The neutral glycosphingolipid globotriaosylceramide promotes fusion mediated by a CD4-dependent CXCR4-utilizing HIV type 1 envelope glycoprotein.
Previously, we showed that the addition of human erythrocyte glycosphingolipids (GSLs) to nonhuman CD4(+) or GSL-depleted human CD4(+) cells rendered those cells susceptible to HIV-1 envelope glycoprotein-mediated cell fusion. Individual components in the GSL mixture were isolated by fractionation on a silica-gel column and incorporated into the membranes of CD4(+) cells. GSL-supplemented target cells were then examined for their ability to fuse with TF228 cells expressing HIV-1LAI envelope glycoprotein. We found that one GSL fraction, fraction 3, exhibited the highest recovery of fusion after incorporation into CD4(+) nonhuman and GSL-depleted HeLa-CD4 cells and that fraction 3 contained a single GSL fraction. Fraction 3 was characterized by MS, NMR spectroscopy, enzymatic analysis, and immunostaining with an antiglobotriaosylceramide (Gb3) antibody and was found to be Gal(alpha1-->4)Gal(beta1-->4)Glc-Cer (Gb3). The addition of fraction 3 or Gb3 to GSL-depleted HeLa-CD4 cells recovered fusion, but the addition of galactosylceramide, glucosylceramide, the monosialoganglioside, GM3, lactosylceramide, globoside, the disialoganglioside, GD3, or alpha-galactosidase A-digested fraction 3 had no effect. Our findings show that the neutral GSL, Gb3, is required for CD4/CXCR4-dependent HIV-1 fusion.
Puri
A
A
Section of Membrane Structure and Function, Laboratory of Experimental and Computational Biology, Division of Basic Sciences, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
Hug
P
P
Jernigan
K
K
Barchi
J
J
Kim
H Y
HY
Hamilton
J
J
Wiels
J
J
Murray
G J
GJ
Brady
R O
RO
Blumenthal
R
R
D000107:Acetylation; D015704:CD4 Antigens / Q000502:physiology*; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002459:Cell Fusion / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D004912:Erythrocytes / Q000737:chemistry / Q000502:physiology; D015686:Gene Products, env / Q000187:drug effects / Q000502:physiology*; D015497:HIV-1 / Q000187:drug effects / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D013058:Mass Spectrometry; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D019718:Receptors, CXCR4 / Q000187:drug effects / Q000502:physiology*; D014281:Trihexosylceramides / Q000097:blood / Q000737:chemistry / Q000494:pharmacology*
D015704:CD4 Antigens; D015686:Gene Products, env; D019718:Receptors, CXCR4; D014281:Trihexosylceramides; C018549:globotriaosylceramide