Dipeptidyl peptidase IV (DP-IV), a member of the prolyl oligopeptidase family of peptidases, is involved in the metabolic inactivation of a glucose-dependent insulinotropic hormone, glucagon-like peptide 1 (GLP-1), and other incretin hormones. Here, we investigated the impact of DP-IV deficiency on body weight control and insulin sensitivity in mice. Whereas WT mice displayed accelerated weight gain and hyperinsulinemia when fed a high-fat diet (HFD), mice lacking the gene encoding DP-IV (DP-IV-/-) are refractory to the development of obesity and hyperinsulinemia. Pair-feeding and indirect calorimetry studies indicate that reduced food intake and increased energy expenditure accounted for the resistance to HFD-induced obesity in the DP-IV-/- mice. Ablation of DP-IV also is associated with elevated GLP-1 levels and improved metabolic control in these animals, resulting in improved insulin sensitivity, reduced pancreatic islet hypertrophy, and protection against streptozotocin-induced loss of beta cell mass and hyperglycemia. Together, these observations suggest that chronic deletion of DP-IV gene has significant impact on body weight control and energy homeostasis, providing validation of DP-IV inhibition as a viable therapeutic option for the treatment of metabolic disorders related to diabetes and obesity.