mTOR complex-2 (mTORC2) controls hydrophobic motif phosphorylation and activation of serum and glucocorticoid induced protein kinase-1 (SGK1)

Juan Manuel García-Martínez , Dario R Alessi (1)

1 MRC Protein Phosphorylation Unit, Department of Biochemistry

SGK1 is a member of the AGC family of protein kinase and is activated by agonists including growth factors. SGK1 regulates diverse effects of extracellular agonists by phosphorylating regulatory proteins that control cellular process such as ion transport and growth. Like other AGC family kinases, activation of SGK1 is triggered by phosphorylation of a Thr residue within the T-loop of the kinase domain and a Ser residue lying within the C-terminal hydrophobic motif (Ser422 in SGK1). PDK1 phosphorylates the T-loop of SGK1. The identity of the hydrophobic motif kinase is unclear. Recent work has established that mammalian Target Of Rapamycin complex-1 (mTORC1) phosphorylates the hydrophobic motif of the S6K whilst mTOR Complex-2 (mTORC2) phosphorylates the hydrophobic motif of Akt. Here we demonstrate that SGK1 hydrophobic motif phosphorylation and activity is ablated in knockout fibroblasts possessing mTORC1 activity but lacking the mTORC2 subunits Rictor, Sin1 or mLST8. Furthermore, phosphorylation of the NDRG1, a physiological substrate of SGK1, was also abolished in Rictor, Sin1 or mLST8 deficient fibroblasts. mTORC2 immunoprecipitated from wild type, but not from mLST8 or Rictor knockout cells, phosphorylated SGK1 at Ser422. Consistent with mTORC1 not regulating SGK1, immunoprecipitated mTORC1 failed to phosphorylate SGK1 at Ser422, under conditions which it phosphorylated the hydrophobic motif of S6K. Moreover, rapamycin treatment of 293, MCF-7 or HeLa cells suppressed phosphorylation of S6K, without affecting SGK1 phosphorylation or activation. These findings indicate that mTORC2 but not mTORC1 plays a vital role in controlling the hydrophobic motif phosphorylation and activity of SGK1. Our findings may explain why in previous studies phosphorylation of substrates such as FOXO that could be regulated by SGK, are reduced in mTORC2 deficient cells. Our data indicate that NDRG1 phosphorylation represents an excellent biomarker for mTORC2 activity.

Mots clés: Life Sciences

Juan Manuel García-Martínez, Dario R Alessi. mTOR complex-2 (mTORC2) controls hydrophobic motif phosphorylation and activation of serum and glucocorticoid induced protein kinase-1 (SGK1). Biochemical Journal, 2008, 416 (3), pp.375-385. ⟨10.1042/BJ20081668⟩. ⟨hal-00479087⟩