Sequential modification of the sialic acid component of cell adhesion molecules and other glycoproteins is one of the mechanisms by which normal cell differentiation proceeds. The sialyl oligosaccharides of aberrant glycoproteins have also been implicated in the pathophysiology of various diseases, including metastatic colorectal cancer. The synthesis of each sialyl linkage is catalyzed by a specific sialyltransferase and fluctuations in the activities of these enzymes may therefore play a critical role in such fundamental biological processes as differentiation and metastatic behavior. Towards an eventual understanding of the mechanisms that coordinate expression of the different sialyltransferases in normal and diseased tissues, objectives of this application are: to express cDNAs for human sialyltransferases in eucaryotic cells and delineate the sialyl linkages that are catalyzed by the different encoded proteins; to perform a tissue survey of human sialyltransferase mRNAs; to compare sialyltransferase mRNA expression and enzyme activity in a panel of human cancer cell lines; and to investigate in the same panel of cell lines the regulatory effects of agents that are likely to influence levels of sialyltransferase expression. Chinese hamster ovary cells will be transfected with constructs of the sialyltransferase cDNAs in plasmid vectors. The activities of expressed sialyltransferases will be characterized by determination of their acceptor substrates and analysis of the oligosaccharides that are synthesized. The tissues survey will be conducted in human surgical specimens through Northern hybridization analysis and S1 protection assays. The panel of cancer cells will include dexamethasone, n-butyrate, retinoic acid and thyroid hormone. The results will form the basis for an understanding of the determinants of the activities of sialyltransferases and future studies of the biological functions of sialylation.
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