The product of the intronless single copy gene RSC1A1, named RS1, is an intracellular 617-amino-acid protein that is involved in the regulation of the Na(+)-d-glucose cotransporter SGLT1. We generated and characterized RS1 knockout (RS1(-/-) mice. In the small intestines of RS1(-/-) mice, the SGLT1 protein was up-regulated sevenfold compared to that of wild-type mice but was not changed in the kidneys. The up-regulation of SGLT1 was posttranscriptional. Small intestinal d-glucose uptake measured in jointly perfused small bowel and liver was increased twofold compared to that of the wild-type, with increased peak concentrations of d-glucose in the portal vein. At birth, the weights of RS1(-/-) and wild-type mice were similar. At the age of 3 months, male RS1(-/-) mice had 5% higher weights and 15% higher food intakes, whereas their energy expenditures and serum leptin concentrations were similar to those of wild-type mice. At the age of 5 months, male and female RS1(-/-) mice were obese, with 30% increased body weight, 80% increased total fat, and 30% increased serum cholesterol. At this age, serum leptin was increased, whereas food intake was the same as for wild-type mice. The data suggest that the removal of RS1 leads to leptin-independent up-regulation of food intake, which causes obesity.