The hand of evolution: conservation and divergence of hand paralogs in cyclostomes and gnathostomes

The Hand family comprises basic helix-loop-helix transcription factors with key roles in the development of the lateral mesoderm, heart, branchial arches and paired appendages in vertebrates. Although Hand1 and Hand2 have been extensively studied in model vertebrates, the evolutionary history and diversification of Hand paralogs in early-diverging vertebrate lineages remain poorly understood. Here, we combined phylogenetic reconstruction, local genomic synteny analysis, and spatiotemporal expression profiling to investigate Hand paralogs in the lamprey Lampetra fluviatilis and the cartilaginous fish Chiloscyllium griseum . Phylogenetic analyses resolve the correspondence of 3 L. fluviatilis paralogs - handα , handβ , and handγ - with their Petromyzon marinus counterparts, whereas deeper relationships between cyclostome Hand paralogs and gnathostome hand1 / hand2 remain unresolved; local genomic synteny provides complementary but insufficient evidence for one-to-one orthology across the cyclostome–gnathostome split. During lamprey embryogenesis, handα and handβ show partially overlapping but distinct spatiotemporal expression patterns, including a previously unrecognized handβ domain in the caudal fin, whereas no specific handγ signal was detected under the conditions examined. In C. griseum , hand1 and hand2 show distinct expression patterns in the branchial apparatus and developing fins: hand2 is expressed in posterior domains of paired and dorsal fins, whereas hand1 is detected predominantly in paired fins by whole-mount in situ hybridization, with substantially lower transcript abundance in dorsal fins supported by targeted qRT-PCR. Together, these findings resolve distinct cyclostome Hand paralogs and reveal lineage- and paralog-specific diversification of Hand expression while supporting an evolutionarily ancient deployment of Hand genes in the vertebrate branchial region and fin-associated developmental programs.