Insights into DEPTOR regulation from in silico analysis of DEPTOR complexes.
DEPTOR is an inhibitor of the mTOR kinase which controls cell growth. DEPTOR consists of two DEP domains and a PDZ domain connected by an unstructured linker, and its stability is tightly regulated through post-translational modifications of its linker region that contains the 286SSGYFS291 degron. Based on the mTORC1 complex, our modelling suggests a possible spatial arrangement of DEPTOR which is characterised to form a dimer. Our model shows that DEPTOR's two PDZ domains bind separately to the dimeric mTOR's FAT domains ∼13 Å apart, while the extended linkers are sufficiently long to span from the FAT domain to the kinase domain and beyond to join a shared dimer of the DEP domains. This places S299 closest to the kinase's catalytic site, indicating that phosphorylation would start with it and successively upstream towards the degron. The CK1α kinase is responsible for phosphorylation of DEPTOR's degron, and our docking analysis further reveals that CK1α contains sites to bind DEPTOR's pS286, pS287 and pT295, which may act as priming phosphates for the phosphorylation of S291 of DEPTOR's degron. DEPTOR's linker can also be ubiquitylated by the UbcH5A-SCFβ-TrCP complex without its PDZ dissociating from mTOR according to the modelling. As the catalytic cleft of mTOR's kinase is restricted, interactions between the kinase's unstructured segment surrounding the cleft and DEPTOR's linker, which may involve S293 and S299, may be critical in controlling DEPTOR's access to the catalytic cleft and hence its phosphorylation by mTOR in a manner dependent on mTOR's activation.