T-bet regulates the maintenance and ASC differentiation potential of lymph node and lung effector memory B cell subsets

While human and mouse memory B cells (Bmem) can express T-bet, its role in regulating Bmem function is largely unknown. We characterized multiple transcriptionally distinct clusters of mature, somatically-mutated nucleoprotein (NP)-specific Bmem in LNs and lungs of influenza-infected mice. Although none of the Bmem expressed the plasma cell (PC) lineage commitment factors Blimp1, one cluster was enriched for Tbx21+ cells. Like the previously described human T-bet+ effector Bmem (eBmem) population, Tbx21+ mouse Bmem upregulated gene networks associated with effector metabolism, protein synthesis and the unfolded protein response. Using constitutive and inducible mouse models to ablate T-bet in B cells, we showed that T-bet expression by Bmem was required for persistence of LN and lung eBmem with rapid in vitro and in vivo PC differentiation potential. Thus, T-bet marked NP+ eBmem that were poised to differentiate and was required for maintenance of eBmem that respond in the lung following viral re-exposure. On two separate days, on D30 following PR8 (H1N1) influenza infection mdLN from Tbx21 reporter were isolated and pooled. Influenza specifc (NP+) and non-specific (NPneg) memory B cells were isolated by fluorescence activated cell sorting (FACS) and stained with CITE-seq antibodies (ADT and HTO). GEX, VDJ, and ADT libraries were prepared using the 10X platform.