The primary object of this proposal is to compare the membrane differentiation of normal (myeloid and lymphoid cells) with that of tumor cells, and to understand the role of carbohydrates and glycoproteins in human leukocyte differentiation. We have shown that leukosialin, a major sialoglycoprotein on leukocytes, exhibits a dramatic structural change in its O-glycans during T-cell differentiation and in leukemia and immunodeficiency. We have also isolated cDNAs and genomic DNA for leukosialin and identified a novel promoter sequence responsible for the expression of the leukosialin gene. Further studies will be to elucidate the role of O-glycans in thymocyte development by isolating thymocyte lectin that binds to immature forms of the O-glycans and to assess the role of O-glycan sialylation in myeloid cell differentiation. In addition, we will elucidate the mechanisms of unique cell type-specific expression of leukosialin by identifying the transcription factor(s) which enhance or suppress its expression, depending on cell types. Transgenic mice will also be utilized to test if ectopic expression of leukosialin gene perturbs development. We identified GalBeta1> 44GlcNAc (GlcNAc to Gal) Beta> 3-N-acetylglucos- aminyltransferase as the key enzyme in the biosynthesis of polylactosaminoglycans which are critically involved in cell-cell interaction under normal and tumorigenic conditions. Further studies will be to clone cDNAs for the Beta1> 3-N-acetylglucosaminyltransferase and determine how the abolishment of this transferase affects organ and tissue formation in transgenic mice and embryoid body formation by embryonic stem cells. The studies proposed will enable us to provide critical information how the expression of specific carbohydrates and glycoproteins containing these carbohydrates control the differentiation and development. In order to achieve this goal, we will draw all our strength in carbohydrate biochemistry, molecular biology, and cell biology, gained in the past.
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