Cdc42 and p190RhoGAP activation by CCN2 regulates cell spreading and polarity and induces actin disassembly in migrating keratinocytes

Abstract

Cell migration requires spatiotemporal integration of signals that regulate cytoskeletal dynamics. In response to a migration-promoting agent, cells begin to polarise and extend protrusions in the direction of migration. These cytoskeletal rearrangements are orchestrated by a variety of proteins, including focal adhesion kinase (FAK) and the Rho family of GTPases. CCN2, also known as connective tissue growth factor, has emerged as a regulator of cell migration but the mechanism by which CCN2 regulates keratinocyte function is not well understood. In this article, we sought to elucidate the basic mechanism of CCN2-induced cell migration in human keratinocytes. Immunohistochemical staining was used to demonstrate that treatment with CCN2 induces a migratory phenotype through actin disassembly, spreading of lamellipodia and re-orientation of the Golgi. In vitro assays were used to show that CCN2-induced cell migration is dependent on FAK, RhoA and Cdc42, but independent of Rac1. CCN2-treated keratinocytes displayed increased Cdc42 activity and decreased RhoA activity up to 12 hours post-treatment, with upregulation of p190RhoGAP. An improved understanding of how CCN2 regulates cell migration may establish the foundation for future therapeutics in fibrotic and neoplastic diseases.

GOLPH3 drives cell migration by promoting Golgi reorientation and directional trafficking to the leading edge

“…Cdc42 has been proposed to be a key signaling molecule that drives cell polarization and cell migration (Nobes and Hall, 1999; Etienne-Manneville and Hall, 2001), although with some disagreement between experiments using dominant-negative, knockout, and small molecule inhibitors in different cell types and culture conditions (Nobes and Hall, 1999; Etienne-Manneville and Hall, 2001; Cau and Hall, 2005; Czuchra et al , 2005; Kiwanuka et al , 2016). We tested the requirement for Cdc42 for GOLPH3-driven cell migration using the specific Cdc42 inhibitor, ML141 (Hong et al , 2013; Kiwanuka et al , 2016).…”

“…We tested the requirement for Cdc42 for GOLPH3-driven cell migration using the specific Cdc42 inhibitor, ML141 (Hong et al , 2013; Kiwanuka et al , 2016). We performed scratch wound healing assays in control (DMSO) or ML141-treated cells (Figure 3E and Supplemental Figure 2D).…”

Vascular Endothelial Cell-Specific Connective Tissue Growth Factor (CTGF) Is Necessary for Development of Chronic Hypoxia-Induced Pulmonary Hypertension

“…However, changes in endothelial cytoskeleton, adherens junctions, and permeability do correlate with Cdc42 activity in a model of hypoxia-induced neonatal pulmonary hypertension (Wojciak-Stothard et al, 2006 ). Importantly, CTGF works in concert with other protein mediators to increase the angiogenic response to injury, including downstream Cdc42 activation (Pi et al, 2012 ; Kiwanuka et al, 2016 ). Thus, it is interesting to speculate—based on our findings—that the spatiotemporal integration of signals that regulate cytoskeletal dynamics through CTGF, specifically Cdc42, are necessary for PH development.…”

Loss of cell polarity regulated by PTEN/Cdc42 enrolled in the process of Hepatopulmonary Syndrome

“…The preparation of GST fusion proteins and the pull down assay for active Rac1, Cdc42 and RhoA was performed according to a previously reported procedure. 23 Briefly, PMVECs were grown in six wells culture-plates for 2 days and stimulated with 5% normal or CBDL rat serum (4 weeks). For treatment groups, PMVECs were added with 0.05 μM PTEN inhibitor BpV(pic) (Abcam) or 5 μM Casin (Sigma-Aldrich) for 24 hours in different groups.…”