Targeting ferroptosis shows considerable promise for diseases through modulation of immune cell function and phenotype. However, the process is regulated by diverse factors making its role in heavy metal immunotoxicity incompletely understood. In this study, we investigated the effect of Cdon ferroptosis in B cells and underlying mechanism. We demonstrated that Cdinduces ferroptosis via iron overload, as supported by rescue with the iron chelator deferoxamine (DFO) and elevated Felevels detected via Ferro Orange flow cytometry. Cdtreatment also increased lipid peroxidation and expression of long-chain acyl-CoA synthetase 4 (ACSL4), while downregulating glutathione peroxidase 4 (GPX4). Functionally, the ferroptosis inducer Erastin pre-sensitized cells to Cd, while the specific inhibitor Ferrostatin-1 robustly restored viability. Mechanistically, Cdenhanced protein interaction between nuclear receptor coactivator 4 (NCOA4) and ferritin heavy chain 1 (FTH1). By siRNA knockdown of NCOA4, Cd-induced FTH1 degradation, iron overload, and lipid peroxidation were significantly attenuated. Autophagy/lysosome inhibitors 3-methyladenine (3-MA) and chloroquine (CQ) partially reversed Cd-mediated suppression of GPX4 and FTH1. Using LysoTracker and acridine orange staining, we found that Cdenhances lysosomal acidification. To further delineate the mechanism of Cd-induced ferritinophagy, we employed the transcriptomic analysis of spleen from Cd-exposed mice. In vivo analysis revealed a marked upregulation of ATP6V0A1, a crucial element of the vacuolar proton pump. This ATP6V0A1 upregulation was recapitulated in human Ramos B cells following Cdexposure at both the protein and transcriptional levels. Knockdown of ATP6V0A1 mitigated Cd-induced lysosomal acidification, FTH1 degradation, iron overload, and lipid peroxidation. These findings, combined murine in vivo screening and in vitro validation in human cells, indicate that Cdupregulates ATP6V0A1 to promote lysosomal acidification, which facilitates NCOA4-mediated ferritinophagy, iron release, and subsequent ferroptosis. This study advances our understanding of heavy metal immunotoxicity and highlights potential therapeutic targets for mitigating Cd-induced immune dysfunction. 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+
