Non-alcoholic fatty liver disease (NAFLD) is a major liver disease subtype worldwide, is commonly associated with insulin resistance and obesity. NAFLD is characterized by an excessive hepatic lipid accumulation, as well as hepatic steatosis. Fenofibrate is a peroxisome proliferator-activated receptor alpha agonist widely used in clinical therapy to effectively ameliorate the development of NAFLD, but its mechanism of action is incompletely understood. Here, we found that fenofibrate dramatically modulate the gut microbiota composition of high-fat diet (HFD)-induced NAFLD mouse model, and the change of gut microbiota composition is dependent on TFEB-autophagy axis. Furthermore, we also found that fenofibrate improved hepatic steatosis, and increased the activation of TFEB, which severed as a regulator of autophagy, thus, the protective effects of fenofibrate against NAFLD are depended on TFEB-autophagy axis. Our study demonstrates the host gene may influence the gut microbiota and highlights the role of TFEB and autophagy in the protective effect of NAFLD. This work expands our understanding of the regulatory interactions between the host and gut microbiota and provides novel strategies for alleviating obesity.