Structural and Mechanistic Analyses of TSC1/2 and Rheb 1/2-Mediated Regulation of the mTORC Pathway

Abstract

Abstract : The multicomponent kinase mTORC1 (mammalian target of rapamycin complex 1) regulates cell growth by coordinating upstream signals from growth factors, intracellular energy levels, and amino acid availability and is deregulated in diseases such as cancer and diabetes. The TSC1 and TSC2 proteins form a tumor suppressor complex that transmits growth factor and energy signals to mTORC1 by regulating the GTP-loading state of Rheb, a Ras-related GTP-binding protein. Additionally, the Rag GTPases interact with mTORC1 and are proposed to activate it in response to amino acids by promoting mTORC1 translocation to a membrane-bound compartment that contains the mTORC1 activator, Rheb. In our recent study of the amino acid sensitivity, we showed that amino acids induce the movement of mTORC1 to lysosomal membranes, where the Rag proteins reside. A complex encoded by the MAPKSP1, ROBLD3, and c11orf59 genes, which we term Ragulator, interacts with the Rag GTPases, recruits them to lysosomes, and is essential for mTORC1 activation. Constitutive targeting of mTORC1 to the lysosomal surface is sufficient to render the mTORC1 pathway amino acid insensitive and independent of Rag and Ragulator, but not Rheb, function. Thus, Rag-Ragulator-mediated translocation of mTORC1 to lysosomal membranes is the key event in amino acid signaling to mTORC1. In a separate study, we determined the three-dimensional (3D) structure of the fully assembled human mTORC1 by cryo-electron microscopy (cryo-EM). Our analyses reveal that mTORC1 is an obligate dimer with an overall rhomboid shape and a central cavity. The dimeric interfaces are formed by interlocking interactions between the mTOR and raptor subunits. Extended incubation with FKBP12-rapamycin compromises the structural integrity of mTORC1 in a stepwise manner, leading us to propose a model in which rapamycin inhibits mTORC1-mediated phosphorylation of 4E-BP1 and S6K1 through different mechanisms.