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Santa Maria Imbaro, Italy

Pankova D.,Charles University | Jobe N.,Charles University | Kratochvilova M.,Charles University | Buccione R.,Tumour Cell Invasion Laboratory | And 2 more authors.
European Journal of Cell Biology | Year: 2012

The aim of this study was to analyze the potential role of NG2 chondroitin sulfate proteoglycan in amoeboid morphology and invasiveness of cancer cells. In the highly metastatic amoeboid cell lines A3 and A375M2, siRNA-mediated down-regulation of NG2 induced an amoeboid-mesenchymal transition associated with decreased invasiveness in 3D collagen and inactivation of the GTPase Rho. Conversely, the expression of NG2 in mesenchymal sarcoma K2 cells as well as in A375M2 cells resulted in an enhanced amoeboid phenotype associated with increased invasiveness and elevated Rho-GTP levels. Remarkably, the amoeboid-mesenchymal transition in A375M2 cells triggered by NG2 down-regulation was associated with increased extracellular matrix-degrading ability, although this was not sufficient to compensate for the decreased invasive capability caused by down-regulated Rho/ROCK signaling. Conversely, in K2 cells with overexpression of NG2, the ability to degrade the extracellular matrix was greatly reduced. Taken together, we suggest that NG2-mediated activation of Rho leading to effective amoeboid invasiveness is a possible mechanism through which NG2 could contribute to tumor cell invasion and metastasis. © 2012 Elsevier GmbH. Source


Genot E.,University of Bordeaux 1 | Genot E.,French Institute of Health and Medical Research | Genot E.,European Institute of Chemistry and Biology | Genot E.,Bordeaux University Hospital Center | And 5 more authors.
Journal of Cell Science | Year: 2012

Disabling mutations in the FGD1 gene cause faciogenital dysplasia (also known as Aarskog-Scott syndrome), a human X-linked developmental disorder that results in disproportionately short stature, facial, skeletal and urogenital anomalies, and in a number of cases, mild mental retardation. FGD1 encodes the guanine nucleotide exchange factor FGD1, which is specific for the Rho GTPase cell division cycle 42 (CDC42). CDC42 controls cytoskeleton-dependent membrane rearrangements, transcriptional activation, secretory membrane trafficking, G1 transition during the cell cycle and tumorigenic transformation. The cellular mechanisms by which FGD1 mutations lead to the hallmark skeletal deformations of faciogenital dysplasia remain unclear, but the pathology of the disease, as well as some recent discoveries, clearly show that the protein is involved in the regulation of bone development. Two recent studies unveiled new potential functions of FGD1, in particular, its involvement in the regulation of the formation and function of invadopodia and podosomes, which are cellular structures devoted to degradation of the extracellular matrix in tumour and endothelial cells. Here, we discuss the hypothesis that FGD1 might be an important regulator of events controlling extracellular matrix remodelling and possibly cell invasion in physiological and pathological settings. Additionally, we focus on how studying the cell biology of FGD1 might help us to connect the dots that link CDC42 signalling with remodelling of the extracellular matrix (ECM) in physiology and complex diseases, while, at the same time, furthering our understanding of the pathogenesis of faciogenital dysplasia. © 2012. Source


Caldieri G.,Tumour Cell Invasion Laboratory | Caldieri G.,FIRC Institute of Molecular Oncology | Capestrano M.,Tumour Cell Invasion Laboratory | Bicanova K.,Tumour Cell Invasion Laboratory | And 5 more authors.
European Journal of Cell Biology | Year: 2012

Invadopodia are proteolytically active protrusions formed by invasive tumoral cells when grown on an extracellular matrix (ECM) substratum. A current challenge is to understand how proteolytic activity is so precisely localised at discrete sites of the plasma membrane to produce focalised ECM degradation at invadopodia. Indeed, a number of components including metalloproteases need to be directed to invadopodia to ensure proper segregation of proteolytic activities.We recently found invadopodia to feature the properties of cholesterol-rich membrane domains (a.k.a. lipid drafts) and that ECM degradation depends on the tight control of cholesterol homeostasis. Since apically directed polarised sorting and transport in epithelial cells relies on segregation of proteins into lipid rafts at the Golgi complex, we hypothesised that invadopodia-dependent ECM degradation might also rely on lipid raft-dependent polarised transport routes.To investigate this issue we undertook a three-pronged approach. First, we found that microtubule depolymerisation, which is known to disrupt polarised transport in polarised cells, strongly inhibited invadopodia formation, while not affecting overall protein transport. In the second approach we found that glycosylphosphatidylinositol-anchored green fluorescent protein (an apical model protein), but not vesicular stomatitis virus G-protein or influenza virus hemagglutinin (both model basolateral model cargoes), was transported to sites of ECM degradation. Finally, RNAi-mediated knock-down of proteins known to specifically regulate polarised apical or basolateral transport in epithelial cells, such as caveolin 1 and annexin XIIIB or clathrin, respectively, demonstrated that the selective inhibition of the apical, but not the basolateral, transport route impairs invadopodia formation and ECM degradation.Taken together, our findings suggest that invadopodia are apical-like membrane domains, where signal transduction and local membrane remodelling events might be temporally and spatially confined via selective raft-dependent apical transport routes. © 2012 Elsevier GmbH. Source


Damiano L.,University of Turin | Le Devedec S.E.,Leiden University | Di Stefano P.,University of Turin | Repetto D.,University of Turin | And 12 more authors.
Oncogene | Year: 2012

We have recently shown that the adaptor protein p140Cap regulates tumor properties in terms of cell motility and growth. Here, by using the highly metastatic rat adenocarcinoma cell line MTLn3-epidermal growth factor receptor (EGFR), we assess the role of p140Cap in metastasis formation. Orthotopic transplantation of MTLn3-EGFR cells over-expressing p140Cap in Rag2 -/- γ c -/- mice resulted in normal primary tumor growth compared with the controls. Strikingly, p140Cap over-expression causes an 80% inhibition in the number of lung metastases. p140Cap over-expressing cells display a 50% reduction in directional cell migration, an increased number and size of focal adhesions, and a strong impairment in the ability to invade in a 3D matrix. p140Cap over-expression affects EGFR signaling and tyrosine phosphorylation of cortactin in response to EGF stimulation. Intriguingly, p140Cap associates with cortactin via interaction with its second proline-rich domain to the cortactin SH3 domain. The phosphomimetic cortactin tyrosine 421 mutant rescues migration and invasive properties in p140Cap over-expressing cells. Taken together, these data demonstrate that p140Cap suppresses the invasive properties of highly metastatic breast carcinoma cells by inhibiting cortactin-dependent cell motility. © 2012 Macmillan Publishers Limited All rights reserved. Source

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