Coding Sequences Upstream of the Human Immunodeficiency Virus Type 1 Reverse Transcriptase Domain in Gag-Pol Are Not Essential for Incorporation of the Pr160
            <sup>
              <i>gag-pol</i>
            </sup>
            into Virus Particles

Chiu, Hsu Chen; Yao, Szu Yung; Wang, Chin Tien

doi:10.1128/jvi.76.7.3221-3231.2002

Cited by 30 publications

(57 citation statements)

References 38 publications

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“…It is generally accepted that Pr160 gag-pol dimerization or multimerization triggers PR activation, and therefore, sequences upstream or downstream of PR may affect PR-mediated virus maturation by interfering with Gag-Pol multimerization. Consistent with this suggestion, deletions of sequences upstream of PR (11,52) or mutations in downstream pol sequences can significantly affect PR-directed virus particle maturation (4, 30,37).…”

Section: Gag-polsupporting

confidence: 50%

“…Amplified DNA fragments were digested with a combination of EcoRV and BsrGI and ligated into HIVgpt. The following constructs have all been described in detail in previous reports: HIV-1 RT mutants (W401A and W401A/W402A) (9); Pol-truncated mutants containing inserted stop codons at the designated IN or RT residue positions (RN198, R560, R515, and R425); plasmids expressing the RT subunits p66 and p51, containing hemagglutinin (HA) tags at the N terminus (9,29,30); and the HIV-1 PR-inactivated mutant HIVgpt D25 (11,48).…”

Section: Methodsmentioning

confidence: 99%

“…The procedure used for this study has been described previously (11). Briefly, transfected 293T cell culture supernatant was harvested, filtered, and pelleted as described for our Western immunoblot analysis.…”

Section: Methodsmentioning

confidence: 99%

“…PR is encoded by pol, which is initially translated as the polyprotein precursor Pr160 gag-pol . It is generally believed that Pr160 gag-pol is incorporated into assembling virions via interaction with Pr55 gag through its N-terminal Gag determinants (10,11,19,22,40,41). However, some researchers have demonstrated that HIV-1 and murine leukemia virus (MLV) Pol can be packaged into virions at a reasonable efficiency despite an absence of Gag-Pol formation (3, 7).…”

mentioning

confidence: 99%

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A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

Chiang

Wang

Pan

et al. 2010

J Virol

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HIV-1 protease (PR) mediates the proteolytic processing of virus particles during or after virus budding. PR activation is thought to be triggered by appropriate Gag-Pol/Gag-Pol interaction; factors affecting this interaction either enhance or reduce PR-mediated cleavage efficiency, resulting in markedly reduced virion production or the release of inadequately processed virions. We previously showed that a Gag-Pol deletion mutation involving the reverse transcriptase tryptophan (Trp) repeat motif markedly impairs PR-mediated virus maturation and that an alanine substitution at W401 (W401A) or at both W401 and W402 (W401A/ W402A) partially or almost completely negates the enhancement effect of efavirenz (a nonnucleoside reverse transcriptase inhibitor) on PR-mediated virus processing efficiency. These data suggest that the Trp repeat motif may contribute to the PR activation process. Here we demonstrate that due to enhanced Gag cleavage efficiency, W402 alanine or leucine substitution significantly reduces virus production. However, W402 replacement with phenylalanine does not significantly affect virus particle assembly or processing, but it does markedly impair viral infectivity in a single-cycle infection assay. Our results demonstrate that a single amino acid substitution at HIV-1 RT can radically affect virus assembly by enhancing Gag cleavage efficiency, suggesting that in addition to contributing to RT biological function during the early stages of virus replication, the HIV-1 RT tryptophan repeat motif in a Gag-Pol context may play an important role in suppressing the premature activation of PR during late-stage virus replication.

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Section: Gag-polsupporting

confidence: 50%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

Chiang

Wang

Pan

et al. 2010

J Virol

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“…gag expression vector, pGAG [47], yielding a set of PR-defective versions Cell culture and transfection 293T cells and HeLa cells were maintained in DMEM supplemented with 10% fetal calf serum. Confluent 293T cells were trypsinized, split 1:10 and seeded onto 10-cm plates 24 h before transfection.…”

Section: Each Of Thegag Mutations Was Also Introduced Into An Hiv-1 Pr55mentioning

confidence: 99%

Mutations in the α-helix Directly C-terminal to the Major Homology Region of Human Immunodeficiency Virus Type 1 Capsid Protein Disrupt Gag Multimerization and Markedly Impair Virus Particle Production

2006

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FRET analysis of HIV‐1 Gag and GagPol interactions

2017

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The Gag protein of HIV multimerizes to form viral particles. The GagPol protein encoding virus‐specific enzymes, such as protease, reverse transcriptase, and integrase, is incorporated into HIV particles via interactions with Gag. The catalytically active forms of these enzymes are dimeric or tetrameric. We employed Förster resonance energy transfer (FRET) assays to evaluate Gag–Gag, Gag–GagPol, and GagPol–GagPol interactions and investigated Gag and Pol interdomains tolerant to fluorescent protein insertion for FRET assays. Our data indicated that the matrix (MA)–capsid (CA) domain junction in the Gag region and the Gag C terminus were equally available for Gag–Gag and Gag–GagPol interaction assays. For GagPol dimerization assays, insertion at the MA–CA domain junction was most favorable.

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Coding Sequences Upstream of the Human Immunodeficiency Virus Type 1 Reverse Transcriptase Domain in Gag-Pol Are Not Essential for Incorporation of the Pr160 ^gag-pol into Virus Particles

Cited by 30 publications

References 38 publications

A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

Mutations in the α-helix Directly C-terminal to the Major Homology Region of Human Immunodeficiency Virus Type 1 Capsid Protein Disrupt Gag Multimerization and Markedly Impair Virus Particle Production

FRET analysis of HIV‐1 Gag and GagPol interactions

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Coding Sequences Upstream of the Human Immunodeficiency Virus Type 1 Reverse Transcriptase Domain in Gag-Pol Are Not Essential for Incorporation of the Pr160 gag-pol into Virus Particles

Cited by 30 publications

References 38 publications

A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

A Single Amino Acid Substitution in HIV-1 Reverse Transcriptase Significantly Reduces Virion Release

Mutations in the α-helix Directly C-terminal to the Major Homology Region of Human Immunodeficiency Virus Type 1 Capsid Protein Disrupt Gag Multimerization and Markedly Impair Virus Particle Production

FRET analysis of HIV‐1 Gag and GagPol interactions

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Product

Resources

About

Coding Sequences Upstream of the Human Immunodeficiency Virus Type 1 Reverse Transcriptase Domain in Gag-Pol Are Not Essential for Incorporation of the Pr160 ^gag-pol into Virus Particles