Transcriptomic Analysis Reveals Potential Candidate Pathways and Genes Involved in Toxin Biosynthesis in True Toads

Synthesized chemical defenses have broadly evolved across countless taxa and are important in shaping evolutionary and ecological interactions within ecosystems. However, the underlying genomic mechanisms by which these organisms synthesize and utilize their toxins are relatively unknown. Herein, we use comparative transcriptomics to uncover potential toxin synthesizing genes and pathways, as well as interspecific patterns of toxin synthesizing genes across 10 species of North American true toads (Bufonidae). Upon assembly and annotation of the 10 transcriptomes, we explored patterns of relative gene expression and possible protein-protein interactions across the species to determine what genes and/or pathways may be responsible for toxin synthesis. We also tested our transcriptome dataset for signatures of positive selection to reveal how selection may be acting upon potential toxin producing genes. We assembled high-quality transcriptomes of the bufonid parotoid gland, a tissue not often investigated in other bufonid-related RNAseq studies. We found several genes involved in metabolic and biosynthetic pathways (e.g., steroid biosynthesis, terpenoid backbone biosynthesis, isoquinoline biosynthesis, and glucosinolate biosynthesis) that were functionally enriched and/or relatively expressed across the 10 focal species that may be involved in the synthesis of alkaloid and steroid toxins, as well as other small metabolic compounds that cause distastefulness in bufonids. We hope that our study lays a foundation for future studies to explore the genomic underpinnings and specific pathways of toxin synthesis in toads, as well as at the macroevolutionary scale across numerous taxa that produce their own defensive toxins.