OBJECTIVE-Obesity and associated pathologies are major global health problems. Transforming growth factor-beta/Smad3 signaling has been implicated in various metabolic processes, including adipogenesis, insulin expression, and pancreatic beta-cell function. However, the systemic effects of Smad3 deficiency on adiposity and insulin resistance in vivo remain elusive. This study investigated the effects of Smad3 deficiency on whole-body glucose and lipid homeostasis and its contribution to the development of obesity and type 2 diabetes.RESEARCH DESIGN AND METHODS-We compared various metabolic profiles of Smad3-knockout and wild-type mice. We also determined the mechanism by which Smad3 deficiency affects the expression of genes involved in adipogenesis and metabolism. Mice were then challenged with a high-fat diet to study the impact of Smad3 deficiency on the development of obesity and insulin resistance.RESULTS-Smad3-knockout mice exhibited diminished adiposity with improved glucose tolerance and insulin sensitivity. Chromatin immunoprecipitation assay revealed that Smad3 deficiency increased CCAAT/enhancer-binding protein beta-C/EBP homologous protein 10 interaction and exerted a differential regulation on proliferator-activated receptor beta/delta and proliferator-activated receptor gamma expression in adipocytes. Focused gene expression profiling revealed an altered expression of genes involved in adipogenesis, lipid accumulation, and fatty acid beta-oxidation, indicative of altered adipose physiology. Despite reduced physical activity with no modification in food intake, these mutant mice were resistant to obesity and insulin resistance induced by a high-fat diet.CONCLUSIONS-Smad3 is a multifaceted regulator in adipose physiology and the pathogenesis of obesity and type 2 diabetes, suggesting that Smad3 may be a potential target for the treatment of obesity and its associated disorders.