Human MUC1 is a membrane glycoprotein that is expressed on the apical surface of many secretory epithelial cells. The cumulative data indicate that MUC1, with its highly conserved cytoplasmic domain and heavily glycosylated ectodomain, functions as a cell surface receptor/sensor and a modifier of epidermal growth factor (EGF) signaling. Overexpression of MUC1 in tumors clearly induces cellular transformation through aberrant signal transduction linked to the EGF receptor (EGFR/ErbB1). Conversely, EGFR phosphorylation of MUC1 enhances MUC1 binding, stabilization and nuclear targeting of [unreadable]-catenin. Although the EGFR is primarily basolateral in polarized epithelial cells, a fraction of the EGFR co-localizes with MUC1 at the apical cell surface. MUC1 and EGFR exhibit common sites for docking of cytoplasmic binding partners required for signaling, internalization and degradation. Thus, the interaction of MUC1 and EGFR plays a significant role both in stabilizing the normal integrity of polarized epithelia, and in disrupting the monolayer in neoplasia when MUC1 is overexpressed. In tumors of epithelial origin, cell polarity is compromised and overexpressed MUC1 is found on all cell surfaces and inside the cell. Recent data indicate that mis- localization of MUC1 correlates with an aggressive tumor phenotype, increased metastasis, and a poor prognosis for the cancer patient. The basis for the aberrant localization of MUC1 is unknown. We previously found that MUC1 internalization by clathrin-mediated endocytosis is slow and modulated by its glycosylation, while recycling is rapid and dependent on its S-palmitoylation. It is now clear that MUC1 is retained on the surface of MDCK cells by galectin binding, whereas the EGFR is retained on the surface of non-polarized tumor cells by binding galectin-3 that is normally secreted at the apical surface of epithelial cells. Galectins are soluble [unreadable] -galactoside-binding proteins secreted by a non-classical pathway from the cytoplasm to cross-link extracellular glycoconjugates. Therefore, we will test the hypothesis that apical targeting of MUC1 and formation of an EGFR-MUC1 complex is dependent on galectin binding to poly-N- acetyllactosamine on terminally processed glycans. We will use a combination of metabolic labeling, surface biotinylation and immunoblotting of endogenous EGFR and MUC1 and recombinant mutants in normal and glycosylation-defective MDCK cells to determine 1) the mechanism of MUC1 apical targeting in polarized epithelial cells, 2) the mechanisms that regulate MUC1 endocytosis and recycling in polarized epithelial cells, and 3) how interaction of MUC1 and the EGFR will influence EGF-dependent signaling at the apical surface of polarized epithelial cells. Our success in these studies will provide novel and fundamental insights into both MUC1-dependent signaling and mislocalization in tumors, which will contribute to a new level of understanding of MUC1 biology in human cancer. [unreadable] [unreadable] [unreadable]
NIH ExPorter
NIH ExPorter