Mix and ChREBP form a heterodimer to regulate glucose-mediated gene expression in the liver. This study was performed to determine if the metabolic syndrome might be improved using dominant negative Mlx (dnMlx). An adenovirus bearing dnMlx was constructed and used to test the inhibitory effect of dnMlx on lipogenesis both in vitro and in vivo. Adenoviral overexpression of dnMlx in rat hepatocytes inhibited expression of glucose-regulated genes, including Chrebp and Transketolase, which constitute a positive feedback loop in the regulation of Chrebp gene expression. Adenoviral overexpression of dnMlx in 25-week-old male C57BL/6J mice reduced hepatic triglyceride contents and improved glucose intolerance by inhibiting expression of Glucose-G-phosphatase and Elov16 mRNA in addition to lipogenic enzymes. In conclusion, overexpression of dnMlx improves glucose intolerance by inhibiting expression not only of lipogenic enzymes but also other important genes such as Glucose-6-phosphatose and Elovl6. (C) 2008 Elsevier Inc. All rights reserved.