ORIGINALARTICLE TheE-cadherin-repressedhNanos1geneinducestumorcellinvasionby upregulatingMT1-MMPexpression A Bonnomet 1,2 , M Polette 1 , K Strumane 3,4 , C Gilles 2 , V Dalstein 1 , C Kileztky 1 , G Berx 3 , F van Roy 3 , P Birembaut 1 and B Nawrocki-Raby 1 1 INSERM UMRS 514, Laboratory of Histology, IFR 53, CHU Maison Blanche, University of Reims Champagne-Ardenne, Reims, France; 2 Laboratory of Developmental and Tumor Biology, University of Lie`ge, CHU Sart-Tilman, Lie`ge, Belgium; 3 Department for Molecular Biomedical Research, VIB-Ghent University, Ghent, Belgium and 4 Department IHB, Leiden University Medical Centre, Leiden, The Netherlands In this study, we examined the role of the E-cadherin- repressed gene human Nanos1 (hNanos1) in tumor invasion process. First, our in vivo study revealed that hNanos1 mRNAs were overexpressed in invasive lung carcinomas. Moreover, hNanos1 was co-localized with MT1-MMP(membranetype1-matrixmetalloproteinase) inE-cadherin-negativeinvasivelungtumorclusters.Using an inducible Tet-on system, we showed that induction of hNanos1 expression in DLD1 cells increased their migratory and invasive abilities in a three-dimensional migration and in a modified Boyden chamber assay. Accordingly, we demonstrated that hNanos1 upregulated MT1-MMPexpressionatthemRNAandproteinlevels. Inversely, using an RNA interference strategy to inhibit hNanos1expressionininvasiveHs578T,BT549andBZR cancercells,weobservedadownregulationofMT1-MMP mRNA and protein and concomitantly a decrease of the invasive capacities of tumor cells in a modified Boyden chamber assay. Taken together, our results demonstrate that hNanos1, by regulating MT1-MMP expression, plays an important role in the acquisition of invasive propertiesbyepithelialtumorcells. Oncogene (2008) 27, 3692–3699; doi:10.1038/sj.onc.1211035; published online 28 January 2008 Keywords: hNanos1; E-cadherin; MT1-MMP; cancer; cell invasion Introduction E-cadherin, a cell surface adhesion molecule essential for maintaining the structural integrity of epithelial tissue, has been previously characterized as a tumor/ invasion suppressor. The loss of intercellular adhesion caused by dysfunction of the E-cadherin/catenin com- plex is correlated with the acquisition of an invasive phenotype (Behrens et al., 1993). The in vitro re- expression of E-cadherin by transfection in highly invasive tumor cells reduces their invasive potential (Frixen etal., 1991; Vleminckx etal., 1991; Mareel etal., 1996; Nawrocki-Raby et al., 2003). Recently, Strumane et al. (2006) have reported that E-cadherin represses human Nanos1 (hNanos1). Indeed, E-cadherin re- expression in invasive tumor cells results in the down- regulation of hNanos1, and high hNanos1 expression was described only in E-cadherin-deficient tumor cell lines. Nanos and nanos-related proteins are character- ized by the presence of a conserved C-terminal (CCHC) 2 zinc-finger RNA-binding domain (Mosquera et al., 1993). Nanos proteins are involved in the regulation of various development processes. In Drosophila, nanos regulates differentiation of the anterior-posterior body axis, primordial germ cell migration, oogenesis and spermatogenesis (Wang and Lehmann, 1991; Wang et al., 1994; Kobayashi et al., 1996; Forbes and Lehmann, 1998; Bhat, 1999; Deshpande et al., 1999). hNanos1 is abundant in germ-line stem cells (Jaruzelska et al., 2003). In tumor, it has also been demonstrated that the E-cadherin-repressed gene hNanos1 is involved in the acquisition of invasive properties by inducing disruption of cell–cell adhesion, cell dispersion and increased migratory and invasive behavior (Strumane et al., 2006). The mechanism by which hNanos1 mediates these functions is, however, not known. Several in vitro studies have shown that E-cadherin’s function as a tumor invasion suppressive resides not only in its participation in cell–cell contacts, but also in its ability to sequester to the cell membrane several co- transcriptional factors (catenins) which can themselves modulate the expression of invasion-associated genes. Among them, matrix metalloproteinases (MMPs) have recently been identified (Behrens, 2005). Indeed, in vitro and in vivo data have shown an inverse correlation between E-cadherin and MMPs (Kitadai et al., 1996; Polette etal., 1998), and the re-expression of E-cadherin has been described to repress MMP expression in invasive cancer cells (Luo et al., 1999; Ara et al., 2000; Nawrocki-Raby et al., 2003). MMPs comprise a large Received 14 June 2007; revised 24 October 2007; accepted 13 December 2007; published online 28 January 2008 Correspondence: Dr B Nawrocki-Raby, INSERM UMRS 514, Laboratory of Histology, IFR 53, CHU Maison Blanche, University of Reims Champagne-Ardenne, 45 rue Cognacq-Jay, Reims, Marne 51100, France. E-mail: beatrice.raby@univ-reims.fr Oncogene (2008) 27, 3692–3699 & 2008 Nature Publishing Group All rights reserved 0950-9232/08 $30.00 www.nature.com/onc