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1.
Mol Med Rep ; 13(5): 4414-20, 2016 May.
Article in English | MEDLINE | ID: mdl-27052944

ABSTRACT

The Ras-related C3 botulinum toxin substrate 1 (Rac1)-WASP-family verprolin-homologous protein-2 (WAVE2)-actin-related protein 2/3 (Arp2/3) signaling pathway has been identified to be involved in cell migration and invasion in various types of cancer cell. Cofilin­1 (CFL­1), which is regulated by the Rac1­WAVE2­Arp2/3 signaling pathway, may promote radioresistance in glioma. Therefore, the present study aimed to investigate the potential role of the Rac1­WAVE2­Arp2/3 signaling pathway in radioresistance in U251 human glioma cells and elucidate its affect on CFL­1 expression. Western blot analysis was performed to evaluate the protein expression of CFL­1. In the present study, Rac1 was inhibited by NSC 23766, WAVE2 was inhibited by transfection with short hairpin (sh)RNA­WAVE2 using Lipofectamine™ 2000 and Arp2/3 was inhibited by CK­666. Cell viability was measured using the 3­(4,5­dimethylthiazol­2­yl)-2,5­diphenyltetrazolium bromide assay, the cell migration ability was examined by a wound­healing assay, and the cell invasion ability was assessed using a Transwell culture chamber system. The results showed that inhibition of the Rac1­WAVE2­Arp2/3 signaling pathway using NSC 23766, shRNA­WAVE2 or CK­666 reduced the cell viability, migration and invasion abilities in U251 human glioma cells, concordant with a reduced expression of CFL­1. Furthermore, the expression of CFL­1 was significantly increased in radioresistant U251 glioma cells when compared with normal U251 human glioma cells. These findings indicate that inhibition of the Rac1­WAVE2­Arp2/3 signaling pathway may promote radiosensitivity, which may partially result from the downregulation of CFL­1 in U251 human glioma cells.


Subject(s)
Actin-Related Protein 2-3 Complex/biosynthesis , Cofilin 1/biosynthesis , Down-Regulation/radiation effects , Gamma Rays , Gene Expression Regulation, Neoplastic/radiation effects , Glioma/metabolism , Neoplasm Proteins/biosynthesis , Radiation Tolerance , Signal Transduction/radiation effects , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , rac1 GTP-Binding Protein/biosynthesis , Actin-Related Protein 2-3 Complex/genetics , Cell Line, Tumor , Cofilin 1/genetics , Glioma/genetics , Glioma/pathology , Glioma/radiotherapy , Humans , Neoplasm Proteins/genetics , Wiskott-Aldrich Syndrome Protein Family/genetics , rac1 GTP-Binding Protein/genetics
2.
Oncogene ; 35(3): 333-43, 2016 Jan 21.
Article in English | MEDLINE | ID: mdl-25823022

ABSTRACT

AAA domain containing 3A (ATAD3A) is an integral mitochondrial membrane protein with unknown function, although we now show that high-level expression is associated with poor survival in breast cancer patients. Using a mass spectrometry approach we have demonstrated that ATAD3A interacts with the WASF3 metastasis-promoting protein. Knockdown of ATAD3A leads to decreased WASF3 protein levels in breast and colon cancer cells. Silencing ATAD3A also results in loss of both cell anchorage-independent growth and invasion and suppression of tumor growth and metastasis in vivo using immuno-compromised mice. HSP70 is responsible for stabilizing WASF3 in the cytoplasm, but inactivation of HSP70 does not lead to the loss of WASF3 stability at the mitochondrial membrane, where presumably it is protected through its interaction with ATAD3A. In response to endoplasmic reticulum (ER) stress, increases in the GRP78 protein level leads to increased WASF3 protein levels. We also show that ATAD3A was present in a WASF3-GRP78 complex, and suppression of GRP78 led to destabilization of WASF3 at the mitochondrial membrane, which was ATAD3A dependent. Furthermore, ATAD3A-mediated suppression of CDH1/E-cadherin occurs through its regulation of GRP78-mediated WASF3 stability. Proteolysis experiments using isolated mitochondria demonstrates the presence of the N-terminal end of WASF3 within the mitochondria, which is the interaction site with the N-terminal end of ATAD3A. It appears, therefore, that stabilization of WASF3 function occurs through its interaction with ATAD3A and GRP78, which may provide a bridge between the ER and mitochondria, allowing communication between the two organelles. These findings also suggest that pharmacologic inhibition of ATAD3A could be an effective therapeutic strategy to treat human cancer.


Subject(s)
Adenosine Triphosphatases/biosynthesis , Breast Neoplasms/genetics , Colonic Neoplasms/genetics , Heat-Shock Proteins/biosynthesis , Membrane Proteins/biosynthesis , Mitochondrial Proteins/biosynthesis , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , ATPases Associated with Diverse Cellular Activities , Adenosine Triphosphatases/genetics , Animals , Breast Neoplasms/pathology , Cell Line, Tumor , Cell Proliferation/genetics , Colonic Neoplasms/pathology , Endoplasmic Reticulum Chaperone BiP , Endoplasmic Reticulum Stress/genetics , Female , Gene Expression Regulation, Neoplastic , Heat-Shock Proteins/genetics , Humans , Membrane Proteins/genetics , Mice , Mitochondria/genetics , Mitochondria/pathology , Mitochondrial Proteins/genetics , Neoplasm Invasiveness/genetics , Neoplasm Invasiveness/pathology , Neoplasm Metastasis , Signal Transduction/genetics , Wiskott-Aldrich Syndrome Protein Family/genetics , Xenograft Model Antitumor Assays
3.
Sci Rep ; 5: 17694, 2015 Dec 04.
Article in English | MEDLINE | ID: mdl-26634693

ABSTRACT

Accumulating evidence suggests that podocytes are direct targets of many classic antiproteinuric drugs. The immunosuppressive drug cyclosporine A (CsA), which is a calcineurin inhibitor, is used to treat proteinuric kidney diseases. One novel mechanism by which CsA reduces proteinuria is by directly stabilizing the podocyte cytoskeleton. Previous studies showed that calcineurin can directly regulate WAVE1 within mouse striatal slices. In this study, WAVE1 was expressed in podocytes and was localized in the podocyte cell bodies and foot processes (FPs). WAVE1 expression increased in both in vivo and in vitro models of puromycin aminonucleoside (PAN)-induced podocyte injury. CsA restored WAVE1 expression and also partially rescued the disordered F-actin arrangement after PAN injury. Co-immunoprecipitation assays showed that calcineurin directly interacted with WAVE1 and regulated WAVE1 phosphorylation in podocytes. Synaptopodin is a well-characterized target of CsA. WAVE1 overexpression and synaptopodin knockdown experiments directly demonstrated that WAVE1 expression is not dependent on synaptopodin expression, and vice versa. Overexpression of WAVE1 using a WAVE1 plasmid disrupted F-actin structure and promoted podocyte migration compared with the empty vector group. Therefore, WAVE1 may be a novel molecular target for the maintenance of podocyte FPs and for antiproteinuric treatment in the future.


Subject(s)
Kidney Diseases/metabolism , Microfilament Proteins/genetics , Podocytes/metabolism , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Actins/chemistry , Actins/metabolism , Animals , Calcineurin/metabolism , Calcineurin Inhibitors/administration & dosage , Cyclosporine/administration & dosage , Gene Expression Regulation/drug effects , Humans , Kidney Diseases/pathology , Mice , Microfilament Proteins/antagonists & inhibitors , Microfilament Proteins/biosynthesis , Phosphorylation/drug effects , Podocytes/drug effects , Podocytes/pathology , Puromycin Aminonucleoside/metabolism , Visual Cortex/drug effects , Visual Cortex/metabolism , Visual Cortex/pathology , Wiskott-Aldrich Syndrome Protein Family/genetics
4.
PLoS One ; 9(10): e110627, 2014.
Article in English | MEDLINE | ID: mdl-25329315

ABSTRACT

The WAVE3 cytoskeletal protein promotes cancer invasion and metastasis. We have shown that the WAVE3-mediated activation of cancer cell invasion is due, in part, to its regulation of expression and activity of key metalloproteinases (MMPs), including MMP9, which is centrally involved in invadopodia-mediated degradation of the extracellular matrix (ECM). MMP9 is also a major NFκB target gene, suggesting a potential linkage of WAVE3 to this pathway, which we sought to investigate. Mechanistically, we found that loss of WAVE3 in cancer cells leads to inhibition of NFκB signaling as a result of a decrease in the nuclear translocation of NFκB and therefore loss of activation of NFκB target genes. Conversely, overexpression of WAVE3 was sufficient to enhance NFκB activity. Both pharmacologic and genetic manipulations of NFκB effector molecules show that the biological consequence of loss of WAVE3 function in the NFκB pathway result the inhibition of invadopodia formation and ECM degradation by cancer cells, and these changes are a consequence of decreased MMP9 expression and activity. Loss of WAVE3 also sensitized cancer cells to apoptosis and cell death driven by TNFα, through the inhibition of the AKT pro-survival pathway. Our results identify a novel function of WAVE3 in NFκB signaling, where its activity is essential for the regulation of invadopodia and ECM degradation. Therefore, targeted therapeutic inhibition of WAVE3 will sensitize cancer cells to apoptosis and cell death, and suppress cancer invasion and metastasis.


Subject(s)
Matrix Metalloproteinase 9/biosynthesis , NF-kappa B/genetics , Neoplasms/genetics , Wiskott-Aldrich Syndrome Protein Family/genetics , Extracellular Matrix/genetics , Extracellular Matrix/metabolism , Gene Expression Regulation, Neoplastic , Humans , MCF-7 Cells , Matrix Metalloproteinase 9/genetics , NF-kappa B/metabolism , Neoplasm Invasiveness/genetics , Neoplasm Metastasis , Neoplasms/pathology , Podosomes/genetics , Signal Transduction/genetics , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/genetics , Wiskott-Aldrich Syndrome Protein Family/biosynthesis
5.
Anticancer Res ; 34(5): 2185-94, 2014 May.
Article in English | MEDLINE | ID: mdl-24778020

ABSTRACT

BACKGROUND: WAVE2 plays a crucial role in actin polymerisation and cell migration. We aimed to investigate the expression and cellular functions of WAVE2 in human gastric cancer (GC). MATERIALS AND METHODS: The level of WAVE2 was determined using quantitative PCR (Q-PCR) in a cohort of human gastric tissues. Expression of WAVE2, ARP2, NWASP, ROCK1 and ROCK2 was examined using RT-PCR in paired tissues. WAVE2 and ARP2 protein co-expression was examined. Anti-WAVE2 transgene ribozymes were constructed and transiently transfected into human GC cells. RESULTS: Down-regulation of WAVE2 expression in GC was significantly correlated with lymph node metastasis. WAVE2 was positively correlated with E-cadherin and negatively with TWIST. Immunohistochemically, WAVE2 and ARP2 were not co-expressed in serial mirror sections. In vitro, WAVE2 knockdown was shown to increase cell motility, whilst ROCK inhibitor treatment reduced this effect in HGC27 cells. CONCLUSION: WAVE2 is down-regulated in GC and loses its metastatic role in GC. Knockdown of WAVE2 could increase metastatic potential by promoting the growth, invasiveness, motility, adhesiveness and suppressing EMT (epithelial-mesenchymal transition) of GC cells.


Subject(s)
Cell Movement , Lymphatic Metastasis/pathology , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Adult , Aged , Aged, 80 and over , Cell Movement/genetics , Down-Regulation , Epithelial-Mesenchymal Transition/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , Immunohistochemistry , Lymphatic Metastasis/genetics , Male , Middle Aged , Neoplasm Invasiveness/genetics , Neoplasm Invasiveness/pathology , Reverse Transcriptase Polymerase Chain Reaction , Stomach Neoplasms/genetics , Transfection , Young Adult
6.
Biochem J ; 452(1): 45-55, 2013 May 15.
Article in English | MEDLINE | ID: mdl-23452202

ABSTRACT

During cell spreading, mammalian cells migrate using lamellipodia formed from a large dense branched actin network which produces the protrusive force required for leading edge advancement. The formation of lamellipodia is a dynamic process and is dependent on a variety of protein cofactors that mediate their local regulation, structural characteristics and dynamics. In the present study, we show that mRNAs encoding some structural and regulatory components of the WAVE [WASP (Wiskott-Aldrich syndrome protein) verprolin homologous] complex are localized to the leading edge of the cell and associated with sites of active translation. Furthermore, we demonstrate that steady-state levels of ArpC2 and Rac1 proteins increase at the leading edge during cell spreading, suggesting that localized protein synthesis has a pivotal role in controlling cell spreading and migration.


Subject(s)
Actin-Related Protein 2-3 Complex/chemistry , Actin-Related Protein 2-3 Complex/genetics , Cell Movement/genetics , Fibroblasts/physiology , RNA, Messenger/metabolism , Wiskott-Aldrich Syndrome Protein Family/chemistry , Wiskott-Aldrich Syndrome Protein Family/genetics , Actin-Related Protein 2-3 Complex/biosynthesis , Cell Line, Transformed , Fibroblasts/chemistry , Fibroblasts/cytology , Humans , Molecular Dynamics Simulation , Protein Biosynthesis , RNA, Messenger/biosynthesis , Wiskott-Aldrich Syndrome Protein Family/biosynthesis
7.
PLoS One ; 7(8): e42895, 2012.
Article in English | MEDLINE | ID: mdl-22952619

ABSTRACT

BACKGROUND: Breast Cancer (BC) is a heterogeneous disease comprised of at least five genetically distinct subtypes, which together form the second leading cause of cancer death in women in the United States. Within BC subtypes, those classified as Triple Negative BCs (TNBCs) exhibit dismal survival rates due to their propensity to develop distant metastases. We have identified the WAVE3 protein, which is a critical regulator of actin cytoskeleton dynamics that are required for the motility and invasion of cancer cells through its activation of the Arp2/3 complex, as a key regulator of the different steps of the invasion-metastasis cascade in BC, especially in the more aggressive TNBCs. Our published studies have also shown that elevated expression levels of WAVE3 in the TNBC cell lines directly contribute to their increased invasion and metastasis potentials both in vitro and in vivo in murine models of BC metastasis. METHODOLOGY/PRINCIPAL FINDINGS: Herein, we utilized both immunohistochemistry (IHC) of primary human BC tumors as well as quantitative real-time RT-PCR of WAVE3 in the peripheral blood of BC patients to clearly establish that WAVE3 is a predictive marker of overall BC patients' survival. High levels of WAVE3 were predictive for reduced distant recurrence-free survival as well as for decreased disease-specific mortality. Our analysis of WAVE3 expression levels in the peripheral blood of BC patients showed that WAVE3 is highly expressed in the blood of patients who developed metastatic breast cancer compared to those who did not. WAVE3 expression was also highly upregulated in the blood of BC patients with the more aggressive TNBC subtype. CONCLUSIONS: Together, these findings establish WAVE3 as a novel marker for increased risk of breast-cancer-specific mortality and for the metastatic potential of the TNBCs, and also identify WAVE3 as an attractive therapeutic target for the treatment of metastatic BC.


Subject(s)
Breast Neoplasms/metabolism , Gene Expression Regulation, Neoplastic , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Actins/metabolism , Biomarkers, Tumor/metabolism , Breast Neoplasms/pathology , Cytoskeleton/metabolism , Disease-Free Survival , Epithelial Cells/cytology , Female , Gene Expression Profiling , Humans , Immunohistochemistry/methods , Neoplasm Metastasis , Polymerase Chain Reaction/methods , Recurrence , Treatment Outcome
8.
J Cancer Res Clin Oncol ; 138(12): 2035-44, 2012 Dec.
Article in English | MEDLINE | ID: mdl-22806308

ABSTRACT

PURPOSE: To investigate matrix metalloproteinase 9 (MMP-9) and WAVE3 expression in human colorectal cancer (CRC) and to evaluate their clinical significance. METHODS: We first performed real-time PCR to evaluate mRNA expression of MMP-9 and WAVE3 in 21 pairs of fresh CRC samples matched with adjacent normal mucosa. Then, MMP-9 and WAVE3 proteins were evaluated by immunohistochemistry on CRC tissue microarrays which included 216 CRC specimens and corresponding normal colorectal mucosa, and their correlation with clinicopathological factors and overall survival after surgery was evaluated. RESULTS: Both real-time PCR and immunohistochemistry evaluation have demonstrated that MMP-9 and WAVE3 were over-expressed in colorectal cancer tissues compared with normal mucosa (p < 0.001). MMP-9 expression was significantly higher in patients with low-grade differentiation and distant metastasis (p = 0.003 and p = 0.005, respectively), and patients with MMP-9-positive expression had a poorer prognosis (p = 0.008). However, patients with WAVE3-positive expression had a better prognosis (p = 0.039) and particularly favorable prognostic factors, including non-lymph node metastasis, non-distant metastasis, and early TNM stage (p = 0.029, 0.021, and 0.003, respectively). In addition, MMP-9-negative/WAVE3-positive patients had the best overall survival (p = 0.021). In multivariate survival analysis, MMP-9 expression and combined expression status of MMP-9/WAVE3 were identified as independent prognostic factors for CRC (p = 0.046 and p = 0.019, respectively). CONCLUSIONS: Combined analysis of MMP-9 and WAVE3 has a significant value for assessing prognosis of CRC patients after surgery.


Subject(s)
Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Matrix Metalloproteinase 9/biosynthesis , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Adult , Aged , Aged, 80 and over , Cell Differentiation/physiology , Colorectal Neoplasms/genetics , Female , Humans , Immunohistochemistry/methods , Intestinal Mucosa/metabolism , Intestinal Mucosa/pathology , Male , Matrix Metalloproteinase 9/genetics , Matrix Metalloproteinase 9/metabolism , Middle Aged , Neoplasm Metastasis , Neoplasm Staging/methods , Prognosis , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Wiskott-Aldrich Syndrome Protein Family/genetics , Wiskott-Aldrich Syndrome Protein Family/metabolism , Young Adult
9.
Gynecol Oncol ; 127(1): 223-30, 2012 Oct.
Article in English | MEDLINE | ID: mdl-22721732

ABSTRACT

OBJECTIVES: Wiskott-Aldrich syndrome protein family verprolin-homologous protein 1 (WAVE1) has been shown to promote cancer invasion and metastasis. However, no evidence has been found to identify the role of WAVE1 in epithelial ovarian cancer (EOC). This study aims to determine the effect of WAVE1 expression and investigate a possible relationship between WAVE1 and prognosis in EOC. METHODS: WAVE1 protein level was measured in 223 EOC specimens by immunohistochemical staining and 46 EOC specimens by Western blot analysis. Expression of WAVE1 in ovarian cancer cell lines was evaluated by Western blot analysis and immunofluorescence. Survival analysis was performed to assess the correlation between WAVE1 expression and survival. RESULTS: Immunohistochemical staining and Western blot analysis showed that WAVE1 was overexpressed in EOC compared with samples from a non-invasive ovarian tumor and normal ovaries (P<0.05). Furthermore, expression of WAVE1 was significantly associated with advanced FIGO stage, poor grade, serum Ca-125 and residual tumor size (P<0.05). By Western blot analysis, WAVE1 expression was detected in four ovarian cancer cell lines. Immunofluorescence was performed to demonstrate WAVE1 expression in SKOV3 and 3AO cell lines. Survival analysis showed that patients with low WAVE1 staining had a significantly better survival compared to patients with high WAVE1 staining (P<0.05). In multivariate analysis, WAVE1 overexpression, advanced stage and suboptimal surgical debulking were independent prognostic factors of poor survival. CONCLUSIONS: Our present study finds that WAVE1 overexpression is associated with an unfavorable prognosis. WAVE1 is an independent prognostic factor for EOC, which suggests that it is a novel and crucial predictor for EOC metastasis.


Subject(s)
Neoplasms, Glandular and Epithelial/metabolism , Neoplasms, Glandular and Epithelial/pathology , Ovarian Neoplasms/metabolism , Ovarian Neoplasms/pathology , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Carcinoma, Ovarian Epithelial , Cell Line, Tumor , Female , Humans , Microscopy, Confocal , Middle Aged , Prognosis
10.
Urol Oncol ; 28(3): 320-7, 2010.
Article in English | MEDLINE | ID: mdl-19395286

ABSTRACT

INTRODUCTION: Wiskott-Aldrich syndrome verprolin-homologous 3 (WAVE3) belongs to Wiskott-Aldrich syndrome family proteins (WASP), which, along with other members, play a critical role in the regulation of actin polymerization and cell motility. We investigated the expression pattern and the effects of manipulating endogenous WAVE3 expression in prostate cancer cells. MATERIALS AND METHODS: Reverse transcriptase-polymerase chain reaction (RT-PCR) of prostate cell lines and immunohistochemical staining of normal and cancer specimens for WAVE3 proteins were done. WAVE3 knockdown clones were synthesized using hammerhead ribozyme transgenes and transfected by electroporation into DU-145 and PC-3 cells. The impact of WAVE3 knockdown was studied using in vitro functional assays. RESULTS: RT-PCR for WAVE3 showed strong expression in both PC-3 and DU-145 cell lines. Immunohistochemistry of prostate tissue specimens showed that the cytoplasm of cancer cells had stronger staining than normal epithelium. The mean [(+/-standard error of mean (SEM)] invading cell number for PC-3(DeltaW3R1) and PC-3(DeltaW3R2) was 5.06 (+/-0.42) and 6.33 (+/-0.19), respectively, compared with PC-3(WT) (12.27 +/- 0.42; P < 0.001). Similarly, the mean (+/-SEM) invading cell numbers for DU-145(DeltaW3R1) and DU-145(DeltaW3R2) were 10.80 (+/-1.33) and 10.20 (+/-0.86) compared with DU-145(WT) (14.80 +/- 0.24, P < 0.05). No significant differences were observed in the adhesiveness and proliferation between the knockdown mutants and the wild types. CONCLUSIONS: This is the first report on the expression patterns and the functions of WAVE3 in prostate cancer cell lines. This study shows that WAVE3 is pivotal in controlling the invasiveness of prostate cancer cells. Further work is needed to assess WAVE3 as a potential marker for predicting tumor aggressiveness.


Subject(s)
Neoplasm Invasiveness/genetics , Prostatic Neoplasms/genetics , Prostatic Neoplasms/pathology , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Blotting, Western , Cell Adhesion/genetics , Cell Line, Tumor , Cell Proliferation , Gene Expression , Gene Knockdown Techniques , Humans , Immunohistochemistry , Male , Reverse Transcriptase Polymerase Chain Reaction , Transfection , Wiskott-Aldrich Syndrome Protein Family/genetics
11.
Neuroscientist ; 13(5): 486-91, 2007 Oct.
Article in English | MEDLINE | ID: mdl-17901257

ABSTRACT

The myelin sheath can be compared to the neuronal growth cone in that the unfurled sheath looks like a giant lamellum. The authors recently tested this hypothesis by examining the importance of WAVE1, a regulator of lamellipodia formation in neurons and other cells, in myelinogenesis. They found that WAVE1 is critical for formation of oligodendrocyte lamellae and myelin sheaths. They review the regulation of WAVE1 and how WAVE1 is transported and localized to lamellipodia. Because they found that some but not all myelination was impaired by knockout of WAVE1 function, they hypothesize that other regulators of actin nucleation help oligodendrocytes produce myelin in parallel with WAVE1 function. Interestingly, they found that oligodendrocyte maturation also is disturbed with WAVE1 knockout and propose that proper localization and transport of signaling molecules relevant to the integrin signaling cascade are disrupted by loss of WAVE1 function.


Subject(s)
Actins/physiology , Active Transport, Cell Nucleus/physiology , Myelin Sheath/physiology , Wiskott-Aldrich Syndrome Protein Family/physiology , Actins/metabolism , Animals , Humans , Phosphorylation , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Wiskott-Aldrich Syndrome Protein Family/genetics
12.
Mod Pathol ; 20(3): 339-43, 2007 Mar.
Article in English | MEDLINE | ID: mdl-17277766

ABSTRACT

Breast carcinoma with a high histologic grade is highly invasive and metastatic. One reason for its irregular morphology is the formation of excessive protrusions due to assemblages of branched actin filament networks. In mammalian cells, the actin-related protein 2 and 3 (Arp2/3) complex initiates actin assembly to form lamellipodial protrusions by binding to the Wiskott-Aldrich syndrome (WASP)/WASP family verproline-homologous protein2 (WAVE2), a member of the WASP protein family. In this study, the localization Arp2 and WAVE2 in breast carcinoma was investigated to clarify whether coexpression of the two proteins is associated with histologic grade or patient outcome. Immunohistochemical staining of Arp2 and WAVE2 was performed on mirror specimens of 197 breast carcinomas, and the association between coexpression of the two proteins and clinicopathologic factors was examined. Kaplan-Meier disease-free survival and overall survival curves were analyzed to determine the prognostic significance of Arp2 and WAVE2 coexpression in breast carcinoma. Coexpression of Arp2 and WAVE2 was detected in 64 (36%) of 179 invasive ductal carcinomas and in 2 (11%) of 18 ductal carcinomas in situ, but was not detected in any adjacent non-cancerous tissue. The proportion of cancer cells expressing both Arp2 and WAVE2 was significantly higher in cases with high histologic grade (P<0.0001), and cases with lymph node metastasis (P=0.0150). The patients whose cancer cells showed such coexpression had shorter disease-free (P=0.0002) and overall survival (P=0.0122) than patients whose cancer cells expressed only one or none of Arp2 and WAVE2. Multivariate Cox regression analysis revealed that coexpression of Arp2 and WAVE2 is an independent factor for both tumor recurrence (P=0.0308) and death (P=0.0455). These results indicate that coexpression of Arp2 and WAVE2 is a significant prognostic factor that is closely associated with aggressive morphology of invasive ductal carcinoma of the breast.


Subject(s)
Actin-Related Protein 2/biosynthesis , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Breast Neoplasms/mortality , Female , Humans , Immunohistochemistry , Kaplan-Meier Estimate , Lymphatic Metastasis/pathology , Prognosis
13.
Neurosci Res ; 56(4): 459-69, 2006 Dec.
Article in English | MEDLINE | ID: mdl-17049400

ABSTRACT

The actin cytoskeleton plays a critical role in the cellular morphological changes. Its organization is essential for neurite extension and synaptogenesis under the processes of neuronal development. Recently, neural Wiskott-Aldrich syndrome protein (N-WASP) and WASP family verprolin-homologous protein (WAVE) have been identified as key molecules, which specifically participate in regulation of actin cytoskeleton through small GTPases. The functions of these factors have been investigated using cultured cells; however, in vivo developmental changes in these factors are not fully understood. In this study, we examined the expression levels and distributions of N-WASP, WAVE and their related proteins in the rat cerebral cortex and hippocampus during postnatal development. Protein levels of these factors were progressively increased during development, and actin was accumulated in membranous fractions. Immunoreactivities for these factors were widely but differentially observed in entire brain. In the developing brain, N-WASP and WAVE seemed to exist in the synapse-rich areas, such as stratum radiatum of hippocampal CA1 subfield. A similar tendency in the distributions of these factors was observed in the mature brain. Taken together, N-WASP, WAVE and their related proteins may participate in normal brain development and synaptic plasticity by regulating the actin cytoskeleton.


Subject(s)
Animals, Newborn/physiology , Cerebral Cortex/metabolism , Hippocampus/metabolism , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Wiskott-Aldrich Syndrome Protein/biosynthesis , Actins/biosynthesis , Animals , Cells, Cultured , Cerebral Cortex/growth & development , Electrophoresis, Polyacrylamide Gel , Hippocampus/growth & development , Immunoblotting , Immunohistochemistry , Neurons/metabolism , Rats , Rats, Wistar , Receptors, Presynaptic/metabolism
14.
J Immunol ; 176(11): 6576-85, 2006 Jun 01.
Article in English | MEDLINE | ID: mdl-16709815

ABSTRACT

The Wiskott-Aldrich syndrome protein (WASP) is a product of the gene defective in an Xid disorder, Wiskott-Aldrich syndrome. WASP expression is limited to hemopoietic cells, and WASP regulates the actin cytoskeleton. It has been reported that monocytes/macrophages from WASP-deficient Wiskott-Aldrich syndrome patients are severely defective in chemotaxis, resulting in recurrent infection. However, the molecular basis of such chemotactic defects is not understood. Recently, the WASP N-terminal region was found to bind to the three mammalian verprolin homologs: WASP interacting protein (WIP); WIP and CR16 homologous protein (WICH)/WIP-related protein (WIRE); and CR16. Verprolin was originally found to play an important role in the regulation of actin cytoskeleton in yeast. We have shown that WASP, WIP, and WICH/WIRE are expressed predominantly in the human monocyte cell line THP-1 and that WIP and WICH/WIRE are involved in monocyte chemotaxis. When WASP binding to verprolins was blocked, chemotactic migration of monocytes was impaired in both THP-1 cells and primary human monocytes. Increased expression of WASP and WIP enhanced monocyte chemotaxis. Blocking WASP binding to verprolins impaired cell polarization but not actin polymerization. These results indicate that a complex of WASP with mammalian verprolins plays an important role in chemotaxis of monocytes. Our results suggest that WASP and mammalian verprolins function as a unit in monocyte chemotaxis and that the activity of this unit is critical to establish cell polarization. In addition, our results also indicate that the WASP-verprolin complex is involved in other functions such as podosome formation and phagocytosis.


Subject(s)
Chemotaxis, Leukocyte/physiology , Monocytes/physiology , Wiskott-Aldrich Syndrome Protein Family/physiology , Actins/metabolism , Amino Acid Sequence , Carrier Proteins/metabolism , Cell Line, Tumor , Gene Expression Regulation/physiology , Humans , Microfilament Proteins , Molecular Sequence Data , Monocytes/metabolism , Phagocytosis , Pseudopodia/metabolism , U937 Cells , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Wiskott-Aldrich Syndrome Protein Family/genetics , Wiskott-Aldrich Syndrome Protein Family/metabolism
15.
Clin Cancer Res ; 12(8): 2449-54, 2006 Apr 15.
Article in English | MEDLINE | ID: mdl-16638851

ABSTRACT

PURPOSE: Highly invasive and metastatic cancer cells, such as adenocarcinoma of the lung cells, form irregular protrusions by assembling a branched network of actin filaments. In mammalian cells, the actin-related protein 2 and 3 (Arp2/3) complex initiates actin assembly to form lamellipodial protrusions by binding to Wiskott-Aldrich syndrome (WASP)/WASP family verproline-homologous protein 2 (WAVE2). In this study, colocalization of Arp2 and WAVE2 in adenocarcinoma of the lung was investigated to elucidate its prognostic value. EXPERIMENTAL DESIGN: Immunohistochemical staining of Arp2 and WAVE2 was done on mirror sections of 115 adenocarcinomas of the lung from pathologic stage IA to IIIA classes. Kaplan-Meier disease-free survival and overall survival curves were analyzed to determine the prognostic significance of the coexpression of Arp2 and WAVE2. RESULTS: Immunoreactivity for both Arp2 and WAVE2 was detected in the same cancer cells in 78 (67.8%) of the 115 lung cancer specimens. The proportion of cancer cells expressing both Arp2 and WAVE2 was significantly higher in cases with lymph-node metastasis (P = 0.0046), and significantly lower in bronchioloalveolar carcinomas (P < 0.0001). The patients whose cancer cells coexpressed them had a shorter disease-free survival time (P < 0.0001) and overall survival time (P < 0.0001). Multivariate Cox regression analysis revealed that coexpression of Arp2 and WAVE2 is an independent risk factor for tumor recurrence. CONCLUSIONS: Coexpression of Arp2 and WAVE2 is correlated with poorer patient outcome, and may be involved in the mechanism of cancer metastasis.


Subject(s)
Actin-Related Protein 2/biosynthesis , Adenocarcinoma/pathology , Lung Neoplasms/pathology , Wiskott-Aldrich Syndrome Protein Family/biosynthesis , Adenocarcinoma/metabolism , Aged , Disease-Free Survival , Female , Humans , Immunohistochemistry , Lung Neoplasms/metabolism , Male , Multivariate Analysis , Neoplasm Staging , Prognosis , Survival Analysis
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