Dithiolethiones, a novel class of adenosine monophosphate‐activated protein kinase (AMPK) activators, prevent insulin resistance through AMPK‐dependent p70 ribosomal S6 kinase‐1 (S6K1) inhibition. There is no known effect of S6K1 for liver X receptor‐alpha (LXRα)‐mediated lipogenic gene expression and steatosis, a cause of chronic liver disease. This study investigated the role of S6K1 in LXRα activation and the effects of oltipraz (prototype) and other dithiolethiones on LXRα‐dependent lipogenesis in hepatocytes and high‐fat diet animal model. Oltipraz prevented the ability of LXRα agonist (T0901317) to activate sterol regulatory element binding protein‐1c (SREBP‐1c), inhibiting its own mRNA and protein induction. Impaired SREBP‐1c activity by oltipraz caused inhibition of LXRα‐induced transcription of the fatty acid synthase, LXRα, acetyl‐CoA carboxylase, stearoyl‐CoA desaturase‐1, and adenosine triphosphate‐binding cassette transporter A1 genes. S6K1 activation antagonized the inhibitory effect of oltipraz on SREBP‐1c activation, whereas dominant negative (DN) mutant S6K1 and rapamycin inhibited the T0901317‐induced SREBP‐1c expression. Oltipraz impaired LXRα DNA binding activity and LXR agonist‐induced CYP7A1‐LXRE‐luciferase (CYP7A1) transactivation. Moreover, in vitro S6K1 directly phosphorylated LXRα at serine residues for gene transactivation, which was antagonized by its DN mutant. S6K1 inhibition antagonized CYP7A1 induction promoted by AMPK inhibition, whereas AMPK activation abrogated S6K1‐dependent CYP7A1 induction, supporting the opposing role of S6K1 and AMPK in LXR activity. Finally, oltipraz was found to inhibit hepatic triglyceride accumulation and lipogenic gene induction in mice fed a high‐fat diet. Other dithiolethiones also inhibited SREBP‐1c induction by T0901317. Conclusion: Our findings showing the role of AMPK‐S6K1 pathway in LXR activity and S6K1‐dependent inhibition of LXRα‐induced lipogenic gene transactivation by a novel class of dithiolethiones led to the identification of S6K1 as a particularly attractive target for intervention in hepatic steatosis. (HEPATOLOGY 2009.)