LIVER GROWTH FACTOR: GENE CLONING AND EXPRESSION

THE HUMAN CDNA FOR HEPATOPOIETIN A WILL BE ISOLATED, USING PUBLISHED SEQUENCE DATA, AND WILL BE INSERTED INTO A RETROVIRAL VECTOR CAPABLE OF EXPRESSING THE GENE IN MAMMALIAN CELLS. THE VECTOR WILL BE INTRODUCED INTO A VARIETY OF MAMMALIAN CELLS, INCLUDING FIBROBLASTS AND HEPATOCYTES, TO MEASURE THE LEVELS OF EXPRESSION OF THE INTRODUCED HEPATOPOIETIN A GENE. BIOLOGICAL ACTIVITY, AS WELL AS THE PHYSICAL PROPERTIES OF THE PRODUCT, WILL BE USEDTO MONITOR EXPRESSION OF THE ACTIVE PRODUCT. BECAUSE HEPATOPOIETIN A IS NOW KNOWN TO BE POSTTRANSITIONALLY MODIFIED BY GLYCOSYLATION, EXPRESSION IN MAMMALIAN CELLS WILL BE USED, RATHER THAN ATTEMPTING TO EXPRESS THE PRODUCT IN ESCHERICHIA COLI. THE PROTEIN PRODUCT, FROM AN OPTIMIZED MAMMALIAN EXPRESSION SYSTEM, WILL BE USED TO TEST THE EFFECTS OF THIS GROWTH FACTOR ON LIVER CELLS AND OTHER CELLS IN VITRO, AS WELL AS ITS EFFECTSIN VIVO IN ANIMAL SYSTEMS.