CdGAP is required for transforming growth factor β- and Neu/ErbB-2-induced breast cancer cell motility and invasion.
ABSTRACT RhoA, Rac1 and Cdc42, the best-characterized members of the Rho family of small GTPases, are critical regulators of many cellular activities. Cdc42 GTPase-activating protein (CdGAP) is a serine- and proline-rich RhoGAP protein showing GAP activity against both Cdc42 and Rac1 but not RhoA. CdGAP is phosphorylated downstream of the MEK-ERK (extracellular signal-regulated kinase) pathway in response to serum and is required for normal cell spreading and polarized lamellipodia formation. In this study, we found that CdGAP protein and mRNA levels are highly increased in mammary tumor explants expressing an activated Neu/ErbB-2 (Neu-NT) receptor. In response to transforming growth factor-β (TGFβ) stimulation, Neu-NT-expressing mammary tumor explants demonstrate a clear induction in cell motility and invasion. We show that downregulation of CdGAP expression by small interfering RNA abrogates the ability of TGFβ to induce cell motility and invasion of Neu-NT-expressing mammary tumor explants. However, it has no effect on TGFβ-mediated cell adhesion on type 1 collagen and fibronectin. Interestingly, protein expression of E-Cadherin is highly increased in Neu-NT-expressing mammary tumor explants depleted of CdGAP. In addition, complete loss of E-Cadherin expression is not observed in CdGAP-depleted cells during TGFβ-mediated epithelial to mesenchymal transition. Downregulation of the CdGAP expression also decreases cell proliferation of Neu-NT-expressing mammary tumor explants independently of TGFβ. Rescue analysis using re-expression of various CdGAP deletion-mutant proteins revealed that the proline-rich domain (PRD) but not the GAP domain of CdGAP is essential to mediate TGFβ-induced cell motility and invasion. Finally, we found that TGFβ induces the expression and phosphorylation of CdGAP in mammary epithelial NMuMG cells. Taken together, these studies identify CdGAP as a novel molecular target in TGFβ signaling and implicate CdGAP as an essential component in the synergistic interaction between TGFβ and Neu/ErbB-2 signaling pathways in breast cancer cells.
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ABSTRACT: The formation and stability of epithelial adhesive systems, such as adherens junctions, desmosomes and tight junctions, rely on a number of cellular processes that ensure a dynamic interaction with the cortical cytoskeleton, and appropriate delivery and turnover of receptors at the surface. Unique signalling pathways must be coordinated to allow the coexistence of distinct adhesive systems at discrete sub-domains along junctions and the specific properties they confer to epithelial cells. Rho, Rac and Cdc42 are members of the Rho small GTPase family, and are well-known regulators of cell-cell adhesion. The spatio-temporal control of small GTPase activation drives specific intracellular processes to enable the hierarchical assembly, morphology and maturation of cell-cell contacts. Here, we discuss the small GTPase regulators that control the precise amplitude and duration of the levels of active Rho at cell-cell contacts, and the mechanisms that tailor the output of Rho signalling to a particular cellular event. Interestingly, the functional interaction is reciprocal; Rho regulators drive the maturation of cell-cell contacts, whereas junctions can also modulate the localisation and activity of Rho regulators to operate in diverse processes in the epithelial differentiation programme.Journal of Cell Science 01/2013; 126(Pt 2):379-91. DOI:10.1242/jcs.097923 · 5.33 Impact Factor
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ABSTRACT: Motile cells are capable of sensing the stiffness of the surrounding extracellular matrix through integrin-mediated focal adhesions and migrate towards regions of higher rigidity in a process known as durotaxis. Durotaxis plays an important role in normal development and disease progression, including tumor invasion and metastasis. However, the signaling mechanisms underlying focal adhesion-mediated rigidity sensing and durotaxis are poorly understood. Utilizing matrix-coated polydimethylsiloxane gels to manipulate substrate compliance, we show that cdGAP, an adhesion-localized Rac1 and Cdc42 specific GTPase activating protein, is necessary for U2OS osteosarcoma cells to coordinate cell shape changes and migration as a function of extracellular matrix stiffness. CdGAP regulated rigidity-dependent motility by controlling membrane protrusion and adhesion dynamics, as well as by modulating Rac1 activity. CdGAP was also found to be necessary for U2OS cell durotaxis. Taken together, these data identify cdGAP as an important component of an integrin-mediated signaling pathway that senses and responds to mechanical cues in the extracellular matrix in order to coordinate directed cell motility.PLoS ONE 03/2014; 9(3):e91815. DOI:10.1371/journal.pone.0091815 · 3.53 Impact Factor
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ABSTRACT: Rho family small GTPases are involved in the spatio-temporal regulation of several physiological processes. They operate as molecular switches based on their GTP- or GDP-bound state. Their GTPase activator proteins (Rho/Rac GAPs) are able to increase the GTP hydrolysis of small GTPases, which turns them to an inactive state. This regulatory step is a key element of signal termination. According to the human genome project the potential number of Rho family GAPs is approximately 70. Despite their significant role in cellular signaling our knowledge on their expression pattern is quite incomplete. In this study we tried to reveal the tissue-distribution of Rho/Rac GAPs based on expressed sequence tag (EST) database from healthy and tumor tissues and microarray experiments. Our accumulated data sets can provide important starting information for future research. However, the nomenclature of Rho family GAPs is quite heterogeneous. Therefore we collected the available names, abbreviations and aliases of human Rho/Rac GAPs in a useful nomenclature table. A phylogenetic tree and domain structure of 65 human RhoGAPs are also presented.Small GTPases 03/2013; 4(2). DOI:10.4161/sgtp.23708