Studies of HIV-1 envelope glycoprotein-mediated fusion using a simple fluorescence assay.
OBJECTIVE
To study HIV envelope glycoprotein (Env)-mediated entry using a sensitive fusion assay.


METHODS
CD4+ lymphocytes or T-cell lines were labelled with fluorescent cytoplasm or membrane markers. Fusion with Env-expressing adherent cells was monitored by observing dye transfer from CD4+ cells to Env cells.


RESULTS
Cell-cell fusion began 20-30 min after co-cultivation at 37 degrees C. Pre-binding at 4 degrees C was observed not to decrease the lag phase before fusion. Cells expressing envelope glycoproteins from non-syncytium-inducing (NSI) HIV strains showed dye transfer between two cells without progression to syncytia. A glycosylphosphatidylinositol anchored Env was found to be incapable of mediating membrane fusion, as measured either by lipid or cytoplasm contents mixing. Primary mouse cells expressing human CD4 and mouse 3T3 cells stably expressing both human CD4 and human CD26 did not support fusion with our Env-expressing cells.


CONCLUSIONS
Env-mediated cell-cell fusion is a relatively slow process, probably reflecting a multi-step process occurring after CD4 binding and requiring the transmembrane domain of gp41. Env proteins are able to mediate cell-cell fusion at least under some experimental conditions, indicating that lack of a syncytia phenotype does not rule out the possibility of fusion occurring between only two or a few cells.
Weiss
C D
CD
Department of Pharmacology, University of California, San Francisco, USA.
Barnett
S W
SW
Cacalano
N
N
Killeen
N
N
Littman
D R
DR
White
J M
JM
D016475:3T3 Cells; D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism*; D016466:CHO Cells; D002459:Cell Fusion; D002460:Cell Line; D006224:Cricetinae; D019302:HIV Envelope Protein gp160 / Q000502:physiology*; D015497:HIV-1 / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D051379:Mice
D019302:HIV Envelope Protein gp160