Therefore, these data support the existence of regulatory functions of in the FoB-MZB fate decision. Overall, the results shown above confirmed that migratory inputs are not affected by the absence of in B cells, and that failure to detect MZB in B cell-specific in B cells caused a block in B cell differentiation, which results in the absence of MZP. MZB, the mitochondrial ETC integrity in FoB, and the efficacy of antiviral humoral immunity. Subject terms: Follicular B cells, Humoral immunity, Marginal zone B cells, Antimicrobial responses Follicular and marginal zone B (FoB and MZB, respectively) cells have divergent metabolic characteristics. Here the authors show that deficiency of glutamate cysteine ligase (Gclc), the enzyme for glutathione synthesis, differentially impacts FoB and MZB homeostasis, while specifically impeding FoB activation and downstream antiviral immunity. Introduction B cells regulate many functions required for immune homeostasis and can present antigens very efficiently through their major histocompatibility complexes to T cells1,2. Furthermore, B cells can release immunomodulatory cytokines that are critical for the normal immune system maintenance, and differentiate into effector subsets that secrete polarized cytokines depending on the environment3. However, the principal function of a B lymphocyte is usually to secrete antibodies that provide humoral immunity. Antibody protection is a key component of the innate and adaptive phases of the immune response and is mediated mainly by two distinct splenic B cell subsets: marginal zone B cells (MZB) and follicular B cells (FoB)4C6. Previous studies have described developmental, phenotypic, functional and transcriptomic differences between MZB and FoB7C14. Moreover, distinct homeostatic control mechanisms regulate FoB and MZB distribution in the spleen15,16. FoB persist in the follicles in a quiescent state as they recirculate until activated by recognition of the antigen by their B cell receptors and the T cell-mediated U18666A cognate help, whereupon they proliferate and undergo germinal center (GC) reactions6,17. In contrast, MZB do not recirculate between lymphoid organs, are proximal to blood vessels, and can self-propagate18. Furthermore, MZB possess innate-like properties and are activated earlier during an immune challenge than FoB8. Nevertheless, like FoB, MZB can undergo GC reactions19,20. Redox balance is essential for maintaining cellular signaling and activation21. Glutathione (GSH) is a key intracellular antioxidant that scavenges excess reactive oxygen species (ROS)22C24, and it is an important molecule for the regulation of lymphocytes activation25C27. Moreover, early studies on HIV-1-infected subjects associated GSH deficiency with a poor lymphocytic response and impaired survival following HIV-1 infections28C30, implying a role for GSH in disease. In B cells, ROS are instrumental in regulating activation31,32, but the contribution to B cell subsets and functions of mitochondrial ROS (mtROS), which are generated principally within mitochondria, is poorly understood. It is, therefore, possible that redox thresholds in B cells are subset-specific, and that these thresholds mediate homeostatic functions that could account for the differing properties of MZB and FoB. To this end, Muri et al. have previously shown that GSH-dependent glutathione peroxidase 4 (Gpx4) activity is critical for MZB compared to FoB33. However, the precise role of the tripeptide GSH in B cell metabolism remains unknown. The mechanistic target of rapamycin (mTOR) signaling is a key modulator of anabolic and catabolic reactions34, which in turn could affect ROS balance. mTOR has emerged as a crucial control point for B cell functions35. Previous reports have shown that relatively high levels of mTOR complex 1 (mTORC1) signaling prevail in MZB36,37, but that the maturation of FoB downregulates the mTORC1/Akt pathway38. However, other groups have shown that efficient mTORC1 suppression is necessary to avoid MZB loss39. Because mTORC1 is a well-known regulator of metabolic functions, these findings imply that specific metabolic programs may underlie the unique characteristics of different B cell subsets. Here, we report that blocking GSH synthesis by ablation of causes loss of MZB. In the absence of GSH synthesis, FoB upregulate mTORC1 and reprogram their metabolism towards glycolysis, which is similar to the metabolic program of wild-type MZB. However, GSH-deficient FoB accumulate defective mitochondria and do not activate upon viral challenge. In summary, our analysis shows that GSH is crucial for the development of MZB and for the control of U18666A mitochondrial metabolic functions in FoB. Therefore, U18666A our results demonstrate a subset-specific role for GSH in controlling the redox balance underlying the metabolic properties between MZB and FoB. Results FoB and MZB exhibit distinct glutathione-based redox dependencies To dissect the redox state of MZB and FoB in relation to the main antioxidant GSH, we studied the expression of mRNA expression in resting = 5 animals examined over three independent experiments). Middle: representative blot of Gclc protein from total cell lysis of resting B6 FoB and MZB (= 3 animals examined over three U18666A independent experiments). Right: relative density of Gclc protein expression in resting B6 FoB and MZB (= 3 animals examined over three independent experiments). b Luminescence-based quantitation of intracellular Rabbit Polyclonal to Akt GSH/GSSG ratio in resting FoB and MZB isolated from B6 mice (= 3 animals examined over three independent experiments). (cCd) Representative histogram and quantitation of DCFDA (c) and MitoSOX (d) staining for intracellular.