► AS160 Thr649→Ala knockin mice are made to study roles of AS160/14-3-3 interaction ► The knockin mice display impaired glucose tolerance and reduced insulin sensitivity ► Deregulated GLUT4 trafficking and glucose uptake in muscles underlie this phenotype ► These findings shed new light on understanding of the pathology of type II diabetes. This evidence concerns the gene TBC1D4 and type 2 diabetes mellitus.