Cell-cell and cell-substratum adhesive interactions are critical to normal morphogenesis and maitenance of tissue architecture. Alterations in these interactions have been documented in malignant cells and in abnormal development. Using broad spectrum antisera which perturb adhesion in mouse mammary tumor epitherlial cells (MMTE), we have purified a complex of cell-substratum adhesion-related integral membrane glycoproteins of 120-160 Kd (called GP120-160) and an 80 Kd soluble fragment of a 120 Kd cell-cell adhesion-related surface membrane glycoprotein that we have called cell-CAM 120/80. Antisera against these glycoproteins interfere with adhesive interactions of preimplantation mouse embryos as well as MMTE. In this application we propose to (1) produce a bank of specific antisera and monoclonal antibodies against these adhesion glycoproteins that will recognize various regions of these adhesion glycoproteins in their native and denatured forms; (2) localize the adhesion glycoproteins in developing mouse embryos and mature tissue using immunofluorescence and immunoelectron microscopy; (3) characterize the peptide structures, phosphorylation state and oligosaccharide composition of these glycoproteins; (4) study the organization of these glycoproteins in the surface membrane asking in particular whether they are transmembrane and how they interact within the membrane; (5) if the adhesion glycoproteins are transmembrane, determine whether they interact with the cytoskeleton using microinjection of antibodies against adhesion glycoproteins and looking for changes in the distribution of the cytoskeleton; (6) purify a second cell-cell adhesion molecule which we have identified in early mouse embryos, using approaches similar to those we used previously to successfully isolate cell-CAM 120/80 and GP120-160; and (7) clone the genes coding for these glycoproteins. As a result of these studies we will increase significantly our knowledge of how integral membrane glycoproteins mediate cell-cell and cell-substratum interactions in both cultured cells and developing embryos.
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