Structural and Mechanistic Analyses of TSC1/2 and Rheb 1/2-Mediated Regulation of the mTORC Pathway
Annual rept. 1 Jul 2008-30 Jun 2009
WHITEHEAD INST FOR BIOMEDICAL RESEARCH CAMBRIDGE MA
Pagination or Media Count:
The mammalian target of rapamycin complex 1 mTORC1 formed by mTOR, Raptor, mLST8 and PRAS40 is a principal regulator of cell growth in response to the nutrient and energy status of the cell, and its deregulation has recently emerged as a key driver in human cancers. Here, we present three-dimensional structures of the intact complex, as well as free Raptor, at nominal resolutions of 26 and 30 , as determined by cryo-electron microscopy cryo-EM. The molecular architecture of the holoenzyme contains a two-fold symmetry, suggesting mTORC1 as an obligate dimeric complex. It is characterized by a flat and rhomboid shape that features a central cavity and peripheral protrusions. Antibody labeling and molecular docking revealed that the smaller subunits mLST8 and PRAS40 localize to the periphery and constitute the distal feet-like structures and the central tips, respectively. In addition, in the intact complex, mTOR and Raptor form a dimeric interface that is disrupted acutely by rapamycin treatment. Dimeric particles accommodate rapamyin at a distal site that is independent of the Raptor-binding region. While binding of a single FKBP12-rapamycin complex has no observable effects on the structural integrity of mTORC1, extended incubation in vitro results in fast and complete disintegration of the complex. Consistent with this observation, loss of the structural integrity of mTORC1 manifests a differential regulation of 4E-BP1 and S6K1 phosphorylation by rapamycin both in vivo and in vitro.
- Medicine and Medical Research