5α-Reductase underscores the development of pectoral fin breeding tubercles in zebrafish

Breeding tubercles (BTs) are keratinized structures that appear on the pectoral fins of male zebrafish, but the evolutionary patterns and genetic mechanisms driving their development remain unclear. Through a survey of Danioninae species, we find BTs on pectoral fins are conserved within Rasborini and Danionini, but absent in Chedrini and outgroups. This survey also highlights pectoral-fin BTs in cypriniforms as a promising model system for investigating the evolution and development of secondary sexual traits in vertebrates. Comparative transcriptome analyses show BTs-bearing regions are enriched for genes involved in vasculature formation, keratin production, and steroid metabolism. Among these, we identify the localized enrichment of srd5a2b, one of the two zebrafish paralogs of steroid 5-α-reductase type 2, as a key candidate regulator. Experimental inhibition of steroid 5-α-reductase block BTs formation, while treatment with Dihydrotestosterone (DHT) partially restores it, indicating local androgen conversion is essential for BTs development. Cross-species comparisons confirm the conservation of this mechanism among species with pectoral fin BTs. Overall, localized androgen metabolism via the steroid 5-α-reductase pathway appears central to the development and evolution of this secondary sexual trait in zebrafish.