Article

Restoration of wild-type infectivity to human immunodeficiency virus type 1 strains lacking nef by intravirion reverse transcription.

Department of Microbiology/Biochemistry/Immunology, Morehouse School of Medicine, Atlanta, Georgia 30310, USA.
Journal of Virology (Impact Factor: 4.65). 01/2002; 75(24):12081-7. DOI: 10.1128/JVI.75.24.12081-12087.2001
Source: PubMed

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) Nef protein exerts several effects, both on infected cells and as a virion protein, which work together to enhance viral replication. One of these activities is the ability to enhance infectivity and the formation of proviral DNA. The mechanism of this enhancement remains incompletely understood. We show that virions with nef deleted can be restored to wild-type infectivity by stimulating intravirion reverse transcription. Particle composition and measures of reverse transcriptase activity remain the same for Nef(+) and Nef(-) virions both before and after natural endogenous reverse transcription (NERT) treatment. The effect of NERT treatment on virions pseudotyped with murine leukemia virus envelope protein was similar to that on particles pseudotyped with HIV-1 envelope protein. However, virions pseudotyped with vesicular stomatitis virus G envelope protein showed no influence of Nef on NERT enhancement of infectivity. These observations suggest that Nef may function at a level prior to reverse transcription. Since NERT treatment results in partial disassembly of the viral core, we speculate that Nef may function at the level of core particle disassembly.

Download full-text

Full-text

Available from: Mahfuz Bano Khan, Jun 17, 2015
0 Followers
 · 
63 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Nef is an important pathogenesis factor of HIV-1 with a multitude of effector functions. We have designed a broad panel of isogenic viruses encoding defined mutants of HIV-1(SF2) Nef and analyzed their biological activity in the context of productive HIV-1 infection. Analysis of subcellular localization, virion incorporation, downregulation of cell surface CD4 and MHC-I, enhancement of virion infectivity and facilitation of HIV replication in primary human T lymphocytes mostly confirmed the mapping of Nef determinants previously reported upon isolated expression of Nef. However, reduced activity in downregulation of CD4, infectivity enhancement and virion incorporation of a Nef variant (Delta12-39) lacking an amphipatic helix required for binding of a cellular kinase complex and the association of Nef with MHC-I/AP-1 suggested a novel role of this N-terminal motif. The SH3 binding motif of Nef was partially required for infectivity enhancement and replication but not for receptor downmodulation. In contrast to previous results obtained using other Nef alleles, non-myristoylated SF2-Nef was only partly defective when expressed during HIV infection and was present in HIV-1 particles. Importantly, incorporation of Nef into HIV-1 virions was not required for any of the tested Nef activities. Altogether, this study provides a broad characterization and mapping of multiple Nef activities in HIV-infected cells. The results emphasize that multiple activities govern Nef's effects on HIV replication and argue against a role of virion incorporation for Nef's activity as pathogenicity factor.
    Virology 09/2006; 351(2):322-39. DOI:10.1016/j.virol.2006.03.044 · 3.28 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: HIV-1 budding is directed primarily by two motifs in Gag p6 designated as late domain-1 and -2 that recruit ESCRT machinery by binding Tsg101 and Alix, respectively, and by poorly characterized determinants in the capsid (CA) domain. Here, we report that a conserved Gag p6 residue, S40, impacts budding mediated by all of these determinants. Whereas budding normally results in formation of single spherical particles ~100 nm in diameter and containing a characteristic electron-dense conical core, the substitution of Phe for S40, a change that does not alter the amino acids encoded in the overlapping pol reading frame, resulted in defective CA-SP1 cleavage, formation of strings of tethered particles or filopodia-like membrane protrusions containing Gag, and diminished infectious particle formation. The S40F-mediated release defects were exacerbated when the viral-encoded protease (PR) was inactivated or when L domain-1 function was disrupted or when budding was almost completely obliterated by the disruption of both L domain-1 and -2. S40F mutation also resulted in stronger Gag-Alix interaction, as detected by yeast 2-hybrid assay. Reducing Alix binding by mutational disruption of contact residues restored single particle release, implicating the perturbed Gag-Alix interaction in the aberrant budding events. Interestingly, introduction of S40F partially rescued the negative effects on budding of CA NTD mutations EE75,76AA and P99A, which both prevent membrane curvature and therefore block budding at an early stage. The results indicate that the S40 residue is a novel determinant of HIV-1 egress that is most likely involved in regulation of a critical assembly event required for budding in the Tsg101-, Alix-, Nedd4- and CA N-terminal domain affected pathways.
    Retrovirology 11/2013; 10(1):143. DOI:10.1186/1742-4690-10-143 · 4.77 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The replication and pathogenicity of lentiviruses is crucially modulated by "auxiliary proteins" which are expressed in addition to the canonical retroviral ORFs gag, pol, and env. Strategies to inhibit the activity of such proteins are often sought and proposed as possible additions to increase efficacy of the traditional antiretroviral therapy. This requires the acquisition of an in-depth knowledge of the molecular mechanisms underlying their function. The Nef auxiliary protein is expressed uniquely by primate lentiviruses and plays an important role in virus replication in vivo and in the onset of AIDS. Among its several activities Nef enhances the intrinsic infectivity of progeny virions through a mechanism which remains today enigmatic. Here we review the current knowledge surrounding such activity and we discuss its possible role in HIV biology.
    Frontiers in Microbiology 05/2014; 5:232. DOI:10.3389/fmicb.2014.00232 · 3.94 Impact Factor