The glycosaminoglycan components of proteoglycans are biosynthesized and modified in the golgi apparatus by highly organized carbohydrate transfer enzymes and sulfotransferases. The purpose of this project is to investigate the functional organization and subcellular localization of these enzyme complexes. Brefeldin A is a chemical which specifically blocks anterograde protein transport within the golgi apparatus. It was used to disrupt the normal biosynthetic processes for adding glycosaminoglycan chains onto proteoglycans. When ovarian granulosa cells were treated with Brefeldin A, dermatan sulfate proteoglycan synthesis was abolished whereas heparan sulfate proteoglycan synthesis was only partially inhibited, suggesting that dermatan sulfate and heparan sulfate assembly on proteoglycans occurs in different subcellular compartments. Topics of present interest include: (1) the use of xylosides as glycosaminoglycan initiators to determine if the galactosyl transferases which synthesize the linkage region can produce (gal)2 xyloside in the presence of a brefeldin A block; (2) determine which core proteins are substituted with heparan sulfate in the presence of a brefeldin A block; and (3) determine the effects of this compound on hyaluronic acid synthesis.
