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High glucose enhances microRNA-26a to activate mTORC1 for mesangial cell hypertrophy and matrix protein expression.

ABSTRACT: High glucose milieu inhibits PTEN expression to activate Akt kinase and induces glomerular mesangial cell hypertrophy and matrix protein expression in diabetic nephropathy. Specific mechanism by which high glucose inhibits PTEN expression is not clear. We found that high glucose increased the expression of the microRNA-26a (miR-26a) in mesangial cells. Using a sensor plasmid with 3'UTR-driven luciferase, we showed PTEN as a target of miR-26a in response to high glucose. Overexpression of miR-26a reduced the PTEN protein levels resulting in increased Akt kinase activity similar to high glucose treatment. In contrast, anti-miR-26a reversed high glucose-induced suppression of PTEN with concomitant inhibition of Akt kinase activity. Akt-mediated phosphorylation of tuberin and PRAS40 regulates mTORC1, which is necessary for mesangial cell hypertrophy and matrix protein expression. Inhibition of high glucose-induced miR-26a blocked phosphorylation of tuberin and PRAS40, which lead to suppression of phosphorylation of S6 kinase and 4EBP-1, two substrates of mTORC1. Furthermore, we show that expression of miR-26a induced mesangial cell hypertrophy and increased fibronectin and collagen I (?2) expression similar to that observed with the cells incubated with high glucose. Anti-miR-26a inhibited these phenomena in response to high glucose. Together our results provide the first evidence for the involvement of miR-26a in high glucose-induced mesangial cell hypertrophy and matrix protein expression. These data indicate the potential therapeutic utility of anti-miR-26a for the complications of diabetic kidney disease.

SUBMITTER: Dey N

PROVIDER: S-EPMC4437875 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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High glucose enhances microRNA-26a to activate mTORC1 for mesangial cell hypertrophy and matrix protein expression.

Dey Nirmalya N Bera Amit A Das Falguni F Ghosh-Choudhury Nandini N Kasinath Balakuntalam S BS Choudhury Goutam Ghosh GG

Cellular signalling 20150320 7

High glucose milieu inhibits PTEN expression to activate Akt kinase and induces glomerular mesangial cell hypertrophy and matrix protein expression in diabetic nephropathy. Specific mechanism by which high glucose inhibits PTEN expression is not clear. We found that high glucose increased the expression of the microRNA-26a (miR-26a) in mesangial cells. Using a sensor plasmid with 3'UTR-driven luciferase, we showed PTEN as a target of miR-26a in response to high glucose. Overexpression of miR-26a ...[more]

PMID: 25797045

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Project description:Transforming growth factor-? (TGF?) promotes glomerular hypertrophy and matrix expansion, leading to glomerulosclerosis. MicroRNAs are well suited to promote fibrosis because they can repress gene expression, which negatively regulate the fibrotic process. Recent cellular and animal studies have revealed enhanced expression of microRNA, miR-21, in renal cells in response to TGF?. Specific miR-21 targets downstream of TGF? receptor activation that control cell hypertrophy and matrix protein expression have not been studied. Using 3'UTR-driven luciferase reporter, we identified the tumor suppressor protein PTEN as a target of TGF?-stimulated miR-21 in glomerular mesangial cells. Expression of miR-21 Sponge, which quenches endogenous miR-21 levels, reversed TGF?-induced suppression of PTEN. Additionally, miR-21 Sponge inhibited TGF?-stimulated phosphorylation of Akt kinase, resulting in attenuation of phosphorylation of its substrate GSK3?. Tuberin and PRAS40, two other Akt substrates, and endogenous inhibitors of mTORC1, regulate mesangial cell hypertrophy. Neutralization of endogenous miR-21 abrogated TGF?-stimulated phosphorylation of tuberin and PRAS40, leading to inhibition of phosphorylation of S6 kinase, mTOR and 4EBP-1. Moreover, downregulation of miR-21 significantly suppressed TGF?-induced protein synthesis and hypertrophy, which were reversed by siRNA-targeted inhibition of PTEN expression. Similarly, expression of constitutively active Akt kinase reversed the miR-21 Sponge-mediated inhibition of TGF?-induced protein synthesis and hypertrophy. Furthermore, expression of constitutively active mTORC1 prevented the miR-21 Sponge-induced suppression of mesangial cell protein synthesis and hypertrophy by TGF?. Finally, we show that miR-21 Sponge inhibited TGF?-stimulated fibronectin and collagen expression. Suppression of PTEN expression and expression of both constitutively active Akt kinase and mTORC1 independently reversed this miR-21-mediated inhibition of TGF?-induced fibronectin and collagen expression. Our results uncover an essential role of TGF?-induced expression of miR-21, which targets PTEN to initiate a non-canonical signaling circuit involving Akt/mTORC1 axis for mesangial cell hypertrophy and matrix protein synthesis.

Project description:Interaction of transforming growth factor-β (TGFβ)-induced canonical signaling with the noncanonical kinase cascades regulates glomerular hypertrophy and matrix protein deposition, which are early features of glomerulosclerosis. However, the specific target downstream of the TGFβ receptor involved in the noncanonical signaling is unknown. Here, we show that TGFβ increased the catalytic loop phosphorylation of platelet-derived growth factor receptor β (PDGFRβ), a receptor tyrosine kinase expressed abundantly in glomerular mesangial cells. TGFβ increased phosphorylation of the PI 3-kinase-interacting Tyr-751 residue of PDGFRβ, thus activating Akt. Inhibition of PDGFRβ using a pharmacological inhibitor and siRNAs blocked TGFβ-stimulated phosphorylation of proline-rich Akt substrate of 40 kDa (PRAS40), an intrinsic inhibitory component of mTORC1, and prevented activation of mTORC1 in the absence of any effect on Smad 2/3 phosphorylation. Expression of constitutively active myristoylated Akt reversed the siPDGFRβ-mediated inhibition of mTORC1 activity; however, co-expression of the phospho-deficient mutant of PRAS40 inhibited the effect of myristoylated Akt, suggesting a definitive role of PRAS40 phosphorylation in mTORC1 activation downstream of PDGFRβ in mesangial cells. Additionally, we demonstrate that PDGFRβ-initiated phosphorylation of PRAS40 is required for TGFβ-induced mesangial cell hypertrophy and fibronectin and collagen I (α2) production. Increased activating phosphorylation of PDGFRβ is also associated with enhanced TGFβ expression and mTORC1 activation in the kidney cortex and glomeruli of diabetic mice and rats, respectively. Thus, pursuing TGFβ noncanonical signaling, we identified how TGFβ receptor I achieves mTORC1 activation through PDGFRβ-mediated Akt/PRAS40 phosphorylation to spur mesangial cell hypertrophy and matrix protein accumulation. These findings provide support for targeting PDGFRβ in TGFβ-driven renal fibrosis.

Dataset Information

High glucose enhances microRNA-26a to activate mTORC1 for mesangial cell hypertrophy and matrix protein expression.

Publications

High glucose enhances microRNA-26a to activate mTORC1 for mesangial cell hypertrophy and matrix protein expression.

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