T Cell–Dependent Affinity Maturation and Innate Immune Pathways Differentially Drive Autoreactive B Cell Responses in Rheumatoid Arthritis

Objective Rheumatoid arthritis ( RA ) is characterized by the activation of B cells that produce anti–citrullinated protein antibodies ( ACPA s) and rheumatoid factors ( RF s), but the mechanisms by which tolerance is broken in these B cells remain incompletely understood. We undertook this study to investigate whether ACPA + and RF + B cells break tolerance through distinct molecular mechanisms. Methods We developed antigen–tetramers to isolate ACPA + and RF + B cells and performed single‐cell RNA sequencing on 2,349 B cells from 6 RA patients and 1 healthy donor to analyze their immunoglobulin repertoires and transcriptional programs. Prominent immunoglobulins were expressed as monoclonal antibodies and tested for autoantigen reactivity. Results ACPA + and RF + B cells were enriched in the peripheral blood of RA patients relative to healthy controls. Characterization of patient‐derived monoclonal antibodies confirmed ACPA and RF targeting of tetramer‐specific B cells at both antigen‐inexperienced and affinity‐matured B cell stages. ACPA + B cells used more class‐switched isotypes and exhibited more somatic hypermutations relative to RF + B cells, and these differences were accompanied by down‐regulation of CD 72 and up‐regulation of genes that promote class‐switching and T cell–dependent responses. In contrast, RF + B cells expressed transcriptional programs that stimulate rapid memory reactivation through multiple innate immune pathways. Coexpression analysis revealed that ACPA + and RF + B cell–enriched genes belong to distinct transcriptional regulatory networks. Conclusion Our findings suggest that ACPA + and RF + B cells are imprinted with distinct transcriptional programs, which suggests that these autoantibodies associated with increased inflammation in RA arise from 2 different molecular mechanisms.