TY - JOUR
T1 - 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
AU - Vukelic, Sasa
AU - Xu, Qian
AU - Seidel-Rogol, Bonnie
AU - Faidley, Elizabeth A.
AU - Dikalova, Anna E.
AU - Hilenski, Lula L.
AU - Jorde, Ulrich
AU - Poole, Leslie B.
AU - Lassègue, Bernard
AU - Zhang, Guogang
AU - Griendling, Kathy K.
N1 - Publisher Copyright:
© 2018 American Heart Association, Inc.
PY - 2018
Y1 - 2018
N2 - 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.
AB - 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.
KW - Actins
KW - Cell adhesion
KW - NADPH oxidase
KW - Reactive oxygen species
KW - Vinculin
UR - http://www.scopus.com/inward/record.url?scp=85055606405&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85055606405&partnerID=8YFLogxK
U2 - 10.1161/ATVBAHA.118.311668
DO - 10.1161/ATVBAHA.118.311668
M3 - Article
C2 - 30354218
AN - SCOPUS:85055606405
SN - 1079-5642
VL - 38
SP - 2423
EP - 2434
JO - Arteriosclerosis, thrombosis, and vascular biology
JF - Arteriosclerosis, thrombosis, and vascular biology
IS - 10
ER -