Intracellular transport and processing of the Marburg virus surface protein in vertebrate and insect cells.
The surface protein (GP) of Marburg virus (MBG) is synthesized as a 90-kDa precursor protein which is cotranslationally modified by the addition of high-mannose sugars (140 kDa). This step is followed by the conversion of the N-linked sugars to endoglycosidase H (endo H)-resistant species and the addition of O-linked oliosaccharides leading to a mature protein of 170-200 kDa approximately 30 min after pulse labelling. The mature form of GP is efficiently transported to the plasma membrane. GP synthesized using the T7 polymerase-driven vaccinia virus expression system was transported with essentially the same kinetics as the authentic GP. However, the protein that is shown to appear 30 min after pulse labeling at the plasma membrane was slighly smaller (160 kDa) than GP incorporated into the virions (170 kDa). Using a recombinant baculovirus, GP was expressed at high levels in insect cells. Three different species could be identified: a 90-kDa unglycosylated GP localized in the cytoplasm and two 140-kDa glycosylated proteins. Characterization of the glycosylated GPs revealed that processing of the oligosaccharides of GP was less efficient in insect cells than in mammalian cells. The majority of GP remained endo H sensitive containing high-mannose type N-linked glycans, whereas only a small fraction became endo 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.
Becker
S
S
Institut fuer Virologie der Philipps-Universitaet-Marburg, Germany. Becker@mailer.uni.marburg.de
Klenk
H D
HD
Mühlberger
E
E
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D006031:Glycosylation / Q000187:drug effects; D006367:HeLa Cells; D006596:Hexosaminidases; D006801:Humans; D037102:Lectins / Q000032:analysis; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D029024:Marburgvirus / Q000378:metabolism*; D008970:Molecular Weight; D017924:Nucleopolyhedroviruses / Q000235:genetics; D009844:Oligosaccharides / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D018411:Spodoptera; D014415:Tunicamycin / Q000494:pharmacology; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D014771:Virion
C081324:GP-protein, Marburg virus; D037102:Lectins; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; D014415:Tunicamycin; D006596:Hexosaminidases; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase