Dataset Information

MTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR.

ABSTRACT: Mammalian target of rapamycin (mTOR) is a core component of raptor-mTOR (mTORC1) and rictor-mTOR (mTORC2) complexes that control diverse cellular processes. Both mTORC1 and mTORC2 regulate several elements downstream of type I insulin-like growth factor receptor (IGF-IR) and insulin receptor (InsR). However, it is unknown whether and how mTOR regulates IGF-IR and InsR themselves. Here we show that mTOR possesses unexpected tyrosine kinase activity and activates IGF-IR/InsR. Rapamycin induces the tyrosine phosphorylation and activation of IGF-IR/InsR, which is largely dependent on rictor and mTOR. Moreover, mTORC2 promotes ligand-induced activation of IGF-IR/InsR. IGF- and insulin-induced IGF-IR/InsR phosphorylation is significantly compromised in rictor-null cells. Insulin receptor substrate (IRS) directly interacts with SIN1 thereby recruiting mTORC2 to IGF-IR/InsR and promoting rapamycin- or ligand-induced phosphorylation of IGF-IR/InsR. mTOR exhibits tyrosine kinase activity towards the general tyrosine kinase substrate poly(Glu-Tyr) and IGF-IR/InsR. Both recombinant mTOR and immunoprecipitated mTORC2 phosphorylate IGF-IR and InsR on Tyr1131/1136 and Tyr1146/1151, respectively. These effects are independent of the intrinsic kinase activity of IGF-IR/InsR, as determined by assays on kinase-dead IGF-IR/InsR mutants. While both rictor and mTOR immunoprecitates from rictor(+/+) MCF-10A cells exhibit tyrosine kinase activity towards IGF-IR and InsR, mTOR immunoprecipitates from rictor(-/-) MCF-10A cells do not induce IGF-IR and InsR phosphorylation. Phosphorylation-deficient mutation of residue Tyr1131 in IGF-IR or Tyr1146 in InsR abrogates the activation of IGF-IR/InsR by mTOR. Finally, overexpression of rictor promotes IGF-induced cell proliferation. Our work identifies mTOR as a dual-specificity kinase and clarifies how mTORC2 promotes IGF-IR/InsR activation.

SUBMITTER: Yin Y

PROVIDER: S-EPMC4816127 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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mTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR.

Yin Yancun Y Hua Hui H Li Minjing M Liu Shu S Kong Qingbin Q Shao Ting T Wang Jiao J Luo Yuanming Y Wang Qian Q Luo Ting T Jiang Yangfu Y

Cell research 20151120 1

Mammalian target of rapamycin (mTOR) is a core component of raptor-mTOR (mTORC1) and rictor-mTOR (mTORC2) complexes that control diverse cellular processes. Both mTORC1 and mTORC2 regulate several elements downstream of type I insulin-like growth factor receptor (IGF-IR) and insulin receptor (InsR). However, it is unknown whether and how mTOR regulates IGF-IR and InsR themselves. Here we show that mTOR possesses unexpected tyrosine kinase activity and activates IGF-IR/InsR. Rapamycin induces the ...[more]

PMID: 26584640

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Project description:AimsEpidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have shown dramatic clinical benefits in advanced non-small cell lung cancer (NSCLC); however, resistance remains a serious problem in clinical practice. The present study analyzed mTOR-associated signaling-pathway differences between the EGFR TKI-sensitive and -resistant NSCLC cell lines and investigated the feasibility of targeting mTOR with specific mTOR inhibitor in EGFR TKI resistant NSCLC cells.MethodsWe selected four different types of EGFR TKI-sensitive and -resistant NSCLC cells: PC9, PC9GR, H1650 and H1975 cells as models to detect mTOR-associated signaling-pathway differences by western blot and Immunoprecipitation and evaluated the antiproliferative effect and cell cycle arrest of ku-0063794 by MTT method and flow cytometry.ResultsIn the present study, we observed that mTORC2-associated Akt ser473-FOXO1 signaling pathway in a basal state was highly activated in resistant cells. In vitro mTORC1 and mTORC2 kinase activities assays showed that EGFR TKI-resistant NSCLC cell lines had higher mTORC2 kinase activity, whereas sensitive cells had higher mTORC1 kinase activity in the basal state. The ATP-competitive mTOR inhibitor ku-0063794 showed dramatic antiproliferative effects and G1-cell cycle arrest in both sensitive and resistant cells. Ku-0063794 at the IC50 concentration effectively inhibited both mTOR and p70S6K phosphorylation levels; the latter is an mTORC1 substrate and did not upregulate Akt ser473 phosphorylation which would be induced by rapamycin and resulted in partial inhibition of FOXO1 phosphorylation. We also observed that EGFR TKI-sensitive and -resistant clinical NSCLC tumor specimens had higher total and phosphorylated p70S6K expression levels.ConclusionOur results indicate mTORC2-associated signaling-pathway was hyperactivated in EGFR TKI-resistant cells and targeting mTOR with specific mTOR inhibitors is likely a good strategy for patients with EGFR mutant NSCLC who develop EGFR TKI resistance; the potential specific roles of mTORC2 in EGFR TKI-resistant NSCLC cells were still unknown and should be further investigated.

Project description:BackgroundType I insulin-like growth factor receptor (IGF-IR) tyrosine kinase induces significant oncogenic effects. Strategies to block IGF-IR signaling are being tested in clinical trials that include patients with aggressive solid malignancies. Mantle cell lymphoma is a B-cell neoplasm with poor prognosis and a tendency to develop resistance. The expression and potential significance of IGF-IR in mantle cell lymphoma are not known.Design and methodsWe used reverse transcriptase polymerase chain reaction, quantitative real-time polymerase chain reaction, immunoprecipitation, western blotting, flow cytometry, and immunohistochemistry to analyze the expression of IGF-IR mRNA, and IGF-IR and pIGF-IR proteins in mantle cell lymphoma cell lines and patients' specimens. Selective and specific blockade of IGF-IR was achieved using picropodophyllin and short-interfering RNA, respectively. Cell viability, apoptosis, cell cycle, cellular morphology, cell proliferation, and target proteins were then analyzed.ResultsWe detected the expression of IGF-IR and pIGF-IR in mantle cell lymphoma cell lines. Notably, IGF-IR molecules/cell were markedly increased in mantle cell lymphoma cell lines compared with human B-lymphocytes. IGF-IR and pIGF-IR were also detected in 78% and 74%, respectively, of 23 primary mantle cell lymphoma specimens. Treatment of serum-deprived mantle cell lymphoma cell lines with IGF-I salvaged these cells from apoptosis. Selective inhibition of IGF-IR by picropodophyllin decreased the viability and proliferation of mantle cell lymphoma cell lines, and induced apoptosis and cell cycle arrest. Selective inhibition of IGF-IR was associated with caspase-3, caspase-8, caspase-9, and PARP cleavage, cytochrome c release, up-regulation of cyclin B1, and down-regulation of cyclin D1, pCdc2, pIRS-1, pAkt, and pJnk. Similar results were obtained by using IGF-IR short-interfering RNA. In addition, picropodophyllin decreased the viability and proliferation of primary mantle cell lymphoma cells that expressed IGF-IR.ConclusionsIGF-IR is up-regulated and frequently activated in mantle cell lymphoma. Our data suggest that IGF-IR could be a molecular target for the treatment of mantle cell lymphoma.

Dataset Information

MTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR.

Publications

mTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR.

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