Scoparone-Induced Phase Separation of ZDHHC12 Drives Ferroptosis via ATG12 S-palmitoylation and Ferritinophagy in Liver Fibrosis.

Activated hepatic stellate cells (HSCs) are central effectors of liver fibrogenesis, and their selective elimination is critical for fibrosis regression. Although ferroptotic clearance of HSCs has emerged as a promising antifibrotic strategy, the upstream mechanisms coupling metabolic remodeling to ferroptosis susceptibility remain unclear. This study investigated whether scoparone (SCO) suppresses liver fibrosis through zinc finger DHHC-type palmitoyltransferase 12 (ZDHHC12) condensate formation, leading to enhanced autophagy-related protein 12 (ATG12) S-palmitoylation, induction of autophagy, and increased ferroptosis susceptibility in HSCs. We found that SCO restrained cell proliferation, migration, and fibrogenic activation while inducing iron overload, lipid peroxidation, glutathione depletion, and ferroptosis-like mitochondrial remodeling. Metabolomic and S-palmitoylation analyses linked these effects to increased palmitic acid levels and enhanced global protein S-palmitoylation. ZDHHC12 emerged as a SCO-responsive palmitoyltransferase associated with autophagy activation, ferritin turnover, ferroptosis induction, and repression of fibrogenic markers. Mechanistically, SCO promoted the formation of liquid-like ZDHHC12 condensates, which enhanced its interaction with ATG12 and facilitated ATG12 S-palmitoylation at the conserved Cys134 residue. This modification was associated with increased ATG12 stability through reduced ubiquitin-dependent proteasomal degradation, thereby promoting autophagy activation and increasing ferroptosis susceptibility in HSCs. Mutation of ATG12 at Cys134 attenuated these effects and diminished HSC ferroptosis. In cholestatic and toxic models of liver fibrosis, SCO alleviated liver injury and collagen deposition, whereas HSC-targeted silencing of ZDHHC12 or ATG12 attenuated these protective effects, supporting biological consistency in vivo. Integrated bioinformatic and clinical cohort analyses suggested that ZDHHC12 expression positively correlated with ATG12 expression and was associated with fibrosis progression in patients with liver fibrosis. Collectively, these findings support a ZDHHC12-dependent ATG12 S-palmitoylation pathway linking lipid metabolic remodeling to autophagy-mediated ferroptosis in HSCs and suggest phase-regulated S-palmitoylation as a potential antifibrotic target.