Cyclophilin a is required for angiotensin II-induced p47phox translocation to caveolae in vascular smooth muscle cells

Nwe Nwe Soe, Mark Sowden, Padmamalini Baskaran, Elaine M. Smolock, Yeonghwan Kim, Patrizia Nigro, Bradford C. Berk

Research output: Contribution to journalArticle

Abstract

Objective-Angiotensin II (AngII) signal transduction in vascular smooth muscle cells (VSMC) is mediated by reactive oxygen species (ROS). Cyclophilin A (CyPA) is a ubiquitously expressed cytosolic protein that possesses peptidyl-prolyl cis-trans isomerase activity, scaffold function, and significantly enhances AngII-induced ROS production in VSMC. We hypothesized that CyPA regulates AngII-induced ROS generation by promoting translocation of NADPH oxidase cytosolic subunit p47phox to caveolae of the plasma membrane. Approach and Results-Overexpression of CyPA in CyPA-deficient VSMC (CyPAVSMC) significantly increased AngII-stimulated ROS production. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitors (VAS2870 or diphenylene iodonium) significantly attenuated AngII-induced ROS production in CyPA and p47phox-overexpressing CyPAVSMC. Cell fractionation and sucrose gradient analyses showed that AngII-induced p47phox plasma membrane translocation, specifically to the caveolae, was reduced in CyPAVSMC compared with wild-type-VSMC. Immunofluorescence studies demonstrated that AngII increased p47phox and CyPA colocalization and translocation to the plasma membrane. In addition, immunoprecipitation of CyPA followed by immunoblotting of p47phox and actin showed that AngII increased CyPA and p47phox interaction. AngII-induced p47phox and actin cell cytoskeleton association was attenuated in CyPAVSMC. Mechanistically, inhibition of p47phox phosphorylation and phox homology domain deletion attenuated CyPA and p47phox interaction. Finally, cyclosporine A and CyPA-peptidyl-prolyl cis-trans isomerase mutant, R55A, inhibited AngII-stimulated CyPA and p47phox association in VSMC, suggesting that peptidyl-prolyl cis-trans isomerase activity was required for their interaction. Conclusions-These findings provide the mechanism by which CyPA is an important regulator for AngII-induced ROS generation in VSMC through interaction with p47phox and cell cytoskeleton, which enhances the translocation of p47phox to caveolae.

Original languageEnglish
Pages (from-to)2147-2153
Number of pages7
JournalArteriosclerosis, Thrombosis, and Vascular Biology
Volume33
Issue number9
DOIs
Publication statusPublished - Sep 2013

Keywords

  • angiotensin II
  • cyclophilin A
  • cytoskeleton
  • p47phox protein, mouse
  • reactive oxygen species

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine

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