Type 2 diabetes mellitus (T2DM) is a progressive disease， accompanied by increased insulin resistance and deteriorating β-cell function. Previous studies have revealed that microRNA (miRNA) plays a crucial role in the treatment of T2DM. Hence， we aim to investigate the role of microRNA-125b-5p (miR-125b-5p) in pancreatic β-cell function and insulin sensitivity of mice with T2DM with the involvement of Dishevelled antagonist Dapper1 (DACT1) and the c-Jun NH2-terminal kinases (JNK) signaling pathway. Firstly， a mouse model of T2DM was established by administering a high-fat diet plus low dosage of streptozotocin， and function of pancreatic β-cell and insulin sensitivity in the normal and T2DM mice were detected. Then， the pancreatic β-cells were collected from pancreatic islet tissues and treated with different mimics， inhibitors and siRNAs. After that， the relationship among miR-125b-5p， DACT1， and the JNK signaling-related factors in T2DM mice was determined. Finally， cell proliferation and apoptosis were determined. Mice with T2DM had lower pancreatic β-cell function and insulin sensitivity， as well as diminished expression of miR-125b-5p but enhanced expressions of DACT1， JNK and c-Jun. miR-125b-5p inhibited DACT1 expression and the activation of the JNK signaling pathway， as well as restrained cell proliferation and promoted cell apoptosis. The current results suggest that up-regulated miR-125b-5p promotes insulin sensitivity and enhances pancreatic β-cell function through inhibiting the JNK signaling pathway by negatively mediating DACT1.