Nox4 (NADPH oxidase 4) and PolDIP2 (polymerase δ-interacting protein 2) induce filamentous actin oxidation and promote its interaction with vinculin during integrin-mediated cell adhesion

Sasa Vukelic, Qian Xu, Bonnie Seidel-Rogol, Elizabeth A. Faidley, Anna E. Dikalova, Lula L. Hilenski, Ulrich Jorde, Leslie B. Poole, Bernard Lassègue, Guogang Zhang, Kathy K. Griendling

Research output: Contribution to journalArticle

6 Scopus citations

Abstract

Objective-Actin cytoskeleton assembly and organization, as a result of focal adhesion (FA) formation during cell adhesion, are dependent on reactive oxygen species and the cellular redox environment. Poldip2 (polymerase δ-interacting protein 2), a novel regulator of NOX4 (NADPH oxidase 4), plays a significant role in reactive oxygen species production and cytoskeletal remodeling. Thus, we hypothesized that endogenous reactive oxygen species derived from Poldip2/NOX4 contribute to redox regulation of actin and cytoskeleton assembly during integrin-mediated cell adhesion. Approach and Results-Using vascular smooth muscle cells, we verified that hydrogen peroxide (H2O2) levels increase during integrin-mediated cell attachment as a result of activation of NOX4. Filamentous actin (F-actin) was oxidized by sulfenylation during cell attachment, with a peak at 3 hours (0.80±0.04 versus 0.08±0.13 arbitrary units at time zero), which was enhanced by overexpression of Poldip2. Depletion of Poldip2 or NOX4 using siRNA, or scavenging of endogenous H2O2 with catalase, inhibited F-actin oxidation by 78±26%, 99±1%, and 98±1%, respectively. To determine the consequence of F-actin oxidation, we examined the binding of F-actin to vinculin, a protein involved in FA complexes that regulates FA maturation. Vinculin binding during cell adhesion as well as migration capacity were inhibited after transfection with actin containing 2 oxidation-resistant point mutations (C272A and C374A). Silencing of Poldip2 or NOX4 also impaired actin-vinculin interaction, which disturbed maturation of FAs and inhibited cell migration. Conclusions-These results suggest that integrin engagement during cell attachment activates Poldip2/Nox4 to oxidize actin, which modulates FA assembly.

Original languageEnglish (US)
Pages (from-to)2423-2434
Number of pages12
JournalArteriosclerosis, thrombosis, and vascular biology
Volume38
Issue number10
DOIs
StatePublished - Jan 1 2018

Keywords

  • Actins
  • Cell adhesion
  • NADPH oxidase
  • Reactive oxygen species
  • Vinculin

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine

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