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Integrity of mTORC2 is dependent on the rictor Gly-934 site

R Aimbetov1, C-H Chen, O Bulgakova

  • 1Department of Molecular and Cellular Oncology, University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.

Oncogene
|September 13, 2011
PubMed

Insights

A specific mutation in rictor protein disrupts mammalian target of rapamycin complex 2 (mTORC2) assembly. This affects phosphatidylinositol-3-OH kinase (PI3K)/Akt signaling, impacting cell proliferation and survival.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Growth factor signaling via phosphatidylinositol-3-OH kinase (PI3K)/Akt pathway regulates cell proliferation and survival.
  • Mammalian target of rapamycin complex 2 (mTORC2) is the key kinase regulating Akt activity at Ser-473.
  • mTORC2 is assembled from mTOR, rictor, Sin1, and mLST8.

Purpose of the Study:

  • To investigate the role of a specific rictor point mutation (G934E) in mTORC2 assembly and function in mammalian cells.
  • To determine if this mutation affects rictor's interaction with Sin1 and Protor.
  • To analyze the impact of the mutation on Akt signaling and cell proliferation.

Main Methods:

  • Site-directed mutagenesis to create the rictor G934E mutant in mammalian cells.
  • Co-immunoprecipitation assays to assess protein-protein interactions (rictor-Sin1, rictor-Protor).
  • Western blotting to detect Akt phosphorylation at Ser-473 and assess mTORC2 signaling.
  • Cell proliferation assays.

Main Results:

  • The rictor G934E mutation prevents the binding of rictor to Sin1, disrupting mTORC2 complex assembly.
  • The mutation does not affect the interaction between rictor and Protor.
  • Cells expressing the G934E mutant show reduced Akt phosphorylation at Ser-473 and decreased cell proliferation rates.
  • Substitution of Gly-934 with a charged amino acid is critical for rictor/Sin1 heterodimer formation.

Conclusions:

  • A single amino acid residue (Gly-934) in rictor is essential for its interaction with Sin1 and subsequent mTORC2 assembly.
  • Disruption of mTORC2 assembly via the rictor G934E mutation impairs Akt signaling and cell proliferation.
  • Full-length rictor is necessary for Sin1 interaction, but Gly-934 specifically controls this critical interaction.

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