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Characterization of Rictor phosphorylation sites reveals direct regulation of mTOR complex 2 by S6K1

Christian C Dibble1, John M Asara, Brendan D Manning

  • 1Department of Genetics and Complex Diseases, Harvard School of Public Health, 665 Huntington Ave., SPH2-117, Boston, MA 02115, USA.

Molecular and Cellular Biology
|September 2, 2009
PubMed

Insights

Researchers identified a key phosphorylation site on Rictor (T1135) regulated by mTORC1. This finding sheds light on the signaling mechanisms controlling mTORC2 activity and Akt phosphorylation, crucial for cell growth and metabolism.

Area of Science:

  • Cellular signaling
  • Molecular biology
  • Biochemistry

Background:

  • The mammalian target of rapamycin (mTOR) pathway is critical for cell growth, proliferation, survival, and metabolism.
  • mTOR functions in two complexes, mTORC1 and mTORC2, with mTORC1 regulation well-understood but mTORC2 regulation less defined.

Purpose of the Study:

  • To elucidate the signaling mechanisms regulating mTORC2.
  • To identify and characterize novel regulatory phosphorylation sites on Rictor, a core component of mTORC2.

Main Methods:

  • Liquid chromatography-tandem mass spectrometry to identify phosphorylation sites on Rictor.
  • Site-directed mutagenesis to create Rictor phosphorylation site mutants (T1135A).
  • Cellular assays in wild-type and Rictor null cells to assess effects on Akt, SGK1, and PKC alpha phosphorylation.

Main Results:

  • Identified 21 phosphorylation sites on Rictor, including T1135, which is growth factor-responsive and rapamycin-sensitive.
  • Rictor-T1135 is phosphorylated by the mTORC1-dependent kinase S6K1.
  • Phosphorylation of Rictor-T1135 increases mTORC2-dependent Akt S473 phosphorylation and stimulates Rictor binding to 14-3-3 proteins, acting in parallel with other feedback mechanisms.

Conclusions:

  • Rictor T1135 phosphorylation by S6K1 is a novel regulatory mechanism for mTORC2.
  • This phosphorylation event influences Akt activation by insulin, independent of mTORC2 integrity or kinase activity.
  • The findings reveal a new layer of crosstalk between mTORC1 and mTORC2, impacting cellular metabolism and growth signaling.

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