11th International AIDS Conference Vancouver, British Columbia — July 7-12, 1996

Int Conf AIDS 1996 Jul 7-12; 11:39 (abstract no. Mo.A.1021)
Chen SS, Terwilliger E; Academia Sinica, Taipei, Taiwan, R.O.C. Fax: 02-782-5573. E-mail: schen@ibms.sinica.edu.tw.

OBJECTIVE: To determine whether an HIV-1 envelope (Env) variant lacking the gp41 cytoplasmic domain can function as an inhibitory mutant in the production of infectious virus, and to study the mechanism underlying the interference effect conferred by this mutant protein.

METHODS: The interference effect conferred by a mutant provirus on infectious virus production was examined by cotransfection of 293 cells with wt and mutant proviruses followed by measuring the infectivity of the resulting supernatant virus on Sup T1 cells. Heterooligomerization between wt and mutant proteins was assayed by isolation of Env immune complexes using the Chessie 8 monoclonal antibody that reacts with an epitope in the gp41 cytoplasmic domain, followed by Western blotting with anti-gp120 antibody. The interference conferred by the mutant protein upon the ability of wt protein to mediate virus entry into cells was determined by a trans-complementation assay.

RESULTS: Coexpression of the wt HIV-1 provirus and a mutant provirus that encoded an Env protein lacking the gp41 cytoplasmic domain inhibited the production of infectious virus in a transient assay. The mutant protein formed a complex with the wt protein when they were coexpressed. The defect in infectivity of the recombinant virus that contained coexpressed wt and mutant proteins did not appear to involve the late steps of virus replication, since coexpression did not affect the synthesis, precursor processing, or intracellular transport of the Env protein, nor did it alter the incorporation of the Env protein into virions and the subsequent release of the virus. Moreover, mutant protein coexpression effectively inhibited the ability of the wt protein to mediate cell-to-cell transmission, as analyzed by a trans-complementation assay.

CONCLUSIONS: These results show that this mutant protein interferes with wt Env protein function by forming a dysfunctional heterooligomer that may impose a defect in the early steps of the virus life cycle.

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MoA1021

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