Human immunodeficiency virus (HIV or AIDS virus) contains two heavily glycosylated envelope proteins-gp41 as a transmembrane protein and gp120 as an outer membrane protein. These glycoproteins mediate attachment of virus particles to cell surface receptors and appear responsible for syncytium formation and associated cytopathic effects of the virus in infected cells. Although these glycoproteins have these obligatory roles in the pathogenesis of HIV, there are no reports describing the significance of HIV envelope protein glycosylation in this process. Therefore, we propose to investigate what role, if any, protein glycosylation has in the pathogenesis of HIV. These investigations will focus utilizing tunicamycin, deoxynorjirimycin, castanospermine, bromoconduritol and swainsonine as specific inhibitors of different steps in the glycoprotein processing pathway. Physical and biochemical effects of these inhibitors on HIV glycoproteins will be examined by (35S) methionine and (3H) mannose radiolabelling coupled with density gradient centrifugation and SDS-PAGE/fluorographic methodologies. Biological effects of these inhibitors will be examined in vitro with respect to viral infectivity, target cell infectability, syncytium formation and cytopathic effects. As an assay of these biological effects, C3 and peripheral blood lymphocytes will be used as target cells for infection with the HIV isolates HTLV-III and LAV produced in H9/HTLV-111 and CEM/LAV cultures, respectively. Infections will be monitored by indirect immunofluorescence of cells for viral antigen synthesis, reverse transcriptase activity in culture fluids for virus production, Giemsa stained microscopic examination for syncytium formation and vital dye (neutral red) uptake for cytopathic effect. Information gained from these studies could provide a new avenue of attack against retroviruses and find great value in the future development of effective anti-HIV therapeutics or vaccines.
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