Analysis of the Assembly of the HIV Core by Electron Microscope Tomography

Höglund, Stefan; Öfverstedt, Lars-Göran; Nilsson, Åsa; Özel, Muhsin; Winkel, Thorsten; Skoglund, Ulf; Gelderblom, Hans R.

doi:10.1007/978-1-349-11907-3_16

Cited by 6 publications

(7 citation statements)

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“…4A and B), demonstrating that the protein sequence within p46 gag is sufficient for particle formation. These results support previous findings that the formation of infectious particles depends on the presence of at least part of the p15 gag protein (13,17,35). The expression of p55M1-10 demonstrated the release of immature particles from transfected cells into the medium (Fig.…”

Section: Resultssupporting

confidence: 92%

Inactivation of the human immunodeficiency virus type 1 inhibitory elements allows Rev-independent expression of Gag and Gag/protease and particle formation

Schneider

Campbell

Nasioulas

et al. 1997

J Virol

236

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The expression of gag, pol, and env of human immunodeficiency virus type 1 (HIV-1) depends on the presence of the viral Rev protein. This dependence is, at least in part, due to the presence of negatively acting sequences (inhibitory or instability elements [INS]) located within unspliced and partially spliced mRNAs. The positive interaction of Rev with the Rev-responsive element in these mRNAs counteracts the negative effects of the inhibitory sequences. Here, we demonstrate that in addition to the previously identified INS1 within p17 gag , several other INS elements exist within the gag/pol region of HIV-1. These elements act independently of each other and were eliminated by mutagenesis after the introduction of multiple point mutations not affecting the coding region, leading to constitutive high levels of Gag expression. Expression vectors containing an intact or nearly intact p55 gag region allowed the production of immature viral particles in mammalian cells in the absence of any other HIV proteins. The introduction of additional mutations in the protease region allowed efficient production of Gag/protease, which resulted in processing of the Pr55 gag precursor and production of mature Gag particles with a lentivirus-like conical-core structure. The elimination of a newly identified INS element within pol and the previously identified CRS located within int was accomplished by the same methodology. Sequence comparisons of the identified inhibitory elements revealed no apparent homologies and demonstrated that these sequences are not splice sites. These results demonstrate that the elimination of INS elements leads to efficient expression of HIV-1 mRNAs in the absence of Rev or any posttranscriptional activating mechanisms.

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Section: Resultssupporting

confidence: 92%

Inactivation of the human immunodeficiency virus type 1 inhibitory elements allows Rev-independent expression of Gag and Gag/protease and particle formation

Schneider

Campbell

Nasioulas

et al. 1997

J Virol

236

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“…It was proved that the core surface a t the narrow end of the core is connected to part of the envelope membrane or matrix-protein ( Fig. 7), which is in agreement with the results of Hoglund et al (1990Hoglund et al ( , 1992. An explanation for the observation of a particle without any membrane connection may be that the core is ultimately isolated from the envelope at a late stage of the viral maturation, or that we observed a section which had been cut at such an angle that the Fig.…”

Section: Discussionsupporting

confidence: 88%

“…In HIV-1, it was reported that this connection might consist of the carboxy-terminal protein p6 and that it might play a n essential role at a late stage of budding (Veronese et al, 1987). Recently, Hoglund et al (1990Hoglund et al ( , 1992 observed the structure of HIV-1 using three-dimensional electron microscopic tomography. We used freeze fracturing to establish that there is continuity between the core and the envelope.…”

Section: Discussionmentioning

confidence: 99%

Inner architecture of human and simian immunodeficiency viruses

Fukui

Imura

Goto

et al. 1993

Microscopy Res & Technique

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The cores of human and simian immunodeficiency viruses (HIV and SIV) were observed by negative staining after isolation of the core with Nonidet P40 and glutaraldehyde. Four kinds of cores were found: asymmetric and symmetric sectoral shapes, a bar shape, and a triangular shape. These results were confirmed by the examination of ultrathin sections of whole virions. In some virions, the connection between the core and the envelope was observed after freeze fracturing. Its structure was considered to be characteristic of an intermediate stage of viral maturation. The HIV-1 core was reacted with anti-HIV-1 p24 mouse monoclonal antibody.

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“…which are characterized by the morphology of the structure found inside the viral envelope. The electron-dense structure seen in immature particles is spherical and lines the inside of the viral envelope, while the structure seen in mature virus is cone-shaped and eccentrically located (6,14,15,23,24,52). Expression of various HIVgag constructs has demonstrated that the Gag precursor polyprotein can, by itself, form spherical structures like those found in immature particles (16).…”

mentioning

confidence: 99%

Assembly of recombinant human immunodeficiency virus type 1 capsid protein in vitro

Ehrlich

Agresta

Carter

1992

J Virol

203

120

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The capsid protein (CA) (p24) of human immunodeficiency virus (HIV) type 1 expressed in Escherichia coli and purified to >90%o homogeneity was used to examine assembly in vitro and to probe the nature of interactions involved in the formation of capsid structures. The protein was detected in dimeric and oligomeric forms as indicated by molecular size measurements by gel filtration column chromatography, sedimentation through sucrose, and nondenaturing gel electrophoresis. Chemical cross-linking of CA molecules was observed with several homobifunctional reagents. Oligomer size was dependent on cross-linker concentration and exhibited a nonrandom pattern in which dimers and tetramers were more abundant than trimers and pentamers. Oligomers as large as dodecamers were detected in native polyacrylamide gels. These were stable in solutions of high ionic strength or in the presence of nonionic detergent, indicating that strong interactions were involved in oligomer stabilization. Limited tryptic digestion converted the putative dodecamers to octamers, suggesting that a region involved in CA protein multimerization was exposed in the structure. This region was mapped to the central portion of the protein. The recombinant CA proteins assembled in vitro into long rodlike structures and were disassembled into small irregular spheres by alterations in ionic strength and pH. The observation that assembly and disassembly of purified HIV type 1 CA protein can be induced in vitro suggests an approach for identifying possible control mechanisms involved in HIV viral core assembly.

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Analysis of the Assembly of the HIV Core by Electron Microscope Tomography

Cited by 6 publications

References 0 publications

Inactivation of the human immunodeficiency virus type 1 inhibitory elements allows Rev-independent expression of Gag and Gag/protease and particle formation

Inactivation of the human immunodeficiency virus type 1 inhibitory elements allows Rev-independent expression of Gag and Gag/protease and particle formation

Inner architecture of human and simian immunodeficiency viruses

Assembly of recombinant human immunodeficiency virus type 1 capsid protein in vitro

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