P5CS-coupled proline metabolism manipulates metabolic dysfunction-associated steatotic liver disease.

Metabolic dysfunction-associated steatotic liver disease (MASLD), characterized by hepatic steatosis, inflammation, and fibrosis, has reached epidemic proportions globally. Emerging evidence highlights a close association between amino acid metabolic dysregulation and MASLD pathogenesis, however, the precise mechanisms remain elusive. In this study, we identify pyrroline-5-carboxylate synthase (P5CS), a pivotal enzyme in proline biosynthesis, as a critical regulator of hepatic proline production and a key driver of MASLD progression. Based on comprehensive analysis of clinical samples from MASLD patients and experimental mouse models, we demonstrate that elevated hepatic and plasma proline levels, resulting from increased P5CS expression, are strongly correlated with disease severity. Genetic overexpression of P5CS in the livers of mice exacerbates diet-induced MASLD, whereas its knockdown exhibits protective profiles. Notably, proline supplementation abolishes the beneficial effects of P5CS knockdown, confirming the causal role of proline overproduction in MASLD pathogenesis. Mechanistically, P5CS-mediated proline accumulation impairs mitochondrial function, thereby disrupting fatty acid oxidation and promoting hepatic lipid accumulation. Pharmacological inhibition of P5CS activity could restore mitochondrial capacity. Thus, our findings establish P5CS-regulated proline metabolism as a novel pathogenic mechanism of MASLD and provide a potential approach for MASLD therapy.