STEPWISE DIFFERENTIATION OF IL21-PRODUCING FOLLICULAR HELPER T CELLS REVEALS DISTINCT DEVELOPMENTAL AND FUNCTIONAL STATES

The interaction of follicular helper T (Tfh) cells with B cells within the germinal center of secondary lymphoid organs is essential for the formation of high affinity antibodies, a key effector mechanism of adaptive immunity. Interleukin (IL)-21 is a proinflammatory cytokine produced by Tfh cells, however the signals that license IL-21 production by these cells during their differentiation are unknown. Here we use fate mapping and reporter strategies to show that, after initial differentiation, Tfh cells transition to an IL-21-expressing Tfh21 stage, which is extrinsically regulated by follicular regulatory T (Tfr) cells. Using single cell RNA sequencing, we uncover additional developmental stages, including an earlyTfh21 stage prior to full Tfh21 differentiation, as well as an exTfh21 stage after Tfh21 development marked by loss of IL-21. The transcriptional and epigenetic landscape is formed in the earlyTfh21 stage and is maintained throughout the exTfh21 state, suggesting terminal differentiation. We also clarify that the transcription factor Foxp1 intrinsically regulates the transition between these stages in Tfh cells. Through selective in vivo deletion of IL21-experienced Tfh cells, we show that these subsets control antigen-specific germinal center formation, maintenance and somatic hypermutation at distinct stages of the germinal center response to a SARS-CoV-2 protein vaccine. Together, these studies demonstrate the transitionary stages of Tfh development and how control of progression through these stages regulates germinal center responses. Regulation and fine-tuning of Tfh developmental stages could be leveraged to promote protective immunity or to restrain pathogenic antibody responses.