Epithelial cell adhesion molecule (EpCAM) is best known as a tumor-associated protein highly expressed in carcinomas. embryonic development. Surprisingly this house is independent of the putative adhesive function of EpCAM and rather relies on a novel signaling function that operates through down-regulation of PKC activity. We display that inhibition of novel PKCs accounts entirely for the invasive phenotype induced by abnormally high levels of EpCAM as well as for its normal function in regulating cell rearrangement during early development. Introduction EpCAM has been long known as a tumor-associated antigen highly expressed in a variety of carcinomas (Koprowski et al. 1979 It is used like a marker for aggressive tumors and has been considered as a potential target for immunotherapy (Osta et al. 2004 In human being and mouse EpCAM is definitely indicated in embryonic epithelia but the levels usually drop as cells reach terminal differentiation (Trzpis et al. 2007 Enhanced manifestation of EpCAM is definitely associated with active proliferations of neoplastic or normal cells (de Boer et al. 1999 The protein can act as a homophilic Ca2+-self-employed cell-cell adhesion molecule (Litvinov et al. 1994 It is not structurally related to any of the major families of CAMs but a potential link to the actin cytoskeleton via α-actinin has been recorded (Balzar et al. 1998 Therefore it was in the beginning proposed that enhanced proliferation and migration in cells expressing high levels of EpCAM resulted from sequestering α-catenin away from E-cadherin (Litvinov et al. 1997 However a recent study has shown that EpCAM is required to maintain the integrity and Rabbit Polyclonal to LMTK3. plasticity of the zebrafish developing epidermis where it works in partial redundancy with E-cadherin to promote cell-cell adhesion (Slanchev et al. 2009 Another study indicates PNU-120596 that this enhancing effect of EpCAM on proliferation rates of carcinoma may in fact largely rely on a signaling activity of its intracellular domain name (Münz et al. 2004 Maetzel et al. 2009 This short segment can be cleaved and is then able to form a complex with FHL2 PNU-120596 β-catenin and Lef-1 that induces gene transcription of oncogenes such as C-myc and cyclins A/E. Thus the role of EpCAM in cell-cell adhesion and the relative contributions of its potential adhesive and signaling activities in morphogenesis and proliferation remain unclear. gastrulation is an established model to study morphogenetic movements. During this phase of development the embryo undergoes massive reorganization. Because there is very little cell division and no increase in total cell mass at this stage the whole process relies purely on rearrangement of preexisting tissues. In particular the ectoderm thins and expands to eventually cover the whole embryo (epiboly) while the mesoderm techniques inside the embryo through involution and migrates along the inner surface of the ectoderm (blastocoel roof [BCR]). We are particularly interested in the mechanisms that maintain the mesoderm separated from your overlying BCR which is essential for proper gastrulation to proceed. This system is also more of interest as it deals with interactions between prototypical forms of epithelial and mesenchymal tissues. The ectoderm-mesoderm boundary can be particularly well analyzed in orthologue of EpCAM in a gain-of-function screen to identify gene products that cause aberrant ectoderm-mesoderm tissue combining at gastrula stages. We show that this overexpression of EpCAM in either the ectoderm PNU-120596 or the mesoderm causes both tissues to mix. More generally we show that EpCAM levels crucially regulate movements of cells in embryonic tissues. We demonstrate that this effect is not due to an adhesive function of EpCAM but to a signaling activity including novel PKC isoforms. Results Identification of EpCAM as a promoter of cell mixing between ectoderm-mesoderm We recognized a orthologue of human EpCAM in a gain-of-function screen for gene products perturbing the ectoderm-mesoderm boundary called Brachets’ cleft. When EpCAM mRNA was injected in the dorsal region (Fig. 1 A′ green area) the embryos displayed a significant reduction of the posterior part of cleft (Fig. 1 B-B′). BLAST search revealed that has two closely related EpCAM genes. Their amino acid sequences are highly similar to each other and to EpCAM from other vertebrate species (Fig. S1). All subsequent experiments were performed using constructs based on the EpCAMa clone originally recognized in our screen. Figure 1. Identification of EpCAM as inhibitor of the.