A novel mechanism of selenium deficiency driving aberrant Th17 cell differentiation via GPX3-targeted PIG3: mitophagy blockade exacerbates mtDNA release

• Selenium deficiency downregulates GPX3 and its target PIG3, triggering redox imbalance, mitochondrial dysfunction, and impaired mitochondrial autophagy flux in splenic lymphocytes. • GPX3-PIG3 axis-mediated mitophagosome-lysosome fusion defects lead to mtDNA leakage into the cytoplasm, thereby promoting abnormal Th17 cell differentiation. • Targeting the GPX3/PIG3 signaling axis to clear damaged mitochondria and prevent mtDNA leakage may offer a novel therapeutic strategy for autoimmune diseases. The biological functions of selenium (Se) are mainly realized through selenoproteins. As the only secreted antioxidant enzyme in the glutathione peroxidase family, the deficiency of glutathione peroxidase 3 (GPX3) can affect lymphocyte function. Th17 cells play an important role in immune regulation, but whether they are regulated by GPX3 remains unclear. We sought to decipher the regulatory role of GPX3 in the differentiation of Th17 cells in porcine spleen. While replicating the dietary Se deficiency porcine spleen model in vivo , we established in vitro models using cultured porcine splenic lymphocytes under conditions of Se deficiency, GPX3 knockdown, and p53-inducible gene 3 (PIG3) overexpression. Comprehensive analyses were performed utilizing a suite of techniques, including molecular docking, Co-Immunoprecipitation (Co-IP), Western blot (WB) and mitochondrial DNA (mtDNA) quantification, etc. Our study revealed an interactive relationship between GPX3 and PIG3. Se deficiency downregulated GPX3, thereby suppressing PIG3 expression, which led to intracellular redox imbalance and mitochondrial dysfunction. Concurrently, mitochondrial fusion-fission dynamics became imbalanced, mitophagic flux was obstructed, and mtDNA leaked into the cytoplasm. These alterations ultimately promoted the aberrant differentiation of Th17 cells in porcine splenic lymphocytes by modulating the expression of key factors. Overexpression of PIG3 significantly alleviated oxidative stress induced by low GPX3 expression, restored mitochondrial homeostasis, reduced mtDNA leakage, and suppressed abnormal Th17 differentiation. However, when the autophagic flux inhibitor Baf-A1 was applied under conditions of combined GPX3 knockdown and PIG3 overexpression, the protective effects of PIG3 were reversed, indicating that unimpeded mitophagic flux is essential for the GPX3-PIG3 axis to suppress Th17 differentiation. Se deficiency impedes mitophagy via the GPX3/PIG3 axis, exacerbates mtDNA leakage, and thereby drives Th17 cell differentiation. Our findings indicate that the GPX3/PIG3 signaling axis may represent a potential therapeutic target for autoimmune diseases.