Dataset Information

Hic-5 Mediates TGF?-Induced Adhesion in Vascular Smooth Muscle Cells by a Nox4-Dependent Mechanism.

ABSTRACT: Focal adhesions (FAs) link the cytoskeleton to the extracellular matrix and as such play important roles in growth, migration, and contractile properties of vascular smooth muscle cells. Recently, it has been shown that downregulation of Nox4, a transforming growth factor (TGF) ?-inducible, hydrogen peroxide (H2O2)-producing enzyme, affects the number of FAs. However, the effectors downstream of Nox4 that mediate FA regulation are unknown. The FA resident protein H2O2-inducible clone (Hic)-5 is H2O2 and TGF? inducible, and a binding partner of the heat shock protein (Hsp) 27. The objective of this study was to elucidate the mechanism, by which Hic-5 and Hsp27 participate in TGF?-induced, Nox4-mediated vascular smooth muscle cell adhesion and migration.Through a combination of molecular biology and biochemistry techniques, we found that TGF?, by a Nox4-dependent mechanism, induces the expression and interaction of Hic-5 and Hsp27, which is essential for Hic-5 localization to FAs. Importantly, we found that Hic-5 expression is required for the TGF?-mediated increase in FA number, adhesive forces and migration. Mechanistically, Nox4 downregulation impedes Smad (small body size and mothers against decapentaplegic) signaling by TGF?, and Hsp27 and Hic-5 upregulation by TGF? is blocked in small body size and mothers against decapentaplegic 4-deficient cells.Hic-5 and Hsp27 are effectors of Nox4 required for TGF?-stimulated FA formation, adhesion strength and migration in vascular smooth muscle cell.

SUBMITTER: Fernandez I

PROVIDER: S-EPMC4426206 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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Hic-5 Mediates TGFβ-Induced Adhesion in Vascular Smooth Muscle Cells by a Nox4-Dependent Mechanism.

Fernandez Isabel I Martin-Garrido Abel A Zhou Dennis W DW Clempus Roza E RE Seidel-Rogol Bonnie B Valdivia Alejandra A Lassègue Bernard B García Andrés J AJ Griendling Kathy K KK San Martin Alejandra A

Arteriosclerosis, thrombosis, and vascular biology 20150326 5

<h4>Objective</h4>Focal adhesions (FAs) link the cytoskeleton to the extracellular matrix and as such play important roles in growth, migration, and contractile properties of vascular smooth muscle cells. Recently, it has been shown that downregulation of Nox4, a transforming growth factor (TGF) β-inducible, hydrogen peroxide (H2O2)-producing enzyme, affects the number of FAs. However, the effectors downstream of Nox4 that mediate FA regulation are unknown. The FA resident protein H2O2-inducible ...[more]

PMID: 25814672

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Project description:Efficient mechanotransduction in vascular smooth muscle cells (VSMCs) is intimately coupled to physical coupling of the cell to extracellular matrix proteins (ECM) by integrins. Integrin adhesion receptors are essential for normal vascular function and defective integrin signaling is associated with cardiovascular disease. However, less is known about the mechanism of integrin activation in VSMCs in relation to vasoregulation. Our laboratory previously demonstrated that the vasoconstrictor Angiotensin II increases VSMC stiffness in concert with enhanced adhesion to fibronectin (FN), indicating an important role for adhesion in contraction. However, the mechanism of this coordination remains to be clarified. In this study, intracellular Ca2+ ([Ca2+]i) was hypothesized to link integrin activation through inside-out signaling pathways leading to enhanced adhesion in response to AII. By using atomic force microscopy (AFM) with an anti-?5 antibody coated AFM probe, we confirmed that cell stiffness was increased by AII, while we observed no change in adhesion to an ?5 integrin antibody. This indicated that increases in cell adhesion to FN induced by AII were occurring through an integrin activation process, as increased membrane integrin expression/receptor density would have been accompanied by increased adhesion to the anti-?5 antibody. Further studies were performed using either KCl or BAPTA-AM to modulate the level of [Ca2+]i. After KCl, VSMCs showed a rapid transient increase in cell stiffness as well as cell adhesion to FN, and these two events were synchronized with superimposed transient increases in the level of [Ca2+]i, which was measured using the Ca2+ indicator, fluo-4. These relationships were unaffected in VSMCs pretreated with the myosin light chain kinase inhibitor, ML-7. In contrast, unstimulated VSMCs incubated with an intracellular calcium chelator, BAPTA-AM, showed reduced cell adhesion to FN as well the expected decrease in [Ca2+]i. These data suggest that in VSMCs, integrin activation is linked to signaling events tied to levels of [Ca2+]i while being less dependent on events at the level of contractile protein activation. These findings provide additional evidence to support a role for adhesion in VSMC contraction and suggest that following cell contractile activation, that adhesion may be regulated in tandem with the contractile event.

Dataset Information

Hic-5 Mediates TGF?-Induced Adhesion in Vascular Smooth Muscle Cells by a Nox4-Dependent Mechanism.

Publications

Hic-5 Mediates TGFβ-Induced Adhesion in Vascular Smooth Muscle Cells by a Nox4-Dependent Mechanism.

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