Expression and characterization of glycophospholipid-anchored human immunodeficiency virus type 1 envelope glycoproteins.
Four chimeric human immunodeficiency virus type 1 (HIV-1) env genes were constructed which encoded the extracellular domain of either the wild-type or a cleavage-defective HIV-1 envelope glycoprotein (gp160) fused at one of two different positions in env to a C-terminal glycosyl-phosphatidylinositol (GPI) attachment signal from the mouse Thy-1.1 glycoprotein. All four of the constructs encoded glycoproteins that were efficiently expressed when Rev was supplied in trans, and the two cleavable forms were processed normally to gp120 and a chimeric "gp41." The chimeric glycoproteins, in contrast to the wild-type glycoprotein, could be cleaved from the surface of transfected cells by treatment with phosphatidylinositol-specific phospholipase C, indicating that they were anchored in the plasma membrane by a GPI moiety. These GPI-anchored glycoproteins were transported intracellularly at a rate only slightly lower than that of the full-length HIV-1 glycoprotein and were present on the cell surface in equivalent amounts. Nevertheless, all four glycoproteins were defective in mediating both cell-cell and virus-cell fusion as determined by syncytium formation in COS-1-HeLa-T4 cell mixtures and trans complementation of an env-defective HIV-1 genome.
Salzwedel
K
K
Department of Microbiology, University of Alabama, Birmingham 35294-0005.
Johnston
P B
PB
Roberts
S J
SJ
Dubay
J W
JW
Hunter
E
E
D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification; D001483:Base Sequence; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D015686:Gene Products, env / Q000032:analysis / Q000096:biosynthesis* / Q000302:isolation & purification; D015752:Genes, env; D015726:Giant Cells / Q000502:physiology; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification; D015183:Restriction Mapping; D018800:Thy-1 Antigens; D014162:Transfection
D000954:Antigens, Surface; D015686:Gene Products, env; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D018800:Thy-1 Antigens