Poldip2 controls vascular smooth muscle cell migration by regulating focal adhesion turnover and force polarization

Abstract

Polymerase-δ-interacting protein 2 (Poldip2) interacts with NADPH oxidase 4 (Nox4) and regulates migration; however, the precise underlying mechanisms are unclear. Here, we investigated the role of Poldip2 in focal adhesion turnover, as well as traction force generation and polarization. Poldip2 overexpression (AdPoldip2) in vascular smooth muscle cells (VSMCs) impairs PDGF-induced migration and induces a characteristic phenotype of long cytoplasmic extensions. AdPoldip2 also prevents the decrease in spreading and increased aspect ratio observed in response to PDGF and slightly impairs cell contraction. Moreover, AdPoldip2 blocks focal adhesion dissolution and sustains H2O2 levels in focal adhesions, whereas Poldip2 knockdown (siPoldip2) significantly decreases the number of focal adhesions. RhoA activity is unchanged when focal adhesion dissolution is stimulated in control cells but increases in AdPoldip2-treated cells. Inhibition of RhoA blocks Poldip2-mediated attenuation of focal adhesion dissolution, and overexpression of RhoA or focal adhesion kinase (FAK) reverses the loss of focal adhesions induced by siPoldip2, indicating that RhoA and FAK mediate the effect of Poldip2 on focal adhesions. Nox4 silencing prevents focal adhesion stabilization by AdPoldip2 and induces a phenotype similar to siPoldip2, suggesting a role for Nox4 in Poldip2-induced focal adhesion stability. As a consequence of impaired focal adhesion turnover, PDGF-treated AdPoldip2 cells are unable to reduce and polarize traction forces, a necessary first step in migration. These results implicate Poldip2 in VSMC migration via regulation of focal adhesion turnover and traction force generation in a Nox4/RhoA/FAK-dependent manner.

Immune Cell Induced Migration of Osteoprogenitor Cells Is Mediated by TGF-β Dependent Upregulation of NOX4 and Activation of Focal Adhesion Kinase

“…Blocking NOX4 activity/ROS formation delayed the gap closure (scratch assay) in the presence of rhTGF-β 1 , as well as the cell migration towards rhTGF-β 1 (under agar spot assay). In migrating vascular smooth muscle cell, induction of NOX4 and associated ROS accumulation lead to an activation of FAK [23]. This is also seen in rhTGF-β 1 treated phOBs in our study.…”

Loss of Smooth Muscle α-Actin Leads to NF-κB–Dependent Increased Sensitivity to Angiotensin II in Smooth Muscle Cells and Aortic Enlargement

“…To identify the source of increased ROS, the expression of the NADPH oxidases were assessed, and expression of Nox4 and p22 phox , along with a known activator of Nox4, Poldip2 , are all significantly increased in the Acta2 −/− SMCs (Figure 4C) (17,18). There is no significant change in expression of Nox1 or Nox2 (Online Figure VIC).…”

Polymerase δ-interacting Protein 2: A Multifunctional Protein

“…Poldip2 is fairly ubiquitously expressed, having been reported in aorta, 3, 4 lung, 3 kidney, 3, 5, 6 heart 7 and thyroid, 8 as well as many cell types including mouse aortic smooth muscle cells, 9 astrocytes, 10 vascular smooth muscle cells (VSMC), 3, 11 brain endothelial cells, 12 epithelial cells, 12 HeLa, 1, 12-15 fibroblasts, 2, 13, 16, 17 kidney fibroblasts, 18 myoblasts, 19 human umbilical vein endothelial cells, 20 human embryonic kidney cells 293 (HEK 293), 13 MRC5 cells, 21, 22 and cortical neurons. 23 There is also evidence of Poldip2 expression in human breast cancer, 24 human breast adenocarcinoma cells 13 and human lung adenocarcinoma cells.…”