March6 Protects Against Acute Kidney Injury by Suppressing Renal Tubular Epithelial Cell Ferroptosis Through the Destabilization of P53 and ACSL4 Proteins

Acute kidney injury (AKI) progression involves significant contributions from renal tubular epithelial cell ferroptosis. Membrane associated RING finger protein 6 (March6) is implicated in modulating ferroptosis by regulating the stability of related proteins, yet its specific role in tubular epithelial cell ferroptosis and AKI progression remains unknown. This work seeks to clarify the regulatory role of March6 in the ferroptosis of tubular epithelial cells and its involvement in AKI using ischemia-reperfusion injury (IRI)-induced animal models and hypoxia/reoxygenation (H/R)-induced cellular models. Our results demonstrated a significant reduction in March6 expression in renal tissues from IRI mice and H/R cells, accompanied by elevated PTGS2 and reduced GPX4 levels, two ferroptosis marker proteins. Overexpression of March6 in HK-2 cells significantly counteracted Erastin-induced ferroptosis, whereas silencing March6 increased susceptibility. In H/R models, March6 overexpression enhanced cell viability and lowered cell death, reversing ferroptosis-related changes, whereas silencing March6 exhibited the opposite effects. Tubular-specific overexpression of March6 in mice with IRI-induced AKI notably mitigated kidney damage and suppressed ferroptotic changes. Mechanistically, March6 inhibited ferroptosis by accelerating the degradation of key pro-ferroptotic proteins ACSL4 and p53. Co-immunoprecipitation (Co-IP) experiments validated the direct interaction between March6 and p53 or ACSL4. Overexpressing ACSL4 or p53 in March6-overexpressing HK-2 cells markedly reversed March6's protective effects against H/R-induced damage and ferroptosis. Collectively, March6 mitigates ferroptosis in renal tubular epithelial cells by promoting the degradation of ACSL4 and p53, thereby alleviating AKI progression. This study not only uncovers a novel regulatory mechanism of tubular epithelial cell ferroptosis in AKI but also provides a potential therapeutic target for its treatment.