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MTOR inhibition by rapamycin increases ceramide synthesis by promoting transforming growth factor-?1/Smad signaling in the skin.

ABSTRACT: Although mammalian target of rapamycin (mTOR) mediates a wide variety of biological functions, little information is available on the effect of mTOR on the functions of skin cells. In this study, we investigated effects of mTOR inhibition by rapamycin on ceramide synthesis in the skin of rats and human keratinocytes and its regulatory mechanisms. The phosphorylation of p70 S6 kinase, which indicates mTOR activation, was induced in the skin of rats fed a high-fat diet, but this abnormality was reversed by supplementation with rapamycin. Ceramide levels and the mRNA levels of serine palmitoyltransferase (SPT) and transforming growth factor (TGF)-?1 were suppressed in the skin of rats fed high-fat diets, but this abnormality was reversed by supplementation with rapamycin. TGF-?1-induced SPT mRNA expression was blocked by SB525334, an inhibitor of TGF-?1-induced Smad2/3 nuclear localization, in human keratinocytes. Rapamycin-induced SPT mRNA expression was blocked by an anti-TGF-?1 antibody or SB525334 in human keratinocytes. These results show that mTOR inhibition by rapamycin increases ceramide synthesis by promoting TGF-?1/Smad signaling in the skin.

SUBMITTER: Yamane T

PROVIDER: S-EPMC4821357 | biostudies-literature | 2016 Apr

REPOSITORIES: biostudies-literature

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mTOR inhibition by rapamycin increases ceramide synthesis by promoting transforming growth factor-β1/Smad signaling in the skin.

Yamane Takumi T Muramatsu Aimi A Yoshino Sawako S Matsui Sho S Shimura Mari M Tsujii Yoshimasa Y Iwatsuki Ken K Kobayashi-Hattori Kazuo K Oishi Yuichi Y

FEBS open bio 20160224 4

Although mammalian target of rapamycin (mTOR) mediates a wide variety of biological functions, little information is available on the effect of mTOR on the functions of skin cells. In this study, we investigated effects of mTOR inhibition by rapamycin on ceramide synthesis in the skin of rats and human keratinocytes and its regulatory mechanisms. The phosphorylation of p70 S6 kinase, which indicates mTOR activation, was induced in the skin of rats fed a high-fat diet, but this abnormality was re ...[more]

PMID: 27239444

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Project description:In fetal growth restriction (FGR), fetal growth is limited by reduced nutrient and oxygen supply. Insulin-like growth factor I (IGF-I) is a key regulator of fetal growth and IGF binding protein -1(IGFBP-1) is the principal regulator of fetal IGF-I bioavailability. Phosphorylation enhances IGFBP-1's affinity for IGF-I. Hypoxia induces IGFBP-1 hyperphosphorylation, markedly decreasing IGF-I bioavailability. We recently reported that fetal liver IGFBP-1 hyperphosphorylation is associated with inhibition of the mechanistic target of rapamycin (mTOR) in a nonhuman primate model of FGR. Here, we test the hypothesis that IGFBP-1 hyperphosphorylation in response to hypoxia is mediated by mTOR inhibition. We inhibited mTOR either by rapamycin or small interfering RNA (siRNA) targeting raptor (mTOR complex [mTORC]1) and/or rictor (mTORC2) in HepG2 cells cultured under hypoxia (1% O2) or basal (20% O2) conditions. Conversely, we activated mTORC1 or mTORC1+mTORC2 by silencing endogenous mTOR inhibitors (tuberous sclerosis complex 2/DEP-domain-containing and mTOR-interacting protein). Immunoblot analysis demonstrated that both hypoxia and inhibition of mTORC1 and/or mTORC2 induced similar degrees of IGFBP-1 phosphorylation at Ser101/119/169 and reduced IGF-I receptor autophosphorylation. Activation of mTORC1+mTORC2 or mTORC1 alone prevented IGFBP-1 hyperphosphorylation in response to hypoxia. Multiple reaction monitoring-mass spectrometry showed that rapamycin and/or hypoxia increased phosphorylation also at Ser98 and at a novel site Ser174. In silico structural analysis indicated that Ser174 was in close proximity to the IGF-binding site. Together, we demonstrate that signaling through the mTORC1 or mTORC2 pathway is sufficient to induce IGFBP-1 hyperphosphorylation in response to hypoxia. This study provides novel understanding of the cellular mechanism that controls fetal IGFBP-1 phosphorylation in hypoxia, and we propose that mTOR inhibition constitutes a mechanistic link between hypoxia, reduced IGF-I bioavailability and FGR.

Dataset Information

MTOR inhibition by rapamycin increases ceramide synthesis by promoting transforming growth factor-?1/Smad signaling in the skin.

Publications

mTOR inhibition by rapamycin increases ceramide synthesis by promoting transforming growth factor-β1/Smad signaling in the skin.

Similar Datasets

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