WWOX Gene Expression Abolishes Ovarian Cancer Tumorigenicity In vivo and Decreases Attachment to Fibronectin via Integrin α3

Charlie Gourley,Karen J Taylor,Adam J.W Paige,Barbara Kuske,Mingjun Sun,Hani Gabra,Carol Ward,David J Harrison,Morwenna Muir,Szymon Janczar,Jieqing Zhang,John F Smyth

doi:10.1158/0008-5472.can-08-2974

Charlie Gourley, Karen J Taylor + Show 10 more

Open Access

https://doi.org/10.1158/0008-5472.can-08-2974

Copy DOI

Abstract

The WW domain-containing oxidoreductase (WWOX) gene is located at FRA16D, a common fragile site involved in human cancer. Targeted deletion of Wwox in mice causes increased spontaneous tumor incidence, confirming that WWOX is a bona fide tumor suppressor gene. We show that stable transfection of WWOX into human PEO1 ovarian cancer cells, containing homozygous WWOX deletion, abolishes in vivo tumorigenicity, but this does not correlate with alteration of in vitro growth. Rather, WWOX restoration in PEO1, or WWOX overexpression in SKOV3 ovarian cancer cells, results in reduced attachment and migration on fibronectin, an extracellular matrix component linked to peritoneal metastasis. Conversely, siRNA-mediated knockdown of endogenous WWOX in A2780 ovarian cancer cells increases adhesion to fibronectin. In addition, whereas there is no WWOX-dependent difference in cell death in adherent cells, WWOX-transfected cells in suspension culture display a proapoptotic phenotype. We further show that WWOX expression reduces membranous integrin alpha(3) protein but not integrin alpha(3) mRNA levels, and that adhesion of PEO1 cells to fibronectin is predominantly mediated through integrin alpha(3). We therefore propose that WWOX acts as an ovarian tumor suppressor by modulating the interaction between tumor cells and the extracellular matrix and by inducing apoptosis in detached cells. Consistent with this, the suppression of PEO1 tumorigenicity by WWOX can be partially overcome by implanting these tumor cells in Matrigel. These data suggest a possible role for the loss of WWOX in the peritoneal dissemination of human ovarian cancer cells.

Highlights

The WWOX gene extends over >1 Mb and spans a common fragile site that has been implicated in human tumorigenesis [1]
We propose that WWOX suppresses the tumorigenicity and regulates the peritoneal dissemination of ovarian cancer cells by modulating the interaction of tumor cells with the extracellular matrix (ECM) and promoting apoptosis in detached cells
Considering the results above where we show decreased cellular adhesion to fibronectin in WWOX-expressing ovarian cancer cells, together with enhanced in vivo survival of these cells when xenografted in the presence of Matrigel, we repeated the apoptosis assay using poly(2-hydroxyethyl methacrylate)–coated plates to prevent cellular adhesion

Summary

Introduction

The WWOX gene extends over >1 Mb and spans a common fragile site that has been implicated in human tumorigenesis [1]. WWOX encodes a 46-kDa protein containing two WW domains and an oxidoreductase domain [2, 3]. WWOX is homozygously deleted in a number of human cancers [4]. Decreased or aberrant WWOX expression has been shown in multiple tumor. Induction of apoptosis has been proposed as the possible mechanism of WWOX tumor suppression. The same group have shown that WWOX can directly bind p73 protein via its WW domains, resulting in enhanced p73-mediated apoptosis [16]. Chang and colleagues [17] reported that WWOX can bind the p53 tumor suppressor protein and enhance the apoptotic response to tumor necrosis factor. Suppression of tumorigenicity in breast cancer was not associated with increased apoptosis [10], suggesting that other mechanisms and contexts of WWOX tumor suppression exist

Methods

Results

Conclusion