The Role of Cellular Factors in Promoting HIV Budding

Program in Gene Function and Expression, University of Massachusetts Medical School, Worcester, MA 01605, USA.
Journal of Molecular Biology (Impact Factor: 4.33). 07/2011; 410(4):525-33. DOI: 10.1016/j.jmb.2011.04.055
Source: PubMed

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) becomes enveloped while budding through the plasma membrane, and the release of nascent virions requires a membrane fission event that separates the viral envelope from the cell surface. To facilitate this crucial step in its life cycle, HIV-1 exploits a complex cellular membrane remodeling and fission machinery known as the endosomal sorting complex required for transport (ESCRT) pathway. HIV-1 Gag directly interacts with early-acting components of this pathway, which ultimately triggers the assembly of the ESCRT-III membrane fission complex at viral budding sites. Surprisingly, HIV-1 requires only a subset of ESCRT-III components, indicating that the membrane fission reaction that occurs during HIV-1 budding differs in crucial aspects from topologically related cellular abscission events.

Download full-text


Available from: Heinrich G Gottlinger, Aug 25, 2015
  • Source
    • "The myristylation of Gly2 in MA targets the Gag protein to the plasma membrane [1] [20], and mutation of this glycine myristylation site prevents virus budding and leads to accumulation of Gag within the host cell [21]. The structure of MA has been determined by both X-ray crystallography and NMR, and comprises 5 alpha helices and a three strand mixed beta sheet [22] [23] [24] which forms a trimer in the virion shell in the mature HIV-1 particle. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The HIV-1 Gag precursor protein, Pr55Gag, is a multi-domain polyprotein that drives HIV-1 assembly. The morphological features of HIV-1 suggested Pr55Gag assumes a variety of different conformations during virion assembly and maturation, yet structural determination of HIV-1 Pr55Gag has not been possible due to an inability to express and to isolate large amounts of full-length recombinant Pr55Gag for biophysical and biochemical analyses. This challenge is further complicated by HIV-1 Gag’s natural propensity to multimerize for the formation of viral particle (with ∼2500 Gag molecules per virion), and this has led Pr55Gag to aggregate and be expressed as inclusion bodies in a number of in vitro protein expression systems. This study reported the production of a recombinant form of HIV-1 Pr55Gag using a bacterial heterologous expression system. Recombinant HIV-1 Pr55Gag was expressed with a C-terminal Hisx6 tag, and purified using a combination of immobilized metal affinity chromatography and size exclusion chromatography. This procedure resulted in the production of milligram quantities of high purity HIV-1 Pr55Gag that has a mobility that resembles a trimer in solution using size exclusion chromatography analysis. The high quantity and purity of the full length HIV Gag will be suitable for structural and functional studies to further understand the process of viral assembly, maturation and the development of inhibitors to interfere with the process.
    Protein Expression and Purification 08/2014; 100. DOI:10.1016/j.pep.2014.04.013 · 1.51 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The Endosomal Sorting Complex Required for Transport (ESCRT) machinery is a set of multi-protein complexes that are well conserved among all eukaryotes and mediate a remarkable array of cellular processes including late endosome/multivesicular body (MVB) formation, retroviral particle release, and membrane abscission during cytokinesis. While the molecular mechanisms underlying ESCRT function have been relatively well characterized in yeasts and mammals, far less is known about ESCRT in plants. In this study, we utilized publicly-available microarray, massively parallel signature sequencing (MPSS) and proteome data sets in order to survey the expression profiles of many of the components of the Arabidopsis thaliana ESCRT machinery. Overall, the results indicate that ESCRT expression in Arabidopsis is highly dynamic across a wide range of organs, tissues and treatments, consistent with the complex interplay that likely exists between the spatial and temporal regulation of the ESCRT machinery and the diverse array of roles that ESCRT participates in during plant growth and development.
    Plant signaling & behavior 12/2011; 6(12):1897-903. DOI:10.4161/psb.6.12.18023
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The detachment of human immunodeficiency type 1 (HIV-1) virions depends on CHPM4 family members, which are late-acting components of the ESCRT pathway that mediate the cleavage of bud necks from the cytosolic side. We now show that in human cells, CHMP4 proteins are to a considerable extent bound to two high-molecular-weight proteins that we have identified as CC2D1A and CC2D1B. Both proteins bind to the core domain of CHMP4B, which has a strong propensity to polymerize and to inhibit HIV-1 budding. Further mapping showed that CC2D1A binds to an N-terminal hairpin within the CHMP4 core that has been implicated in polymerization. Consistent with a model in which CC2D1A and CC2D1B regulate CHMP4 polymerization, the overexpression of CC2D1A inhibited both the release of wild-type HIV-1 and the CHMP4-dependent rescue of an HIV-1 L domain mutant by exogenous ALIX. Furthermore, small interfering RNA against CC2D1A or CC2D1B increased HIV-1 budding under certain conditions. CC2D1A and CC2D1B possess four Drosophila melanogaster 14 (DM14) domains, and we demonstrate that these constitute novel CHMP4 binding modules. The DM14 domain that bound most avidly to CHMP4B was by itself sufficient to inhibit the function of ALIX in HIV-1 budding, indicating that the inhibition occurred through CHMP4 sequestration. However, N-terminal fragments of CC2D1A that did not interact with CHMP4B nevertheless retained a significant level of inhibitory activity. Thus, CC2D1A may also affect HIV-1 budding in a CHMP4-independent manner.
    Journal of Virology 01/2012; 86(7):3746-56. DOI:10.1128/JVI.06539-11 · 4.65 Impact Factor
Show more