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

97 Citations (Scopus)

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

Fingerprint

Raptors
Phosphorylation
Sirolimus
Cues
Chemical activation
Phospho-Specific Antibodies
Cell proliferation
Cell growth
Phosphatidylinositol 3-Kinases
Epidermal Growth Factor

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Foster, K. G., Acosta-Jaquez, H. A., Romeo, Y., Ekim, B., Soliman, G. A., Carriere, 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

Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation. / Foster, Kathryn G.; Acosta-Jaquez, Hugo A.; Romeo, Yves; Ekim, Bilgen; Soliman, Ghada A.; Carriere, Audrey; Roux, Philippe P.; Ballif, Bryan A.; Fingar, Diance C.

In: Journal of Biological Chemistry, Vol. 285, No. 1, 01.01.2010, p. 80-94.

Research output: Contribution to journalArticle

Foster, KG, Acosta-Jaquez, HA, Romeo, Y, Ekim, B, Soliman, GA, Carriere, A, Roux, PP, Ballif, BA & Fingar, DC 2010, 'Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation', Journal of Biological Chemistry, vol. 285, no. 1, pp. 80-94. https://doi.org/10.1074/jbc.M109.029637
Foster KG, Acosta-Jaquez HA, Romeo Y, Ekim B, Soliman GA, Carriere A et al. Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation. Journal of Biological Chemistry. 2010 Jan 1;285(1):80-94. https://doi.org/10.1074/jbc.M109.029637
Foster, Kathryn G. ; Acosta-Jaquez, Hugo A. ; Romeo, Yves ; Ekim, Bilgen ; Soliman, Ghada A. ; Carriere, Audrey ; Roux, Philippe P. ; Ballif, Bryan A. ; Fingar, Diance C. / Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation. In: Journal of Biological Chemistry. 2010 ; Vol. 285, No. 1. pp. 80-94.
@article{fdc3b4be0982435089b1e252bb8919ee,
title = "Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation",
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.",
author = "Foster, {Kathryn G.} and Acosta-Jaquez, {Hugo A.} and Yves Romeo and Bilgen Ekim and Soliman, {Ghada A.} and Audrey Carriere and Roux, {Philippe P.} and Ballif, {Bryan A.} and Fingar, {Diance C.}",
year = "2010",
month = "1",
day = "1",
doi = "10.1074/jbc.M109.029637",
language = "English (US)",
volume = "285",
pages = "80--94",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "1",

}

TY - JOUR

T1 - Regulation of mTOR complex 1 (mTORC1) by raptor Ser863 and multisite phosphorylation

AU - Foster, Kathryn G.

AU - Acosta-Jaquez, Hugo A.

AU - Romeo, Yves

AU - Ekim, Bilgen

AU - Soliman, Ghada A.

AU - Carriere, Audrey

AU - Roux, Philippe P.

AU - Ballif, Bryan A.

AU - Fingar, Diance C.

PY - 2010/1/1

Y1 - 2010/1/1

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=73649098283&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=73649098283&partnerID=8YFLogxK

U2 - 10.1074/jbc.M109.029637

DO - 10.1074/jbc.M109.029637

M3 - Article

C2 - 19864431

AN - SCOPUS:73649098283

VL - 285

SP - 80

EP - 94

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 1

ER -