Type 1 diabetes (T1D) is an autoimmune disease resulting from the destruction of pancreatic-β cells by the immune system involving innate and adaptive immune cells. Mucosal-associated invariant T (MAIT) cells are innate-like T-cells recognizing bacterial riboflavin-precursor derivatives presented by the MHC-I related molecule, MR1. Since T1D is associated with gut microbiota modification, we investigated MAIT cells in this pathology. In T1D patients and non-obese (NOD) diabetic mice, we detected MAIT cell alterations, including increased granzyme B production, which occur before disease onset. Analysis of NOD mice deficient for MR1, and therefore lacking MAIT cells, revealed a loss of gut integrity and increased anti-islet responses associated with exacerbated diabetes. Altogether our data highlight the role of MAIT cells in the maintenance of gut integrity and the control of anti-islet autoimmune responses. MAIT cell monitoring could represent a new