MOLECULAR BIOLOGY & FUNCTION OF PLATELET MEMBRANE GP130

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Of the 40 or more proteins visible on a two-dimensional SDS gel of platelet membrane proteins, only a handful have names and assigned functions. Most of the currently characterized glycoproteins, including membrane glycoprotein (GP) Ib and the GPIIb-IIIa complex, are missing or defective in a particular pathological state, a feature that has greatly aided in delineating their structure and function. The recent advent of recombinant DNA technology, however, has made it possible to study many other previously uncharacterized and potentially very important membrane proteins. We have used these techniques to investigate membrane glycoproteins that are shared by platelets and endothelial cells, and have obtained a number of interesting complementary DNA (cDNA) clones. The purpose of the study is to characterize completely a newly described membrane glycoprotein, termed GP130, that we have identified in human platelets and umbilical vein endothelial cells. A combination of molecular biological, biochemical, and immunological techniques will be used to provide a detailed understanding of the structure and function of this molecule. To complete these objectives, the following studies will be conducted. First, the complete nucleotide sequence of full-length cDNA clones that encode GP130 will be determined. These clones will permit the amino acid sequence of the glycoprotein to be deduced. Second, the cDNA clones will be used as hybridization probes to characterize the size, chromosomal organization, chromosomal location, and number of genes that encode GP130. Finally, the role of GP130 in platelet function will be investigated. Polyclonal and monoclonal antibodies specific for GP130 will be produced and used to gain insight into the precise subcellular location and function of this membrane glycoprotein in human platelets. The possible association of GP130 with other membrane components or with the underlying cytoskeleton will also be explored. Biochemical properties, including limited amino acid sequence data, as well as the degree of glycosylation of GP130, will be analyzed. Together, these three interrelated projects constitute a research program that should yield valuable, timely new information about the detailed structure and function of a newly described platelet membrane glycoprotein. Importantly, this investigation is expected to serve as as prototype for the study of other important membrane antigens present in platelets and other cells that have not been amenable to study by more classical biochemical techniques. Studies such as these will further our knowledge of platelet membrane architecture and function, which should in turn lead to improvements in transfusion therapy, platelet storage, and our understanding and management of platelet functional disorders.