The Rana sylvatica skin-secreted antimicrobial peptide (AMP) gene repertoire highlights broader patterns in anuran AMP evolution

Antimicrobial peptides (AMPs) are a diverse class of short peptides with therapeutic potential as antimicrobial agents and immune regulators. Many anurans (frogs and toads) produce diverse sets of AMPs via specialized skin glands, yet their biological significance remains debated. To better understand the evolution and expression of anuran AMPs, we examined the Wood Frog (Rana sylvatica) genome, assembled a skin transcriptome to identify skin-secreted AMP genes, and confirmed the presence of predicted AMPs in skin secretions using mass spectrometry. Along with discovering nine novel AMPs, we found that the AMP genes of R. sylvatica cluster in a manner similar to other AMP gene families and that analogous gene clusters appear in other Rana species and more distant relatives. These genes possess a consistent structure, and all encode a conserved signal peptide which is subject to strong purifying selection, contrasting with the varied associated AMPs. We examined the effects of season and pathogen exposure on AMP gene expression, observing large differences in transcript and peptide levels between genes. Overall, our findings provide insight into broad patterns of AMP evolution in anurans, reinforcing the importance of the signal peptide as a functional feature of skin-secreted peptide precursors which requires further study.