IDDF2024-ABS-0074 Myeloid NCOA4 sequesters KEAP1 to reduce ferroptosis for protection against salmonellosis in mice

<h3>Background</h3> Salmonellosis, caused by <i>Salmonella enterica</i> serovar Typhimurium, is a significant global threat. Host immunity limits bacterial replication by inducing hepcidin, which degrades ferroportin, reducing iron transfer. However, this boosts macrophage iron storage, aiding intracellular pathogens like <i>Salmonella</i>. Mice lacking ferritin heavy chain (FTH1) in myeloid cells suffer worsened <i>Salmonella</i> infection. Myeloid Nuclear receptor co-activator 4 (NCOA4) regulates iron release via FTH1 autophagic degradation during low iron, but its role in salmonellosis is unclear. <h3>Methods</h3> We generated transgenic mice with myeloid cell-specific loss-of-function (LysM-Cre <i>Ncoa4</i>^F/F, KO) and gain-of-function (LysM-Cre <i>NCOA4</i>^{Loxp-Stop-Loxp (LSL)/LSL}, OE) of NCOA4. These mice were treated with <i>Salmonella</i> to induce colitis. The effects of altered NCOA4 expression on disease progression were assessed using real-time qPCR, immunoblot, and ImageStream assays. Co-immunoprecipitation was employed to examine protein interactions and molecular pathways. <h3>Results</h3> Here, we reveal that myeloid NCOA4 deficiency augments spleen iron levels and increases cellular iron accumulation, oxidative stress, and ferroptosis in bone marrow-derived macrophages (BMDM) cells. This deficiency also increases susceptibility to Salmonella-induced colitis in mice due to exacerbated oxidative stress and ferroptosis. Mechanistically, NCOA4 suppresses oxidative stress by directly binding to the E3 ubiquitin ligase Kelch-like ECH-associated protein 1 (KEAP1) and stabilizing the antioxidant transcription factor nuclear factor-erythroid 2-related factor 2 (NRF2). Activation of NRF2 protects myeloid NCOA4 knockout mice from Salmonella-induced colitis. Antioxidants Tempol and myeloid cell-targeted curcumin offer protection against colitis in myeloid NCOA4-deficient mice. A low iron diet and ferroptosis inhibition also mitigate the heightened colitis in these mice. Overexpression of myeloid cell-specific NCOA4 confers protection against Salmonella-induced colitis via upregulating NRF2 signaling. <h3>Conclusions</h3> Together, this study not only advances our understanding of the NCOA4/KEAP1/NRF2/ferroptosis axis but also paves the way for novel myeloid cell-targeted therapies to combat salmonellosis.