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Journal ArticleDOI

Positive regulation of migration and invasion by vasodilator-stimulated phosphoprotein via Rac1 pathway in human breast cancer cells

01 Jan 1994-Oncology Reports (Oncol Rep)-Vol. 20, Iss: 4, pp 929-939
TL;DR: The data showed that the higher expression level of VASP contributed to a higher invasive migration capacity of human breast cancer cells which was controlled by the Rac1 pathway.
Abstract: This study aimed to investigate the role of the cytoskeleton-associated protein vasodilator-stimulated phosphoprotein (VASP) on the migration and invasion of human breast cancer cells and its relationship to Rac1 which is a member of the Rho family and has been found to be implicated in tumorigenesis, invasion and metastasis. We detected the mRNA and protein expression levels of VASP and Rac1 of the non-invasive breast cancer cell line MCF-7 as well as the invasive cell line MDA-MB-231 by RT-PCR and Western blotting. GST pull-down assay was used to examine the activity of Rac1. Accordingly, the cell invasive migration ability was analyzed in a wound-healing assay (2D) and transwell assays (3D migration and invasion). We then used VASP-siRNA to inhibit the expression of VASP in breast cancer cells in order to study the relationship between the VASP expression level and the invasive migration ability of breast cancer cells. Rac1-siRNA was used to decrease the expression of Rac1, and observe its effect on the VASP expression level together with the migration and invasion ability of MCF-7 and MDA-MB-231 cells. Our results revealed that the invasive breast cancer cell line MDA-MB-231 showed a higher Rac1 activity and VASP expression level compared with the non-invasive MCF-7. Inhibition of Rac1 or VASP by siRNA, respectively, decreased the migration and invasion ability of breast cancer cells and the transfection of Rac1 siRNA-mediated reduction of VASP expression at mRNA and protein levels. Collectively, our data showed that the higher expression level of VASP contributed to a higher invasive migration capacity of human breast cancer cells which was controlled by the Rac1 pathway.

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Citations
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Journal ArticleDOI
TL;DR: In this article, the authors observed that tumor cells produce a secretion that modifies their microenvironment to facilitate tumor angiogenesis and metastasis under hypoxia, and the secreted proteins were predominantly cytoplasmic and membrane proteins.

483 citations


Cites background from "Positive regulation of migration an..."

  • ...Up-regulated proteins include filamins (FLNA and FLNB), actin cross-linkers that anchor membrane proteins to the actin cytoskeleton (33); F11 receptor, an important regulator of tight junction assemblies (34); plectin-1, a cross-linker of actin, microtubules, and intermediate filaments (35); VASP (vasodilator-stimulated phosphoprotein), involved in invasive migration of cancer cells (36); S100A4, a metastasis promoter involved in up-regulation of matrix metalloproteases (MMPs) and down-regulation of tissue inhibitors of matrix metalloprotease (TIMPs) (37); VCL (vinculin), involved in anchoring F-actin to the membrane (38); and LGALS3 (lectin, galactoside-binding, soluble-3), a protein that cross-links Mgat-5 to the cell surface....

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Journal ArticleDOI
TL;DR: The role of GPCR mediated signal transduction and their importance in the regulation of actin remodeling leading to cell migration are reviewed.

238 citations

Journal ArticleDOI
TL;DR: It is demonstrated that EHop-016 inhibits Rac activity in the MDA-MB-435 metastatic cancer cells that overexpress Rac and exhibits high endogenous Rac activity, and holds promise as a targeted therapeutic agent for the treatment of metastatic cancers with high Rac activity.

192 citations

Journal ArticleDOI
TL;DR: This review of recent literature focuses on aspects of cell biology related to motility and metastasis, and suggests some directions for future breast cancer research.

160 citations

Journal ArticleDOI
TL;DR: VASP was defined as an oncogene of HCC pathogenesis and metastasis with the potential to serve as a prognostic biomarker after a variety of hypoxia-induced molecular mechanisms contributed to the upregulation of VASP at transcriptional and post-transcriptional levels.
Abstract: Rational: Patients with hepatocellular carcinoma (HCC) have a poor prognosis mostly due to intrahepatic as well as distal metastasis. Vasodilator-stimulated phosphoprotein (VASP), a regulator of actin cytoskeleton and cell migration, is overexpressed in HCC and correlated with its malignant features and poor prognosis. Very little is known about its function in HCC. Methods: qRT-PCR, Western blot and IHC were used to detect the VASP expression in tissues and cells. Transwell and wound healing assays were used to measure the migration and invasion of HCC cells. Immunoblotting and immunofluorescence were used for detection of epithelial-to-mesenchymal transition (EMT) progression in HCC cells. A lung metastasis mouse model was used to evaluate metastasis of HCC in vivo. The putative targets of miR-204 were disclosed by public databases and a dual-luciferase reporter assay. IP was used to show the interaction between VASP and CRKL. ChIP was used to analyze the binding of HIF-1α to VASP promoter region. Results: Our data involving both gain- and loss-of-function studies revealed that VASP activated AKT and ERK signaling and promoted HCC migration and invasion in vitro and in vivo by altering the EMT phenotype and expression of MMPs. We investigated the positive correlation between VASP and an adapter protein, CRKL. VASP dynamically co-localized at the SH3N domain of CRKL and mediated its function. Mechanistically, VASP overexpression at the transcriptional level was mediated by HIF-1α through direct binding to two hypoxia response elements (HRE) in the VASP promoter region. Furthermore, we identified hypoxia-induced down-regulation of miR-204, which functioned as the regulator of VASP overexpression at the post-transcriptional level. Also, hypoxia-activated p-Smad3 dependent TGF-β signaling indirectly promoted VASP expression. Conclusion: A variety of hypoxia-induced molecular mechanisms contributed to the upregulation of VASP at transcriptional and post-transcriptional levels. These mechanisms involved CRKL, HIF-1α, miR-204, and TGF-β activating the AKT and ERK signaling to promote EMT and expression of MMPs. Taken together, our results defined VASP as an oncogene of HCC pathogenesis and metastasis with the potential to serve as a prognostic biomarker.

100 citations


Cites background from "Positive regulation of migration an..."

  • ...VASP was involved in migfilin-mediated cell-matrix adhesions and migration; however, VASP exerted its positive modulation of migration and invasion via Rac1 in human breast cancer cells [21-23]....

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References
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Journal ArticleDOI
24 May 2001-Nature
TL;DR: 21-nucleotide siRNA duplexes provide a new tool for studying gene function in mammalian cells and may eventually be used as gene-specific therapeutics.
Abstract: RNA interference (RNAi) is the process of sequence-specific, post-transcriptional gene silencing in animals and plants, initiated by double-stranded RNA (dsRNA) that is homologous in sequence to the silenced gene. The mediators of sequence-specific messenger RNA degradation are 21- and 22-nucleotide small interfering RNAs (siRNAs) generated by ribonuclease III cleavage from longer dsRNAs. Here we show that 21-nucleotide siRNA duplexes specifically suppress expression of endogenous and heterologous genes in different mammalian cell lines, including human embryonic kidney (293) and HeLa cells. Therefore, 21-nucleotide siRNA duplexes provide a new tool for studying gene function in mammalian cells and may eventually be used as gene-specific therapeutics.

10,451 citations


"Positive regulation of migration an..." refers background in this paper

  • ...It has been found that 21 short nucleotide (nt) ds RNA molecules, known as short interfering RNAs (siRNAs), can mediate RNAi in mammalian cell lines (19)....

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Journal ArticleDOI
TL;DR: This work challenges previous assumptions about how the G1/S transition of the mammalian cell cycle is governed, helps explain some enigmatic features of cell cycle control that also involve the functions of the retinoblastoma protein (Rb) and the INK4 proteins, and changes the thinking about how either p16 loss or overexpression of cyclin D-dependent kinases contribute to cancer.
Abstract: Mitogen-dependent progression through the first gap phase (G1) and initiation of DNA synthesis (S phase) during the mammalian cell division cycle are cooperatively regulated by several classes of cyclin-dependent kinases (CDKs) whose activities are in turn constrained by CDK inhibitors (CKIs). CKIs that govern these events have been assigned to one of two families based on their structures and CDK targets. The first class includes the INK4 proteins (inhibitors of CDK4), so named for their ability to specifically inhibit the catalytic subunits of CDK4 and CDK6. Four such proteins [p16 (Serrano et al. 1993), p15 (Hannon and Beach 1994), p18 (Guan et al. 1994; Hirai et al. 1995), and p19 (Chan et al. 1995; Hirai et al. 1995)] are composed of multiple ankyrin repeats and bind only to CDK4 and CDK6 but not to other CDKs or to D-type cyclins. The INK4 proteins can be contrasted with more broadly acting inhibitors of the Cip/Kip family whose actions affect the activities of cyclin D-, E-, and A-dependent kinases. The latter class includes p21 (Gu et al. 1993; Harper et al. 1993; El-Deiry et al. 1993; Xiong et al. 1993a; Dulic et al. 1994; Noda et al. 1994), p27 (Polyak et al. 1994a,b; Toyoshima and Hunter 1994), and p57 (Lee et al. 1995; Matsuoka et al. 1995), all of which contain characteristic motifs within their amino-terminal moieties that enable them to bind both to cyclin and CDK subunits (Chen et al. 1995, 1996; Nakanishi et al. 1995; Warbrick et al. 1995; Lin et al. 1996; Russo et al. 1996). Based largely on in vitro experiments and in vivo overexpression studies, CKIs of the Cip/Kip family were initially thought to interfere with the activities of cyclin D-, E-, and A-dependent kinases. More recent work has altered this view and revealed that although the Cip/Kip proteins are potent inhibitors of cyclin Eand A-dependent CDK2, they act as positive regulators of cyclin Ddependent kinases. This challenges previous assumptions about how the G1/S transition of the mammalian cell cycle is governed, helps explain some enigmatic features of cell cycle control that also involve the functions of the retinoblastoma protein (Rb) and the INK4 proteins, and changes our thinking about how either p16 loss or overexpression of cyclin D-dependent kinases contribute to cancer. Here we focus on the biochemical interactions that occur between CKIs and cyclin Dand E-dependent kinases in cultured mammalian cells, emphasizing the manner by which different positive and negative regulators of the cell division cycle cooperate to govern the G1-to-S transition. To gain a more comprehensive understanding of the biology of CDK inhibitors, readers are encouraged to refer to a rapidly emerging but already extensive literature (for review, see Elledge and Harper 1994; Sherr and Roberts 1995; Chellappan et al. 1998; Hengst and Reed 1998a; Kiyokawa and Koff 1998; Nakayama 1998; Ruas and Peters 1998).

6,076 citations


"Positive regulation of migration an..." refers background in this paper

  • ...As a significant member of the Rho family, Rac1 has been implicated in tumorigenesis (2), tumor angiogenesis (3), invasion and metastasis (4), cell-cycle control and apoptosis (5)....

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Journal ArticleDOI
TL;DR: In this review, functions of small G proteins and their modes of activation and action are described.
Abstract: Small GTP-binding proteins (G proteins) exist in eukaryotes from yeast to human and constitute a superfamily consisting of more than 100 members. This superfamily is structurally classified into at least five families: the Ras, Rho, Rab, Sar1/Arf, and Ran families. They regulate a wide variety of cell functions as biological timers (biotimers) that initiate and terminate specific cell functions and determine the periods of time for the continuation of the specific cell functions. They furthermore play key roles in not only temporal but also spatial determination of specific cell functions. The Ras family regulates gene expression, the Rho family regulates cytoskeletal reorganization and gene expression, the Rab and Sar1/Arf families regulate vesicle trafficking, and the Ran family regulates nucleocytoplasmic transport and microtubule organization. Many upstream regulators and downstream effectors of small G proteins have been isolated, and their modes of activation and action have gradually been elucidated. Cascades and cross-talks of small G proteins have also been clarified. In this review, functions of small G proteins and their modes of activation and action are described.

2,520 citations


Additional excerpts

  • ...progression and cell-cell adhesion (1)....

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Journal ArticleDOI
07 Oct 1999-Nature
TL;DR: Pure components of the actin cytoskeleton are used to reconstitute sustained movement in Listeria and Shigella in vitro and have implications for the understanding of the mechanism of actin-based motility in cells.
Abstract: Actin polymerization is essential for cell locomotion and is thought to generate the force responsible for cellular protrusions. The Arp2/3 complex is required to stimulate actin assembly at the leading edge in response to signalling. The bacteria Listeria and Shigella bypass the signalling pathway and harness the Arp2/3 complex to induce actin assembly and to propel themselves in living cells. However, the Arp2/3 complex alone is insufficient to promote movement. Here we have used pure components of the actin cytoskeleton to reconstitute sustained movement in Listeria and Shigella in vitro. Actin-based propulsion is driven by the free energy released by ATP hydrolysis linked to actin polymerization, and does not require myosin. In addition to actin and activated Arp2/3 complex, actin depolymerizing factor (ADF, or cofilin) and capping protein are also required for motility as they maintain a high steady-state level of G-actin, which controls the rate of unidirectional growth of actin filaments at the surface of the bacterium. The movement is more effective when profilin, alpha-actinin and VASP (for Listeria) are also included. These results have implications for our understanding of the mechanism of actin-based motility in cells.

959 citations


"Positive regulation of migration an..." refers background in this paper

  • ...Furthermore, the addition of VASP resulted in an increase in bacterial speed in the in vitro reconstitution assays of Listeria motility (34)....

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  • ...That VASP can affect the cell movements of bacteria, mouse melanoma cells, human platelet, fibroblasts and neurons by regulating cytoskeleton has been detected (14,15,34,35)....

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Journal ArticleDOI
TL;DR: A spatial inventory of the many molecular players in this dynamic domain of the actin cytoskeleton is given in order to highlight the open questions and the challenges ahead.

939 citations


"Positive regulation of migration an..." refers background in this paper

  • ...During cell migration, activated Rac and Cdc42 induce reorganization of the actin cytoskeleton at the leading edge (6)....

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