RAG1-Independent Atypical B Cell Development in Chickens: Redefining the Role of RAG1 in Avian Immunology
The importance of RAG1 in avian B cell development has been a topic of significant debate, as chickens exhibit limited antibody diversity through RAG1-mediated V(D)J recombination compared to mammals. In chickens, antibody diversity is predominantly achieved through gene conversion in the bursa of Fabricius. However, the absence of RAG1 disrupts gene conversion and impairs B cell signaling pathways, underscoring its importance. In this study, we utilized a RAG1-deficient chicken model to uncover a classical RAG1-dependent and an alternative RAG1-independent B cell developmental pathway unique to avians. Unlike mammals, where RAG1-induced double-strand breaks (DSBs) activate ATM-dependent DNA damage responses (DDR) for B cell development, we found that ATR mediates this process in chickens. In RAG1-deficient chicken bursa, normal development of B cells is halted due to the absence of V(D)J recombination and reduced ATR-mediated B cell development signals. Although the absence of RAG1 significantly reduced B cell numbers, a small number of B cell subset was still observed. These atypical B cells (ABCs), which developed in the bursa, exhibited reduced ATR-mediated signaling but elevated expression of inflammatory and migration-related genes, allowing them to migrate into the bursa. ABCs were also specifically observed in peripheral organ (spleen) of RAG1-deficient chicken and expressed IgM with only harboring J and C segments. Additionally, ABCs displayed autoimmune-like phenotypes, including lysosome, TLR signaling activation, autophagy, and inflammation. This study highlights avian-specific alternative RAG1-independent B cell development and provides insights into autoantibody-independent mechanisms of autoimmune diseases, contributing to comparative immunology and therapeutic development. Overall design: Bulk RNAseq of embryonic day 18 bursa from wildtype and RAG1 knock out chickens, three biological replicates each