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MTOR-dependent regulation of PHLPP expression controls the rapamycin sensitivity in cancer cells.

ABSTRACT: PHLPP belongs to a novel family of protein phosphatases that serve as negative regulators of Akt. There are two isoforms, PHLPP1 and PHLPP2, identified in this family. Our previous studies indicated a tumor suppressor role of both PHLPP isoforms in colon cancer. Here we report that the expression of PHLPP is controlled by mTOR-dependent protein translation in colon and breast cancer cells. Treating cells with rapamycin or knockdown of mTOR using RNAi results in a marked decrease of PHLPP protein expression. In contrast, stable knockdown of TSC2, a negative regulator of mTOR activity, increases PHLPP expression. The rapamycin-mediated down-regulation of PHLPP is blocked by expression of a rapamycin-insensitive mutant of p70S6K. In addition, depletion of 4E-BP1 expression by RNAi results in an increase of PHLPP expression and resistance to rapamycin-induced down-regulation. Moreover, inhibition of mTOR activity by amino acid or glucose starvation reduces PHLPP expression in cells. Functionally, we show that rapamycin-mediated inhibition of PHLPP expression contributes to rapamycin resistance in colon cancer cells. Thus, our studies identify a compensatory feedback regulation in which the activation of Akt is inhibited by up-regulation of PHLPP through mTOR, and this mTOR-dependent expression of PHLPP subsequently determines the rapamycin sensitivity of cancer cells.

SUBMITTER: Liu J

PROVIDER: S-EPMC3057781 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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mTOR-dependent regulation of PHLPP expression controls the rapamycin sensitivity in cancer cells.

Liu Jianyu J Stevens Payton D PD Gao Tianyan T

The Journal of biological chemistry 20101222 8

PHLPP belongs to a novel family of protein phosphatases that serve as negative regulators of Akt. There are two isoforms, PHLPP1 and PHLPP2, identified in this family. Our previous studies indicated a tumor suppressor role of both PHLPP isoforms in colon cancer. Here we report that the expression of PHLPP is controlled by mTOR-dependent protein translation in colon and breast cancer cells. Treating cells with rapamycin or knockdown of mTOR using RNAi results in a marked decrease of PHLPP protein ...[more]

PMID: 21177869

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Project description:ObjectivesPatients with mammalian target of rapamycin (mTOR)-dependent malformations of cortical development (MCDs) associated with seizures display hyperperfusion and increased vessel density of the dysmorphic cortical tissue. Some studies have suggested that the vascular defect occurred independently of seizures. Here, we further examined whether hypervascularization occurs in animal models of global and focal MCD with and without seizures, and whether it is sensitive to the mTOR blocker, rapamycin, that is approved for epilepsy treatment in tuberous sclerosis complex.MethodsWe used two experimental models of mTOR-dependent MCD consisting of conditional transgenic mice containing Tsc1null cells in the forebrain generating a global malformation associated with seizures and of wild-type mice containing a focal malformation in the somatosensory cortex generated by in utero electroporation (IUE) that does not lead to seizures. Alterations in blood vessels and the effects of a 2-week-long rapamycin treatment on these phenotypes were assessed in juvenile mice.ResultsBlood vessels in both the focal and global MCDs of postnatal day 14 mice displayed significant increase in vessel density, branching index, total vessel length, and decreased tissue lacunarity. In addition, rapamycin treatment (0.5 mg/kg, every 2 days) partially rescued vessel abnormalities in the focal MCD model, but it did not ameliorate the vessel abnormalities in the global MCD model that required higher rapamycin dosage for a partial rescue.SignificanceHere, we identified hypervascularization in mTOR-dependent MCD in the absence of seizures in young mice, suggesting that increased angiogenesis occurs during development in parallel to alterations in corticogenesis. In addition, a predictive functional outcome is that dysplastic neurons forming MCD will have better access to oxygen and metabolic supplies via their closer proximity to blood vessels. Finally, the difference in rapamycin sensitivity between a focal and global MCD suggest that rapamycin treatment will need to be titrated to match the type of MCD.

Project description:BackgroundThe AKT/mammalian target of rapamycin (mTOR) signaling pathway is regulated by 17α-estradiol (E2) signaling and mediates E2-induced proliferation and progesterone receptor (PgR) expression in breast cancer.Methods and resultsHere we use deep sequencing analysis of previously published data from The Cancer Genome Atlas to demonstrate that expression of a key component of mTOR signaling, rapamycin-insensitive companion of mTOR (Rictor), positively correlated with an estrogen receptor-α positive (ERα+) breast tumor signature. Through increased microRNA-155 (miR-155) expression in the ERα+ breast cancer cells we demonstrate repression of Rictor enhanced activation of mTOR complex 1 (mTORC1) signaling with both qPCR and western blot. miR-155-mediated mTOR signaling resulted in deregulated ERα signaling both in cultured cells in vitro and in xenografts in vivo in addition to repressed PgR expression and activity. Furthermore we observed that miR-155 enhanced mTORC1 signaling (observed through western blot for increased phosphorylation on mTOR S2448) and induced inhibition of mTORC2 signaling (evident through repressed Rictor and tuberous sclerosis 1 (TSC1) gene expression). mTORC1 induced deregulation of E2 signaling was confirmed using qPCR and the mTORC1-specific inhibitor RAD001. Co-treatment of MCF7 breast cancer cells stably overexpressing miR-155 with RAD001 and E2 restored E2-induced PgR gene expression. RAD001 treatment of SCID/CB17 mice inhibited E2-induced tumorigenesis of the MCF7 miR-155 overexpressing cell line. Finally we demonstrated a strong positive correlation between Rictor and PgR expression and a negative correlation with Raptor expression in Luminal B breast cancer samples, a breast cancer histological subtype known for having an altered ERα-signaling pathway.ConclusionsmiRNA mediated alterations in mTOR and ERα signaling establishes a new mechanism for altered estrogen responses independent of growth factor stimulation.

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MTOR-dependent regulation of PHLPP expression controls the rapamycin sensitivity in cancer cells.

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mTOR-dependent regulation of PHLPP expression controls the rapamycin sensitivity in cancer cells.

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