Targeted Sequencing of Venom Genes from Cone Snail Genomes Improves Understanding of Conotoxin Molecular Evolution

Abstract

To expand our capacity to discover venom sequences from the genomes of venomous organisms, we applied targeted sequencing techniques to selectively recover venom gene superfamilies and nontoxin loci from the genomes of 32 cone snail species (family, Conidae), a diverse group of marine gastropods that capture their prey using a cocktail of neurotoxic peptides (conotoxins). We were able to successfully recover conotoxin gene superfamilies across all species with high confidence (> 100Â coverage) and used these data to provide new insights into conotoxin evolution. First, we found that conotoxin gene superfamilies are composed of one to six exons and are typically short in length (mean ¼ $85 bp). Second, we expanded our understanding of the following genetic features of conotoxin evolution: 1) positive selection, where exons coding the mature toxin region were often three times more divergent than their adjacent noncoding regions, 2) expression regulation, with comparisons to transcriptome data showing that cone snails only express a fraction of the genes available in their genome (24-63%), and 3) extensive gene turnover, where Conidae species varied from 120 to 859 conotoxin gene copies. Finally, using comparative phylogenetic methods, we found that while diet specificity did not predict patterns of conotoxin evolution, dietary breadth was positively correlated with total conotoxin gene diversity. Overall, the targeted sequencing technique demonstrated here has the potential to radically increase the pace at which venom gene families are sequenced and studied, reshaping our ability to understand the impact of genetic changes on ecologically relevant phenotypes and subsequent diversification.

Conidae phylogenomics and evolution

“…The same relationships were recovered by Puillandre et al (), although these authors consider Miliariconus and Africonus as synonyms of Virroconus and Lautoconus , respectively. The recent study by Phuong and Mahardika () based on multiple exons recovered the reciprocal monophylies of Miliariconus and Virroconus , supporting the validity of both genera. The paraphyly of Lautoconus , which is recovered in all our trees, was already reported in a specific phylogenetic study of West African and Mediterranean cones based on mt genomes and a large taxon sampling of endemic species of the region (Abalde, Tenorio, Afonso, Uribe, et al, ).…”

“…Understanding the relative role of different evolutionary processes leading to the extraordinary morphological, ecological and species diversity of cone snails requires a robust phylogeny, which thus far has been elusive (Aman et al, ; Phuong & Mahardika, ; Puillandre et al, ; Uribe, Puillandre, et al, ). Several reasons make particularly challenging the reconstruction of the phylogeny of cone snails, including the difficulty of obtaining thorough taxon samplings and the need of gathering large sequence data sets able to accumulate the phylogenetic signal needed to resolve the typical short nodes associated to evolutionary radiations.…”

“…Here, we propose an intermediate approach similar to that of Phuong and Mahardika () based on the analysis of concatenated exons (>4,000; >500,000 bp) from 32 cone species, but complementary and now possible thanks to the ongoing active sequencing of cone venom gland transcriptomes. We gathered up to 41 different species representing 27 genera of Conidae in the mt genome data set and 22 different species belonging to 19 genera of Conidae in the nuclear and combined data sets.…”

“…Therefore, the challenge is going beyond phylogenies of cones based on few genes (Aman et al, ; Puillandre et al, ) and reconstruct robust phylogenetic relationships among cone genera sensu Tucker and Tenorio () based on multilocus sequences data sets such as the recent one based on concatenating hundreds to thousands of exon sequences (Phuong & Mahardika, ). An additional source of molecular markers for phylogenetic inference is mitochondrial (mt) genomes, which have been widely used in gastropods and proven to be particularly useful for resolving phylogenies at the family level (Osca, Templado, & Zardoya, ; Uribe, Williams, Templado, Abalde, & Zardoya, ) as well as for disentangling recent radiation events (Abalde, Tenorio, Afonso, & Zardoya, ; Abalde, Tenorio, Afonso, Uribe, et al, ).…”

Lack of signal for the impact of venom gene diversity on speciation rates in cone snails

“…2015; Chang & Duda 2016; Phuong et al . 2016; Phuong & Mahardika 2018). Future work in other venomous animal systems may shed light on whether or not the ability to adapt to different prey through venom evolution translates to the long-term evolutionary success of taxa.…”