Role of IκB Kinase-β in the Growth-Promoting Effects of Angiotensin II In Vitro and In Vivo

Doyon, Priscilla; Zuylen, Wendy J. van; Servant, Marc J.

doi:10.1161/atvbaha.113.302487

Cited by 6 publications

(7 citation statements)

References 43 publications

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“…Protein extracts were prepared and analyses were performed as described previously (50). The primary antibodies used were mouse anti-ROR2 (QED Bioscience; 1:1,000), rabbit anti-JNK (Cell Signaling; 1:1,000), mouse anti-RhoA (1B12; Abcam; 1:1,000), mouse anti-CMV immediate early/early (IE/E) (Dako clone CCH2ϩDDG9; 1:1,000), and mouse anti-␤-actin (Sigma; 1:2,000).…”

Section: Methodsmentioning

confidence: 99%

Human Cytomegalovirus Modulates Expression of Noncanonical Wnt Receptor ROR2 To Alter Trophoblast Migration

Zuylen

Ford

Wong

et al. 2016

J Virol

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Maternal primary cytomegalovirus (CMV) infection, reactivation, or reinfection with a different viral strain may cause fetal injury and adverse pregnancy outcomes. Increasing evidence indicates that fetal injury results not only from direct viral cytopathic damage to the CMV-infected fetus but also from indirect effects through placental infection and dysfunction. CMV alters Wingless (Wnt) signaling, an essential cellular pathway involved in placentation, as evidenced by reduced transcription of canonical Wnt target genes and decreased Wnt3a-induced trophoblast migration. Whether CMV affects the noncanonical Wnt signaling pathway has been unclear. This study demonstrates for the first time that CMV infection inhibits Wnt5a-stimulated migration of human SGHPL-4 trophoblasts and that inhibition of the pathway restores normal migration of CMV-infected cells. Western blot and real-time PCR analyses show increased expression of noncanonical Wnt receptor ROR2 in CMV-infected trophoblasts. Mimicking the CMV-induced ROR2 protein expression via ectopic expression inhibited Wnt5a-induced trophoblast migration and reduced T cell-specific factor (TCF)/lymphoid enhancer-binding factor (LEF)-mediated transcription as measured using luciferase reporter assays. Gene silencing using small interfering RNA (siRNA) duplexes decreased ROR2 transcript and protein levels. In contrast, proliferation of SGHPL-4 trophoblasts, measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was not affected. The siRNA-mediated downregulation of ROR2 in trophoblasts rescued CMV-induced reduction in trophoblast migration. These data suggest a mechanism where CMV alters the expression of the Wnt receptor ROR2 to alter Wnt5a-mediated signaling and inhibit trophoblast motility. Inhibition of this mechanism may be a target for therapeutic intervention for CMV-induced placental damage and consequent fetal damage in congenital CMV infections. IMPORTANCEMaternal primary cytomegalovirus (CMV) infection, reactivation, or reinfection with a different viral strain may cause fetal injury and adverse pregnancy outcomes. Increasing evidence indicates that fetal injury results not only from direct viral cytopathic damage to the CMV-infected fetus but also from indirect effects through placental infection and placental dysfunction. No effective therapy is currently proven to prevent or treat congenital CMV infection. Understanding the molecular underpinnings of CMV infection of the placenta is essential for therapeutic innovations and vaccine design. CMV alters canonical Wingless (Wnt) signaling, an essential cellular pathway involved in placental development. This study suggests a mechanism in which CMV alters the expression of noncanonical Wnt receptor ROR2 to alter motility of placental cells, which has important implications in the pathogenesis of CMV-induced placental dysfunction. Inhibition of this mechanism may be a target for therapeutic intervention for CMV-induced placental damage and consequent fetal damage in con...

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Section: Methodsmentioning

confidence: 99%

Human Cytomegalovirus Modulates Expression of Noncanonical Wnt Receptor ROR2 To Alter Trophoblast Migration

Zuylen

Ford

Wong

et al. 2016

J Virol

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“…Our findings underscore the importance of studying such complex cellular responses in different cell types, in particular relevant physiological ones like VSMCs. Despite VSMCs expressing AT 1 R and being responsive to Ang II (even in late passage tissue culture) (42)(43)(44)(62)(63)(64) for MAPK activation, the signaling pathway leading to cell migration differs from HEK 293 cells as observed here. A detailed analysis in multiple cell lines may indeed reveal that signaling pathways engaged by receptors to promote a specific cellular response can vary between cell types but also for the same cell type depending on its origin as shown previously for MAPK in HEK 293 cells (65).…”

Section: Discussionmentioning

confidence: 65%

“…VSMCs were isolated from rat aorta as described under "Experimental Procedures." We and others have shown that these cells, when cultured in the conditions described, retained the expression of AT 1 R and that Ang II stimulation resulted in the activation of numerous cellular events, which could be blocked by selective AT 1 R antagonists (42)(43)(44)49). As illustrated in Fig.…”

Section: Volume 291 • Number 8 • February 19 2016mentioning

confidence: 70%

“…Aortic VSMCs, isolated from Wistar rats by explant, were provided by Dr. Marc Servant (Université de Montréal, Montreal, Canada) and used at passages 7-10. These cells were characterized and shown to be responsive to Ang II as well as to express AT 1 R until passage 16 (41)(42)(43)(44). Here, experiments were performed and repeated in cells from passages 7-16 to ensure reproducibility of the results in all assays.…”

Section: Reagents and Antibodiesmentioning

confidence: 99%

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β-Arrestin-mediated Angiotensin II Signaling Controls the Activation of ARF6 Protein and Endocytosis in Migration of Vascular Smooth Muscle Cells

Charles

Namkung

Cotton

et al. 2016

Journal of Biological Chemistry

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Angiotensin II (Ang II) is a vasopressive hormone but is also a potent activator of cellular migration. We have previously shown that it can promote the activation of the GTPase ARF6 in a heterologous overexpressing system. The molecular mechanisms by which receptors control the activation of this small G protein remain, however, largely unknown. Furthermore, how ARF6 coordinates the activation of complex cellular responses needs to be further elucidated. In this study, we demonstrate that Ang II receptors engage ␤-arrestin, but not G q , to mediate ARF6 activation in HEK 293 cells. To further confirm the key role of ␤-arrestin proteins, we overexpressed ␤-arrestin2-(1-320), a dominant negative mutant known to block receptor endocytosis. We show that expression of this truncated construct does not support the activation of the GTPase nor cell migration. Interestingly, ␤-arrestin2 can interact with the ARF guanine nucleotide exchange factor ARNO, although the C-terminally lacking mutant does not. We finally examined whether receptor endocytosis controlled ARF6 activation and cell migration. Although the clathrin inhibitor PitStop2 did not impact the ability of Ang II to activate ARF6, cell migration was markedly impaired. To further show that ARF activation regulates key signaling events leading to migration, we also examined MAPK activation. We demonstrate that this signaling axis is relevant in smooth muscle cells of the vasculature. Altogether, our findings show for the first time that Ang II receptor signaling to ␤-arrestin regulates ARF6 activation. These proteins together control receptor endocytosis and ultimately cell migration.

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“…Peroxisome PPAR-γ coactivator 1α deletion caused vascular dysfunction and inflammation during chronic angiotensin II infusion by increasing mitochondrial ROS production. 30 Furthermore, the findings by Doyon et al 31 provided new insights into the molecular mechanism of the pathological role of angiotensin II and assist in identifying the beneficial effects of IκB kinase-β inhibition for the treatment of cardiovascular diseases. Finally, perivascular adipose tissue wraps blood vessels and modulates vasoreactivity by secretion of vasoactive molecules.…”

Section: Adventitial Tissues and Inflammatory Cells For Vascular Funcmentioning

confidence: 99%

Vascular Function

Shimokawa

Satoh

2014

ATVB

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Role of IκB Kinase-β in the Growth-Promoting Effects of Angiotensin II In Vitro and In Vivo

Cited by 6 publications

References 43 publications

Human Cytomegalovirus Modulates Expression of Noncanonical Wnt Receptor ROR2 To Alter Trophoblast Migration

Human Cytomegalovirus Modulates Expression of Noncanonical Wnt Receptor ROR2 To Alter Trophoblast Migration

β-Arrestin-mediated Angiotensin II Signaling Controls the Activation of ARF6 Protein and Endocytosis in Migration of Vascular Smooth Muscle Cells

Vascular Function

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