INTRACELLULAR-TRANSPORT AND PROCESSING OF THE MARBURG VIRUS SURFACE PROTEIN IN VERTEBRATE AND INSECT CELLS

The surface protein (GP) of Marburg virus (MEG) is synthesized as a 90 -kDa precursor protein which is cotranslationally modified by the addi tion of high-mannose sugars (140 kDa). This step is followed by the co nversion of the N-linked sugars to endoglycosidase H (endo H)-resistan t species and the addition of O-linked oligosaccharides leading to a m ature protein of 170-200 kDa approximately 30 min after pulse labeling . The mature form of GP is efficiently transported to the plasma membr ane. GP synthesized using the T7 polymerase-driven vaccinia virus expr ession system was transported with essentially the same kinetics as th e authentic GP. However, the protein that is shown to appear 30 min af ter pulse labeling at the plasma membrane was slightly smaller (160 kD a) than GP incorporated into the virions (170 kDa). Using a recombinan t baculovirus, GP was expressed at high levels in insect cells. Three different species could be identified: a 90-kDa unglycosylated GP loca lized in the cytoplasm and two 140-kDa glycosylated proteins. Characte rization of the glycosylated GPs revealed that processing of the oligo saccharides of GP was less efficient in insect cells than in mammalian cells. The majority of GP remained endo H sensitive containing high-m annose type N-linked glycans, whereas only a small fraction became end o H resistant carrying processed N-glycans and O-glycans. Tunicamycin treatment of the GP-expressing cells demonstrated that N-glycosylation is essential for the transport of the MBG surface protein. (C) 1996 A cademic Press, Inc.