Structural Basis of DEPTOR to Recognize Phosphatidic Acid Using its Tandem DEP Domains

[Display omitted] •Structure of the tandem DEP domains of DEPTOR was determined at 1.5-Å resolution.•Several positively charged patches on the surface of the structure are shown.•The extensional DDEX motif of DEPTOR-DEP2 interacts with DEPTOR-DEP1.•DEPTOR binds directly to phosphatidic acid with positively charged patches.•The interaction of DEPTOR and PA provides valuable insights into mTOR activation. DEP domain containing mTOR-interacting protein (DEPTOR) plays pivotal roles in regulating metabolism, growth, autophagy and apoptosis by functions as an endogenous inhibitor of mTOR signaling pathway. Activated by phosphatidic acid, a second messenger in mTOR signaling, DEPTOR dissociates from mTORC1 complex with unknown mechanism. Here, we present a 1.5 Å resolution crystal structure, which shows that the N-terminal two tandem DEP domains of hDEPTOR fold into a dumbbell-shaped structure, protruding the characteristic β-hairpin arms of DEP domains on each side. An 18 amino acids DDEX motif at the end of DEP2 interacts with DEP1 and stabilizes the structure. Biochemical studies showed that the tandem DEP domains directly interact with phosphatidic acid using two distinct positively charged patches. These results provide insights into mTOR activation upon phosphatidic acid stimulation.