Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation

Kathryn G. Foster, Hugo A. Acosta-Jaquez, Yves Romeo, Bilgen Ekim, Ghada A. Soliman, Audrey Carriere, Philippe P. Roux, Bryan A. Ballif, Diance C. Fingar

Research output: Contribution to journalArticle

102 Scopus citations

Abstract

The rapamycin-sensitive mTOR complex 1 (mTORC1) promotes protein synthesis, cell growth, and cell proliferation in response to growth factors and nutritional cues. To elucidate the poorly defined mechanisms underlying mTORC1 regulation, we have studied the phosphorylation of raptor, an mTOR-interacting partner. We have identified six raptor phosphorylation sites that lie in two centrally localized clusters (cluster 1, Ser696/Thr706 and cluster 2, Ser855/Ser859/ Ser863/Ser877) using tandem mass spectrometry and generated phosphospecific antibodies for each of these sites. Here we focus primarily although not exclusively on raptor Ser863 phosphorylation. We report that insulin promotes mTORC1-associated phosphorylation of raptor Ser863 via the canonical PI3K/TSC/ Rheb pathway in a rapamycin-sensitive manner. mTORC1 activation by other stimuli (e.g. amino acids, epidermal growth factor/MAPK signaling, and cellular energy) also promote raptor Ser863 phosphorylation. Rheb overexpression increases phosphorylation on raptor Ser863 as well as on the five other identified sites (e.g. Ser859, Ser855, Ser877, Ser696, and Thr706). Strikingly, raptor Ser863 phosphorylation is absolutely required for raptor Ser859 and Ser855 phosphorylation. These data suggest that mTORC1 activation leads to raptor multisite phosphorylation and that raptor Ser863 phosphorylation functions as a master biochemical switch that modulates hierarchical raptor phosphorylation (e.g. on Ser859 and Ser855). Importantly, mTORC1 containing phosphorylation site-defective raptor exhibits reduced in vitro kinase activity toward the substrate 4EBP1, with a multisite raptor 6A mutant more strongly defective that single-site raptor S863A. Taken together, these data suggest that complex raptor phosphorylation functions as a biochemical rheostat that modulates mTORC1 signaling in accordance with environmental cues.

Original languageEnglish (US)
Pages (from-to)80-94
Number of pages15
JournalJournal of Biological Chemistry
Volume285
Issue number1
DOIs
StatePublished - Jan 1 2010

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Foster, K. G., Acosta-Jaquez, H. A., Romeo, Y., Ekim, B., Soliman, G. A., Carriere, A., Roux, P. P., Ballif, B. A., & Fingar, D. C. (2010). Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation. Journal of Biological Chemistry, 285(1), 80-94. https://doi.org/10.1074/jbc.M109.029637