Eye-Transcriptome and Genome-Wide Sequencing for Scolecophidia: Implications for Inferring the Visual System of the Ancestral Snake

Abstract

Molecular genetic data have recently been incorporated in attempts to reconstruct the ecology of the ancestral snake, though this has been limited by a paucity of data for one of the two main extant snake taxa, the highly fossorial Scolecophidia. Here we present and analyze vision genes from the first eye-transcriptomic and genome-wide data for Scolecophidia, for Anilios bicolor, and A. bituberculatus, respectively. We also present immunohistochemistry data for retinal anatomy and visual opsin-gene expression in Anilios. Analyzed in the context of 19 lepidosaurian genomes and 12 eye transcriptomes, the new genome-wide and transcriptomic data provide evidence for a much more reduced visual system in Anilios than in non-scolecophidian (=alethinophidian) snakes and in lizards. In Anilios, there is no evidence of the presence of 7 of the 12 genes associated with alethinophidian photopic (cone) phototransduction. This indicates extensive gene loss and many of these candidate gene losses occur also in highly fossorial mammals with reduced vision. Although recent phylogenetic studies have found evidence for scolecophidian paraphyly, the loss in Anilios of visual genes that are present in alethinophidians implies that the ancestral snake had a better-developed visual system than is known for any extant scolecophidian.

Keywords: Squamata; gene loss; opsins; phylogeny; regressive evolution; vision.

Fig. 1.

Network (modified from Emerling and Springer 2014: fig. 1) for 34 phototransduction proteins indicating absences in squamates and fossorial mammals as determined from genomic data. Note that loss of PDE6B and GRK1 in fossorial mammals is provisional, based on negative BLAST results rather than synteny analyses (Emerling and Springer 2014). Of the 36 phototransduction genes in table 1, protein products for the cone phototransduction (visual opsins) genes sws2 and rh2 are not shown because they are absent in all snakes and mammals.

Fig. 2.

Retinal photoreceptor labelling in Anilios bituberculatus (a, c) and A. bicolor (b, d, e). (a, b) Immunolabeling for RH1 in green. The transverse sections show strong labeling of rod outer segments, indicating a high rod density. Some of the rod somata in the ONL show fainter labeling. (c, d) Immunolabeling for LWS in red, with sections showing strong labeling of a substantial population of outer segments. (e) Double labeling of a section for RH1 (e1) and LWS (e2), the merge (e3) shows colocalization of the two opsins in many photoreceptor outer segments (yellow to orange), but also clear examples of outer segments showing exclusive RH1 label (green arrowhead) or LWS label (red arrowheads). The sections are counterstained with DAPI (blue) to show the retinal layers. All sections are oblique with artificially thicker nuclear layers, (c) shows a retinal fold. Overall, outer segment preservation is poor, many outer segments are ruffled, lumped, or torn. (a–d) Maximum-intensity projections of confocal image stacks; (e) single-focus image. OS, IS, photoreceptor outer and inner segments; ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer. Scale bar in (d) applies to (a–d), scale bar in (e3) applies to (e1–e3).

Fig. 3.

Proposed pattern of visual-gene loss during lepidosaurian evolution. Losses are indicated by red transverse bars on phylogenetic branches for cone (blue), rod (black), and cone and rod (purple) phototransduction genes, and visual cycle (orange) genes. Lower scale indicates approximate inferred age of phylogenetic divergences in millions of years. Phylogenetic relationships and mean estimates of divergence age estimates from www.vertlife.org.