BREFELDIN-A AND MONENSIN ARREST CELL-SURFACE EXPRESSION OF MEMBRANE-GLYCOPROTEINS AND RELEASE OF RUBELLA-VIRUS

The maturation of rubella virus (RV) glycoproteins E2 and E1 was exami ned by using brefeldin A (BFA) and monensin. BFA, which induces the ra pid redistribution of Golgi enzymes residing in the Golgi complex into the endoplasmic reticulum (ER), was used to locate the intracellular site for the modification of carbohydrate side-chains on RV E1 and E2 proteins. The monovalent ionophore monensin, which inhibits intracellu lar transport of proteins through the ER-Golgi complex, was used to bl ock the transport of E1 and E2 glycoproteins through the Golgi complex . BFA and monensin effectively blocked the cell surface expression of RV E2 and El proteins, secretion of an anchor-free form of E2 and budd ing of RV from the plasma membrane. For O-linked glycosylation, additi on of N-acetylgalactosamine and galactose to E2 protein was found to t ake place in the medial to the trans Golgi. A dramatic change in the i ntracellular distribution of RV structural proteins was observed when transfected COS cells were treated with BFA or monensin, although the proteolytic processing of RV structural protein precursor was not affe cted. In the presence of BFA or monensin, virus release from infected Vero cells was only 0.1% of the intracellular virus, and the intracell ular virus titre decreased as well. Our results suggest that O-linked glycosylation on the E2 protein occurred in the post-ER region and the transport of RV structural proteins to the Golgi complex and post-Gol gi compartment may be a rate-limiting step in RV assembly and budding.