Decoding immunoglobulin repertoires in zebrafish reveals distinct paradigms for adaptive immune diversification in...

The evolutionary origins of adaptive immune diversification remain a central question in immunology. Using zebrafish as an early vertebrate model, we systematically dissect the distinct diversification strategies of ancient IgM and IgZ antibody repertoires. While both isotypes share a common V gene pool, they exhibit lineage-specific V/J usage biases during independent V(D)J recombination. Heavy-chain rearrangement employs the recombination-activating gene-non-homologous end joining (RAG-NHEJ) pathway, with IgM demonstrating greater junctional diversity than IgZ. Antigen-driven somatic hypermutation (SHM) primarily targets IgM via activation-induced cytidine deaminase (AID)-primed base excision repair/mismatch repair within primitive germinal center-like clusters. However, unbiased targeting across complementarity determining and framework regions, together with constrained CXCR5-CXCL13 signaling, limits mutation frequency and affinity maturation. Notably, we identify an AID-independent SHM pathway mediated by APOBEC2a that specifically targets TCG/CGT motifs. Furthermore, light-chain V-J recombination is uniquely processed via microhomology-mediated end joining, restricting CDR3 diversity compared with heavy-chain. Conversely, IgZ⁺ B cells are biased toward plasma cell differentiation, exhibit loosely organized distribution, and undergo minimal antigen-driven SHM, aligning with rapid mucosal defense. Our findings reveal both conserved and distinct diversification mechanisms in teleosts, illuminating layered evolutionary strategies balancing receptor diversity and self-tolerance in early vertebrates.