ORIGINAL ARTICLE Expression of inappropriate cadherins by epithelial tumor cells promotes endocytosis and degradation of E-cadherin via competition for p120 ctn M Maeda 1,5 , E Johnson 1,5 , SH Mandal 2,5 , KR Lawson 1,3 , SA Keim 4 , RA Svoboda 1 , S Caplan 2,3 , JK Wahl III 1,4 , MJ Wheelock 1,2,3,4 and KR Johnson 1,2,3,4 1 Department of Oral Biology, University of Nebraska Medical Center, Omaha, NE, USA; 2 Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE, USA; 3 Eppley Cancer Center, University of Nebraska Medical Center, Omaha, NE, USA and 4 Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical Center, Omaha, NE, USA Cadherin cell–cell adhesion proteins play an important role in modulating the behavior of tumor cells. E-cadherin serves as a suppressor of tumor cell invasion, and when tumor cells turn on the expression of a non-epithelial cadherin, they often express less E-cadherin, enhancing the tumorigenic phenotype of the cells. Here, we show that when A431 cells are forced to express R-cadherin, they dramatically downregulate the expression of endogenous E- and P-cadherin. In addition, we show that this downregulation is owing to increased turnover of the endogenous cadherins via clathrin-dependent endocytosis. p120 ctn binds to the juxtamembrane domain of classical cadherins and has been proposed to regulate cadherin adhesive activity. One way p120 ctn may accomplish this is to serve as a rheostat to regulate the levels of cadherin. Here, we show that the degradation of E-cadherin in response to expression of R-cadherin is owing to competition for p120 ctn . Oncogene (2006) 25, 4595–4604. doi:10.1038/sj.onc.1209396; published online 19 June 2006 Keywords: cadherin; endocytosis; p120 ctn Introduction Cells undergo a transition from an epithelial phenotype to a more mesenchymal phenotype to set in motion a number of normal developmental processes, including gastrulation in the vertebrate embryo (Thiery, 2002). An important aspect of this epithelial to mesenchymal transition is the loss of E-cadherin expression and an increase in the expression of non-epithelial cadherins, such as N-cadherin, cadherin-11 or R-cadherin. For example, expression of R-cadherin is necessary for the epithelial to mesenchymal transition that occurs in developing kidneys (Dahl et al., 2002). It has been suggested that the epithelial to mesenchymal transition that occurs in some tumor cells as they become invasive is a recapitulation of the normal processes that occur during development (Thiery, 2002). Our lab and others have shown that the expression of N-cadherin by epithelial tumor cells results in an epithelial to mesench- ymal transition that is accompanied by increased motility and invasion (Islam et al., 1996; Nieman et al., 1999b; Hazan et al., 2000; Suyama et al., 2002). In addition, non-epithelial cadherins, including N- cadherin, R-cadherin and cadherin-11, are expressed by some tumors in situ, and their expression has been correlated with high tumor grade and with an aggres- sive, metastatic phenotype (Islam et al., 1996; Pishvaian et al., 1999; Bussemakers et al., 2000; Giroldi et al., 2000; Tomita et al., 2000; Feltes et al., 2002). The dynamic nature of the adherens junctional complex allows epithelial cells to make and break connections with their neighbors as needed. One means cells use to ensure the dynamic nature of this junction is to continuously endocytose and recycle their cadherin/ catenin complexes (Le et al., 1999, 2002; Akhtar and Hotchin, 2001; Palacios et al., 2001; Xiao et al., 2003b). Various mechanisms and pathways have been proposed for cadherin recycling. It has been shown that confluent monolayers of Madin–Darby canine kidney (MDCK) cells constantly recycle a pool of detergent-soluble E- cadherin through an endocytic pathway that is clathrin dependent (Le et al., 1999). These authors further showed that semiconfluent cells have an even higher proportion of recycling cadherin and that recycling of E- cadherin in MDCK cells is regulated by protein kinase C (Le et al., 2002). Other investigations have shown that isolated cells, which cannot make contact because they have been plated at very low density, have increased cadherin recycling that is clathrin independent (Paterson et al., 2003). Others have shown that when Src- transfected MDCK cells undergo an epithelial to mesenchymal transition, E-cadherin is internalized and routed to the lysosome for degradation, rather than being recycled, thus ensuring that contacts do not reform (Palacios et al., 2005). In addition, when cells Received 2 June 2005; revised 8 December 2005; accepted 9 December 2005; published online 19 June 2006 Correspondence: Professor MJ Wheelock, The University of Nebraska Medical Center, 987696 Nebraska Medical Center, Omaha, NE 68198- 7696, USA. E-mail: mwheelock@unmc.edu 5 These authors contributed equally to this work. Oncogene (2006) 25, 4595–4604 & 2006 Nature Publishing Group All rights reserved 0950-9232/06 $30.00 www.nature.com/onc