Locus-specific Convergent Evolution and Interchromosomal Rearrangements Contribute Type I Interferon Diversification in Amniotes

Type I interferons (IFN-Is) play essential roles in antiviral immune responses. The extensive diversification of IFN-Is into multiple subtypes and redundant gene copies has posed significant challenges for evolutionary reconstruction. To address this, we developed an integrated analytical pipeline combining the IFN-SCOPE classification model and GENE-GRADE algorithm. Through comprehensive synteny-guided phylogenetic analysis, we identified three evolutionarily conserved IFN-I loci (HACD4, MOB3B, and UBAP2) that maintain chromosomal colocalization across all major amniote lineages - mammals, squamates, and Archelosauria (turtles, crocodilians, and birds). While MOB3B-locus maintains a single IFN-κ ortholog, the HACD4 (IFN-H) and UBAP2 (IFN-U) loci show lineage-specific expansion patterns: IFN-H proliferated in mammals/reptiles but remained single-copy in birds, whereas IFN-U expanded in birds but not in other lineages. Phylogenetic analysis reveals these independently evolved multicopy genes nevertheless cluster into two conserved subgroups (IFN-H2/H1 and IFN-U2/U1), suggesting convergent functional specialization. Within IFN-H and IFN-U clusters, the single-copy IFN-H2 and IFN-U2 genes - positioned at the ancestral ends of their respective genomic arrays - likely represent the progenitor sequences of each locus. Notably, among mammalian IFN-H2 genes, we identified the poorly characterized IFN-υ (rather than IFN-β) as the ancestral form of mammalian IFN-H subtypes. Furthermore, we identified IFN-I genes at non-canonical loci resulting from interchromosomal duplication events in tortoises and diving ducks and provide clear evidence that interchromosomal duplications contributed to IFN-I gene diversity. These discoveries advance our understanding of the evolutionary mechanisms shaping intronless IFN-I genes in amniotes, and potentially beneficial for developing novel IFN-I based antiviral treatments through comparative immunological approaches.