TM4SF5-mediated KEAP1 Regulation in Hepatocytes Irrelevant to NRF2 Expression and Activity Promotes Oxidative Stress and Inflammation to Develop Metabolic Dysfunction-Associated Steatotic Liver Disease.

Metabolic dysfunction-associated steatohepatitis (MASH)-associated fibrosis involves inflammation accompanied by reactive oxygen species (ROS), in addition to abnormal lipid metabolism and extracellular matrix (ECM) deposition. ROS levels are regulated by the NRF2-KEAP1 pathway. Transmembrane 4 L six family member 5 (TM4SF5) is implicated in metabolic dysfunction-associated steatotic liver disease (MASLD). However, it remains unknown how hepatocyte TM4SF5 modulates abnormal lipid and ROS accumulations during MASLD development. Here we assessed the influence of TM4SF5 on NRF2-KEAP1 pathway utilizing various in vitro and in vivo MASLD models. Our results indicate that hepatocyte TM4SF5 downregulates KEAP1 in physiological states and stabilizes KEAP1 in pathological conditions, without altering NRF2 expression. However, TM4SF5-dependent stabilization of KEAP1 was not observed in Tm4sf5 (-/-) KO mice. At least the cytosolic TM4SF5 C-terminus could bind to KEAP1 for proteosomal degradation. TM4SF5-driven biphasic KEAP1 regulation was associated with increased CD36 levels in normal livers, whereas in hyperlipidemic states, it contributed to oxidative stress and hepatic inflammation. Genetically engineered mice with altered Tm4sf5 and Nrf2 displayed TM4SF5-induced MASLD phenotypes characterized by elevated Keap1, regardless of Nrf2 expression or activity. These findings were more obvious than for mice with Nrf2 mutation alone. Notably, suppression of Keap1 alone nullified the MASLD-promoting effects of TM4SF5. Taken together, these data demonstrate that TM4SF5 can modulate KEAP1 independently of NRF2, identifying TM4SF5-mediated KEAP1 stabilization as a potential therapeutic target for MASLD.