially amenable to field and laboratory science. Slender anoles are highly abundant, short-lived (nearly 100% annual turnover), easy to manipulate in both the lab and field, and are ubiquitous in the forests surrounding the Smithsonian Tropical Research Institute in Panama, where researchers have access to high-quality laboratory facilities. Here, we present a high-quality genome for the slender anole, which is an important new resource for studying this model species. We assembled and annotated the slender anole genome by combining three technologies; Oxford Nanopore, 10X Genomics linked-reads, and Dovetail Omni-C. We compared this genome with the recently published brown anole (Anolis sagrei) and the canonical green anole (Anolis carolinensis) genomes. Our genome is the first assembled for an Anolis lizard from mainland Central or South America, the regions that host the majority of diversity in the genus. This new reference genome is one of the most complete genomes of any anole assembled to date and should facilitate deeper studies of slender anole evolution, as well as broader scale comparative genomic studies of both mainland and island species. In turn, such studies will further our understanding of the well-known adaptive radiation of Anolis lizards.

And here is a slightly longer summary of what we did (and some results): We used a hybrid genome assembly by combining three technologies: Oxford Nanopore, 10X Genomics linked-reads, and Dovetail Omni-C. We annotated our slender anole genome using the Dovetail Genomics annotation pipeline and compared our genome with the recently published brown anole (Anolis sagrei) and the canonical green anole (Anolis carolinensis) genomes. We also estimated the repeat elements composition and repetitive landscape using the RepeatModeler and RepeatMasker pipelines.

After several rounds of improvement, our final genome assembly for the slender anole was ~2.4 Gbp in size with with a scaffold N50 of 154.6 Kbp and a GC content of 43.8%. The slender anole genome was thus substantially larger than both the green anole (1.89 Gbp) and brown anole (1.93 Gbp) genomes. Our annotation using the Dovetail pipeline identified a total of 46,763,836 bp coding regions and a total of 33,912 gene models. The number of gene models identified for the slender anole was higher than that of both the green anole (22,292) and brown anole (20,033).

Authors: Renata M. Pirani^1,2*†, Carlos F. Arias^2,3, Kristin Charles¹, Albert K. Chung^2,4, John David Curlis^2,5, Daniel J. Nicholson^2,6, Marta Vargas², Christian L. Cox^2,7, W. Owen McMillan², Michael L. Logan^1,2

Affiliation:

(1) Department of Biology and program in Ecology, Evolution, and Conservation Biology, University of Nevada, Reno, Reno, 89557, United States

(2) Smithsonian Tropical Research Institute, Panama City, Panama

(3) Data Science Lab, Office of the Chief Information Officer, Smithsonian Institution, Washington, 20013, United States

(4) Department of Ecology and Evolutionary Biology, Princeton University, Princeton, 08544-2016, United States

(5) Department of Ecology and Evolution, University of Michigan, Ann Arbor, 48109-1085, United States

(6) University of Texas, Arlington, Arlington, 76019, United States

(7) Florida International University, Miami, 33199, United States

*Corresponding author: renatampirani@gmail.com

† Present address: Department of Ecology and Evolutionary Biology, University of California, Los Angeles, 90095, USAFigure 1 Figure_2

This year at SICB, I had the great opportunity to talk about part of my work as a postdoctoral researcher in the lab of Dr. Michael Logan at the University of Nevada, Reno. In collaboration with John David Curlis (University of Michigan), Christian Cox (Florida International University), W. Owen McMillan (Smithsonian Tropical Research Institute), and Carlos Arias (STRI), we have been studying the Panamanian slender anole Anolis apletophallus, which has a dewlap polymorphism: males either have a solid orange dewlap (solid morph) or a white dewlap with an orange spot (bicolor morph). Preliminary results from John David Curlis’ PhD dissertation research suggests that, in our mainland study population, the frequencies of these morphs change in conjunction with understory light levels—the solid morph is more frequently observed in brighter areas where more light reaches the understory, whereas  the opposite is true for the bicolor dewlap, which is more frequently observed in darker areas of the forest. Thus, it seems possible that selection is maintaining this polymorphism following the predictions of the sensory drive hypothesis, which states that sexual signals should have characteristics that make them the most transmissible given the physical characteristics of the local habitat.

As part of an effort to understand how this trait is evolving in the wild, I set out to understand the genetic basis of this dewlap polymorphism. To do this, my collaborators and I first assembled the full slender anole genome which we then used as a reference for a pooled population sequencing (Pool-Seq) approach using half individuals with solid dewlaps and half individuals with bicolor dewlaps to identify the genomic region underlying this dewlap polymorphism.

Our genome assembly showed pretty good results (Scaffold N50 154,613,287). The Pool-Seq results presented a clear peak of differentiation between solid and bicolor morph groups that corresponded to a region on Scaffold 3. We have a promising candidate gene within this region that may underly the dewlap polymorphism, but will continue to explore these data further to understand the genetic basis of this charismatic trait.