Background and Purpose Squalene epoxidase (SQLE) is a key enzyme involved in cholesterol biosynthesis, but increasing evidence reveals that SQLE is abnormally expressed in some types of malignant tumors, and the underlying mechanism remains poorly understood. Experimental Approach Bioinformatics analysis and RNA sequencing were applied to detect to differentially expressed genes in clinical HCC tumors. AnnexinV/PI, EdU assay, transwell, western blot, qRT-PCR, IHC staining, RNA sequencing, dual-luciferase reporters and HE staining were evaluated to investigate the pharmacological effects and possible mechanisms of SQLE in vitro and in vivo. Key Results We found that SQLE expression is specifically elevated in HCC tumors, correlating with poor clinical outcomes. SQLE promoted HCC growth, EMT, and metastasis both in vitro and in vivo. In contrast, silencing of SQLE expression prevented HCC development. Both RNA-seq and functional experiments revealed that the protumorigenic effect of SQLE on HCC is closely related to the activation of cellular TGF-β/SMAD signaling, but interestingly, the stimulatory effect of SQLE on TGF-β/SMAD signaling and HCC development is also critically dependent on STRAP. SQLE expression is well correlated with STRAP in HCC, and further, to amplify TGF-β/SMAD signaling, SQLE even transcriptionally increased STRAP expression mediated by the trans-acting factor AP-2α. Finally, as a chemical inhibitor of SQLE, NB-598 markedly inhibited HCC cell growth and tumor development in mouse models. Conclusions and Implications Taken together, SQLE serves as a novel oncogene in HCC development by activating TGF-β/SMAD signaling, and targeting SQLE could be useful in drug development and therapy for HCC.