Intracellular synthesis of Epstein-Barr virus membrane antigen gp350/220. Inhibitory effect of monensin on its expression.
We have defined the intracellular expression and localization of gp350/220, one of the Epstein-Barr virus (EBV) induced membrane antigens, on 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and n-butyrate-treated P3HR-1 cells. 1B6 monoclonal antibody (mAb) immunoprecipitated gp350/220 from [35S]-methionine-labeled cells, as confirmed with other mAbs (2L10, 72A1, and C1), to the same membrane antigen. The appearance of gp350/220 was observed about 14 h after TPA and n-butyrate activation and reached a maximal level at about 48 h. 1B6 mAb membrane immunofluorescence-positive and cytoplasmic fluorescence-positive cells appeared progressively in cell populations at the same frequencies. Cytoplasmic immunofluorescent staining with 1B6 mAb demonstrated a paranuclear complex which was identical to a rhodamine-labeled wheat germ agglutinin-stained pattern which has been ascribed to the Golgi apparatus. We investigated the effect of monensin on gp350/220 expression and processing. Monensin at 10(-7) M significantly inhibited membrane antigen expression in the Golgi apparatus and on the cell surface, but had a negligible effect on synthesis of viral capsid antigen, early antigen, and viral DNA. The inhibition of gp350/220 with monensin was further characterized by the immunoprecipitation of gp350/220 with anti-MA-positive human sera and mAbs. Monensin treatment resulted in the accumulation of a 165-kD molecule which was judged to be a precursor of gp350/220. These results were consistent with the view that the Golgi apparatus plays an important role as a place of synthesis, processing, and maturation of gp350/220.
Bertoni
G
G
Department of Pharmacology, University of Massachusetts Medical School, Worcester.
Nguyen
Q V
QV
Humphreys
R E
RE
Sairenji
T
T
D000956:Antigens, Viral / Q000032:analysis / Q000096:biosynthesis*; D002087:Butyrates / Q000494:pharmacology; D020148:Butyric Acid; D036022:Capsid Proteins; D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology; D002470:Cell Survival / Q000187:drug effects; D003593:Cytoplasm / Q000276:immunology; D004279:DNA, Viral / Q000096:biosynthesis; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D004854:Herpesvirus 4, Human / Q000276:immunology* / Q000378:metabolism; D008985:Monensin / Q000494:pharmacology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014763:Viral Matrix Proteins
D000956:Antigens, Viral; D002087:Butyrates; D036022:Capsid Proteins; D004279:DNA, Viral; C037662:EBV-associated membrane antigen, Epstein-Barr virus; C041788:Epstein-Barr viral capsid antigen; C033305:Epstein-Barr virus early antigen; D014763:Viral Matrix Proteins; D020148:Butyric Acid; D008985:Monensin; D013755:Tetradecanoylphorbol Acetate