Positive regulation of migration and invasion by vasodilator-stimulated phosphoprotein via Rac1 pathway in human breast cancer cells
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.
Citations
More filters
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....
[...]
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
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
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
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]....
[...]
References
More filters
TL;DR: It is shown that Ratl fibroblasts expressing activated Val-12 Racl (Racl with valine at residue 12) display all the hallmarks of malignant transformation and that Rac is essential for transformation by Ras, indicating that oncogenic Ras drives both the Rac and MAP-kinase pathways, which cooperate to cause transformation.
Abstract: The GTPase Rac1 is a key component in the reorganization of the actin cytoskeleton that is induced by growth factors or oncogenic Ras1. Here we investigate the role of Rac1 in cell transformation and show that Rat1 fibroblasts expressing activated Val-12 Rac1 (Rac1 with valine at residue 12) display all the hallmarks of malignant transformation. In a focus-forming assay in NIH3T3 fibroblasts to measure the efficiency of transformation, we found that dominant-negative Asn-17 Rac1 inhibited focus formation by oncogenic Ras, but not by RafCAAX, a Raf kinase targeted to the plasma membrane by virtue of the addition of a carboxyterminal localization signal from K-Ras. This indicates that Rac is essential for transformation by Ras. In addition, Val-12 Rac1 synergizes strongly with RafCAAX in focus-formation assays, indicating that oncogenic Ras drives both the Rac and MAP-kinase pathways, which cooperate to cause transformation.
889 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)....
[...]
TL;DR: A method called FLAIR (fluorescence activation indicator for Rho proteins) was developed to quantify the spatio-temporal dynamics of the Rac1 nucleotide state in living cells, revealing precise spatial control of growth factor-induced Rac activation, in membrane ruffles and in a gradient of activation at the leading edge of motile cells.
Abstract: Signaling proteins are thought to be tightly regulated spatially and temporally in order to generate specific and localized effects. For Rac and other small guanosine triphosphatases, binding to guanosine triphosphate leads to interaction with downstream targets and regulates subcellular localization. A method called FLAIR (fluorescence activation indicator for Rho proteins) was developed to quantify the spatio-temporal dynamics of the Rac1 nucleotide state in living cells. FLAIR revealed precise spatial control of growth factor-induced Rac activation, in membrane ruffles and in a gradient of activation at the leading edge of motile cells. FLAIR exemplifies a generally applicable approach for examining spatio-temporal control of protein activity.
703 citations
"Positive regulation of migration an..." refers background in this paper
...Activated Rac1 localizes to the plasma membrane where it is thought to function in actin remodeling which contributes to membrane ruffling (26,27)....
[...]
TL;DR: Phosphorylation by PKA/PKG serine/threonine kinases appears to modulate Ena/VASP function within cells, although the mechanism underlying this regulation remains to be determined.
Abstract: Ena/VASP proteins are a conserved family of actin regulatory proteins made up of EVH1, EVH2 domains, and a proline-rich central region. They have been implicated in actin-based processes such as fibroblast migration, axon guidance, and T cell polarization and are important for the actin-based motility of the intracellular pathogen Listeria monocytogenes. Mechanistically, these proteins associate with barbed ends of actin filaments and antagonize filament capping by capping protein (CapZ). In addition, they reduce the density of Arp2/3-dependent actin filament branches and bind Profilin at sites of actin polymerization. Vertebrate Ena/VASP proteins are substrates for PKA/PKG serine/threonine kinases. Phosphorylation by these kinases appears to modulate Ena/VASP function within cells, although the mechanism underlying this regulation remains to be determined.
689 citations
"Positive regulation of migration an..." refers background in this paper
...We suspect that the VASP-Rac interaction is mediated by VASP binding a Pak polyproline motif because Pak contains five polyproline domains, and VASP often associates with prolinerich domains through its EVH1 domain (32)....
[...]
...The vasodilator-stimulated phosphoprotein (VASP), which was first described in human platelets, is one member of the Ena/VASP family of cytoskeletal regulatory proteins (32)....
[...]
TL;DR: Like neutrophilic leukocytes, differentiated HL-60 cells respond to chemoattractant by adopting a polarized morphology, with F-actin in a protruding pseudopod at the leading edge and contractile actin-myosin complexes at the back and sides.
689 citations
"Positive regulation of migration an..." refers background in this paper
...Xu et al showed a rapid and transient increase in Rac1 activity ~1 min after exposure to fMLP in HL-60 cells (23)....
[...]
TL;DR: Using overexpression, loss-of-function, and inhibitory approaches, it is found that Ena/VASP proteins negatively regulate fibroblast motility.
486 citations
"Positive regulation of migration an..." refers background in this paper
...However, fibroblasts devoid of Ena/VASP proteins exhibit increased rates of cell motility (15)....
[...]
...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)....
[...]
...Fibroblasts devoid of Ena/VASP proteins exhibit increased rates of cell motility (15)....
[...]