Systemic 4-1BB stimulation augments extrafollicular memory B cell formation and recall responses during Plasmodium infection [scATAC-seq]

T-dependent germinal center (GC) output, comprising plasma cells (PC) and memory B cells (MBC), is crucial for the clearance of Plasmodium infection and protection against reinfection. In this study, we examined the effect of an agonistic antibody targeting 4-1BB (CD137), a member of the Tumor Necrosis Factor Receptor Superfamily (TNFRSF), during experimental malaria. We found that exogenous 4-1BB stimulation dramatically enhanced humoral immune memory and protection from reinfection, despite delaying the effector GC response. Although fewer in number, single cell RNA and ATAC sequencing of MBCs from mice that received 4-1BB stimulation revealed clusters with a transcriptional and epigenetic signature indicative of superior recall and proliferative potential. Importantly, our results indicate that these effects are independent of parasite load or the inflammatory milieu but are dependent on IL-9R signaling in B cells. Our study proposes an immunomodulatory approach to enhance the quality of the MBC pool, providing superior protection during infection and vaccination, particularly in the context of malaria. C57BL/6 mice were infected with Plasmodium yoelii and treated with 3H3 (4-1BB agonist antibody) or isotype control on days 5 and 7 post infection. On 38 days post infection, the spleens were harvested and MSP-1 specific (B220+IgD-GL7-CD38+Decoy-MSP1+) and polyclonal (B220+IgD-GL7-CD38+) memory B cells were flow sorted. Nuclei were isolated and transposition, barcoding on the 10X Chromium (10X Genomics) and library construction were performed according to the manufacturer’s protocol for Chromium Next GEM Single Cell ATAC Reagents Kits v2 (10X Genomics).