Critical role of 4-1BB:4-1BBL axis in humoral immune responses against Plasmodium 2242516

Abstract Introduction Several costimulatory and coinhibitory signaling axis have been reported to modulate anti-Plasmodium immune response, albeit the precise role of 4-1BB:4-1BBL axis has yet to be investigated. Methods We used 4-1BB KO and 4-1BBL KO mice to investigate the effects of each receptor during Plasmodium infection. To temporally modulate 4-1BBL signaling, we treated WT mice with a 4-1BBL blocking antibody (TKS-1) at different timepoints. Considering the scenario of 4-1BB binding to Galectin-9 (GAL9) as previously reported, certain experiments involve treating WT mice with GAL9 blocking antibody. To understand the functional significance of 4-1BB-4-1BBL axis, we also performed single cell RNA-, V(D)J-, and CITE-seq on sort-purified WT, 4-1BB and 4-1BBL KO T and B cells sourced from three-way bone marrow chimeric mice infected with a rodent-specific Plasmodium strain. Results Here, we show that complete loss of 4-1BB resulted in delayed parasite control, impaired GC responses and memory B cell recall. The absence of 4-1BBL, however, resulted in enhanced parasite control without influencing the GC response. Early blockade of 4-1BB:4-1BBL signaling by using TKS-1 recapitulated 4-1BB KO data, an effect that was absent when 4-1BBL was blocked at effector time points. While exploring the possibility of other 4-1BB binding partners such as GAL9, we observed that combined blockade of GAL9 and 4-1BBL resulted in loss of parasite control as well as loss of protection following lethal challenge, similar to 4-1BB KO mice. Conclusion While these data show a preferential binding of 4-1BB to 4-1BBL mediating an inhibitory effect, our data suggest that 4-1BB can bind to GAL9 in the absence of 4-1BBL and potentiate stimulatory signals. Our future experiments are directed towards understanding the precise and differential role of ligands of 4-1BB and their temporal effects in governing anti-Plasmodium humoral immunity. Funding Source This work was supported by 1R01GM125907 to S.E.L., the RFUMS startup funds, Schweppe Scholar Award, 1R01AI182217 to R.V., and the AAI 2024 Careers in Immunology Fellowship to C.C. and R.V. Topic Categories Microbial, Parasitic, and Fungal Immunology (MPF)