Genome‐wide association studies identified GLIS3 as a susceptibility locus for type 1 and type 2 diabetes. Global Glis3 deficiency in mice leads to congenital diabetes and neonatal lethality. In this study, we explore the role of Glis3 in adulthood using Glis3^+/− and conditional knockout animals. We challenged Glis3^+/− mice with high fat diet for 20 weeks and found that they developed diabetes because of impaired beta cell mass expansion. GLIS3 controls beta cell proliferation in response to high‐fat feeding at least partly by regulating Ccnd2 transcription. To determine if sustained Glis3 expression is essential to normal beta cell function, we generated Glis3^fl/fl/Pdx1Cre^ERT+ animal by intercrossing Glis3^fl/fl mice with Pdx1Cre^ERT+ mice and used tamoxifen (TAM) to induce Glis3 deletion in adults. Adult Glis3^fl/fl/Pdx1Cre^ERT+ mice are euglycaemic. TAM‐mediated beta cell‐specific inactivation of Glis3 in adult mice downregulates insulin expression, leading to hyperglycaemia and subsequently enhanced beta cell apoptosis. We conclude that normal Glis3 expression is required for pancreatic beta cell function and mass maintenance during adulthood, which impairment leads to diabetes in adults.