Endoplasmic reticulum stress-mediated PPARα-DDX3X interaction promotes immune-induced liver injury

Dysfunction of endoplasmic reticulum (ER) stress is involved in the pathogenesis of immune-mediated liver injury, but its molecular mechanism has not been fully elucidated. This study explored PPARα-DDX3X interactions in regulating ER stress-related dysfunction. Liver injury was induced in C57BL/6 J mice using Con A. DDX3X hepatocyte-specific knockout (DDX3X ΔHep ) and control (DDX3X fl/fl ) mice were used. ER stress was inhibited with 4-PBA, PPARα expression modulated via siRNA, AAV-shRNA, and LV, and ER stress induced with tunicamycin in primary hepatocytes. Tunicamycin was used to induce ER stress, and AIH patients were included for validation. Hepatic DDX3X expression and phosphorylation increase while PPARα expression decreases, with DDX3X translocating to nucleus and losing colocalization with PPARα during the progression from AIH to AIH-LF. Inhibition of ER stress alleviates ConA-induced liver injury by increasing PPARα expression and reducing both DDX3X levels and phosphorylation in the liver. PPARα activation attenuates liver injury and suppresses the phosphorylation of DDX3X, which is regulated by ER stress. A phospho-deficient DDX3X mutant (p.T323A) significantly reduces ConA-induced liver injury in mice. Mechanistically, phosphorylation of DDX3X promotes its cytoplasmic–nuclear translocation and hepatocyte apoptosis during ER stress. Moreover, PPARα interacts with the DDX3X helicase domain in the cytoplasm, masking the phosphorylation site (p.T323) which inhibits its nuclear translocation leading to downregulation of CHOP expression. The PPARα-DDX3X interaction regulates DDX3X phosphorylation during ER stress, promoting DDX3X nuclear translocation and hepatocyte apoptosis. Targeting the ER stress-PPARα-DDX3X pathway offers molecular targets for immune-mediated liver injury. Under ER homeostasis, PPARα sequesters DDX3X in the cytoplasm to block its phosphorylation-driven nuclear translocation and suppress CHOP transcription, whereas ER stress-induced PPARα downregulation enables DDX3X phosphorylation and active nuclear import, leading to CHOP transactivation and subsequent hepatocyte apoptosis and liver injury.