γ-Secretase is a transmembrane protease complex responsible for processing multiple type I transmembrane proteins, including the amyloid precursor protein (APP) and NOTCH. Using a proteomics-based screening approach, we found that dedicator of cytokinesis protein 2 (DOCK2) interacts with the γ-secretase complex component Nicastrin (NCSTN), regulating NCSTN N45-mannosylation and the effects of γ-secretase on NOTCH receptors. Both genetically depleting DOCK2 and pharmacologically targeting NCSTN mannosylation via Kifunensine ameliorate Notch-dependent leukemia progression in vivo. Together, these results reveal a mechanism by which γ-secretase specifically recognizes its substrate and provide a direction for treating Notch-related diseases.