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TGF?-induced deptor suppression recruits mTORC1 and not mTORC2 to enhance collagen I (?2) gene expression.

ABSTRACT: Enhanced TGF? activity contributes to the accumulation of matrix proteins including collagen I (?2) by proximal tubular epithelial cells in progressive kidney disease. Although TGF? rapidly activates its canonical Smad signaling pathway, it also recruits noncanonical pathway involving mTOR kinase to regulate renal matrix expansion. The mechanism by which chronic TGF? treatment maintains increased mTOR activity to induce the matrix protein collagen I (?2) expression is not known. Deptor is an mTOR interacting protein that suppresses mTOR activity in both mTORC1 and mTORC2. In proximal tubular epithelial cells, TGF? reduced deptor levels in a time-dependent manner with concomitant increase in both mTORC1 and mTORC2 activities. Expression of deptor abrogated activity of mTORC1 and mTORC2, resulting in inhibition of collagen I (?2) mRNA and protein expression via transcriptional mechanism. In contrast, neutralization of endogenous deptor by shRNAs increased activity of both mTOR complexes and expression of collagen I (?2) similar to TGF? treatment. Importantly, downregulation of deptor by TGF? increased the expression of Hif1? by increasing translation of its mRNA. TGF?-induced deptor downregulation promotes Hif1? binding to its cognate hypoxia responsive element in the collagen I (?2) gene to control its protein expression via direct transcriptional mechanism. Interestingly, knockdown of raptor to specifically block mTORC1 activity significantly inhibited expression of collagen I (?2) and Hif1? while inhibition of rictor to prevent selectively mTORC2 activation did not have any effect. Critically, our data provide evidence for the requirement of TGF?-activated mTORC1 only by deptor downregulation, which dominates upon the bystander mTORC2 activity for enhanced expression of collagen I (?2). Our results also suggest the presence of a safeguard mechanism involving deptor-mediated suppression of mTORC1 activity against developing TGF?-induced renal fibrosis.

SUBMITTER: Das F

PROVIDER: S-EPMC4198127 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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TGFβ-induced deptor suppression recruits mTORC1 and not mTORC2 to enhance collagen I (α2) gene expression.

Das Falguni F Bera Amit A Ghosh-Choudhury Nandini N Abboud Hanna E HE Kasinath Balakuntalam S BS Choudhury Goutam Ghosh GG

PloS one 20141015 10

Enhanced TGFβ activity contributes to the accumulation of matrix proteins including collagen I (α2) by proximal tubular epithelial cells in progressive kidney disease. Although TGFβ rapidly activates its canonical Smad signaling pathway, it also recruits noncanonical pathway involving mTOR kinase to regulate renal matrix expansion. The mechanism by which chronic TGFβ treatment maintains increased mTOR activity to induce the matrix protein collagen I (α2) expression is not known. Deptor is an mTO ...[more]

PMID: 25333702

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Project description:PurposeIt was previously demonstrated that opticin (OPTC) inhibits the collagen-induced promotion of bioactivities of human retinal vascular endothelial cells (hRVECs). The present in vivo study aimed to further investigate the regulatory role of opticin in vitreous collagen-mediated retinal neovascularization and to elucidate its regulatory mechanisms with regard to integrin α2-I domain-GXXGER complex formation and RhoA/ROCK1 signal change. The regulatory role of Mg2+ on integrin α2-I domain-GXXGER complex formation in the above process was also investigated.MethodsThe zebrafish model of hypoxia-induced retinopathy was established, and OPTC-overexpressing plasmids were intravitreally injected to assess the antiangiogenesis effect of opticin. The regulatory role of opticin in integrin α2-I domain-GXXGER complex formation in vivo was analyzed by mass spectrometry. The mRNA and protein expression of RhoA/ROCK1 were examined. The concentration of Mg2+ as an activator of the integrin α2-I domain-GXXGER complex was measured. Solid-phase binding assays were performed to investigate the interference of opticin in integrin α2 collagen binding and the regulatory role of Mg2+ in that process.ResultsOpticin and OPTC-overexpressing plasmid injection reduced retinal neovascularization in the zebrafish model of hypoxia-induced retinopathy. Mass spectrometry revealed that opticin could inhibit integrin α2-I domain-GXXGER complex formation. The Mg2+ concentration was also decreased by opticin, which was another indication of the complex activation. Injection of OPTC-overexpressing plasmids inhibited mRNA and the protein expression of RhoA/ROCK1 in the zebrafish model of hypoxia-induced retinopathy. The solid-phase binding assay revealed that opticin could block integrin α2-collagen I binding in the presence of Mg2+.ConclusionsOpticin exerts its antiangiogenesis effect by interfering in the Mg2+-modulated integrin α2-I domain-collagen complex formation and suppressing the downstream RhoA/ ROCK1 signaling pathway.

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TGF?-induced deptor suppression recruits mTORC1 and not mTORC2 to enhance collagen I (?2) gene expression.

Publications

TGFβ-induced deptor suppression recruits mTORC1 and not mTORC2 to enhance collagen I (α2) gene expression.

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