IL-21R signal reprogramming cooperates with CD40 and BCR signals to select and differentiate germinal center B cells

Both B cell receptor (BCR) and CD40 signaling are rewired in germinal center (GC) B cells (GCBCs) to synergistically induce c-MYC and phosphorylated S6 ribosomal protein (p-S6), markers of positive selection. How interleukin-21 (IL-21), a key T follicular helper (T_FH)-derived cytokine, affects GCBCs is unclear. Like BCR and CD40 signals, IL-21 receptor (IL-21R) plus CD40 signals also synergize to induce c-MYC and p-S6 in GCBCs. However, IL-21R plus CD40 stimulation differentially affects GCBC fate compared with BCR plus CD40 ligation-engaging unique molecular mechanisms-as revealed by bulk RNA sequencing (RNA-seq), single-cell RNA-seq, and flow cytometry of GCBCs in vitro and in vivo. Whereas both signal pairs induced BLIMP1 in some GCBCs, only the IL-21R/CD40 combination induced IRF4^hi/CD138⁺ cells, indicative of plasma cell differentiation, along with CCR6⁺/CD38⁺ memory B cell precursors. These findings reveal a second positive selection pathway in GCBCs, document rewired IL-21R signaling in GCBCs, and link specific T_FH- and Ag-derived signals to GCBC differentiation.