Single-amino acid mutation 66SR in Gag-matrix enhances viral single-cycle infectivity of R5-tropic HIV-1rmt

Doi, Naoya; Sakai, Yosuke; Miyazaki, Yasuaki; Adachi, Akio; Nomaguchi, Masako

doi:10.2152/jmi.62.228

Cited by 2 publications

(9 citation statements)

References 21 publications

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“…These new clones were analyzed for their single -cycle infectivity and multi-cycle replication ability. We have demonstrated here that two proviral clones grow better than a standard CCR5 -tropic HIV-1rmt clone designated MN5/ LSDQgtu (5gtu) (1,9,10). We have also shown, together with our previous results, that growth-promoting mutations in HIV-1 Env exert their positive effects in an Env-dependent manner.…”

Section: Introductionsupporting

confidence: 77%

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Generation and characterization of new CCR5-tropic HIV-1rmt clones

et al. 2017

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: To develop effective non -human primate models for coping with numerous HIV-1/AIDS studies, rhesus macaque-tropic HIV-1 (HIV-1rmt) clones with a variety of biological properties are required. Such clones, if available, are powerful tools to experimentally elucidate HIV-1 replication and pathogenicity in host individuals, and also to develop anti-HIV-1 drugs/vaccines. However, only limited numbers of HIV-1rmt clones have been currently reported. In the present study, we generated new HIV-1rmt clones carrying various CCR5-tropic env (envelope) genes by standard recombinant DNA and intracellular homologous recombination techniques. Resultant virus clones contain the env sequences derived from an AIDS-inducible laboratory or two clinically isolated viral strains. We further constructed their variant clones bearing N160K, S304G, or G310R mutation in Env that potentially can change the viruses to better grow. Newly generated clones were analyzed for their virological properties such as Env expression, single-cycle infectivity, and multi -cycle replication ability. Out of a number of new clones examined, two were found to grow better in macaque cells than the previously constructed clone used for comparison. Our study described here constitutes the initial and essential step towards obtaining CCR5-tropic HIV-1rmt clones useful for various basic and clinical research projects on infected individuals.

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Section: Introductionsupporting

confidence: 77%

“…Construction and characterization of a CCR5 -tropic HIV-1rmt clone designated MN5/LSDQgtu (5gtu) have been previously described (1,9,10). New CCR5 -tropic proviral clones were generated as outlined in Fig.…”

Section: Proviral Molecular Clonesmentioning

confidence: 99%

Generation and characterization of new CCR5-tropic HIV-1rmt clones

et al. 2017

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“…A human kidney cell line 293T and a HeLa-derived reporter cell line TZM-bl carrying a luciferase gene under control of HIV-1 long terminal repeat (LTR) were cultured in Eagle's minimal essential containing 10% heat-inactivated fetal bovine serum (20). Lymphocyte cell lines H9 (human) and HSC-F (cynomolgus macaque) were maintained in RPMI1640 medium containing 10% heat-inactivated fetal bovine serum (19,21).…”

Section: Cellsmentioning

confidence: 99%

“…To monitor multi-cycle infectivity (virus growth ability/potential), equal RT units (10 5 for H9 cells and 10 6 for HSC-F cells) of virus samples were infected into H9 (10 5 ) or HSC -F cells (2 x 10 5 ), and virus growth kinetics were determined by RT assays as previously described (17,20). H9 and HSC -F cell lines show phenotypes very similar to that of their corresponding primary cells such as peripheral blood mononuclear cells and lymphocytes with respect to the susceptibility to infections with various HIV/SIVs and/or their mutant viruses.…”

Section: Assays For Viral Infectivitymentioning

confidence: 99%

“…Progeny viruses produced in infected cells were monitored at intervals (every 3 days) during 15 days following infection. To determine single-cycle infectivity, equal RT units (10 4 ) of virus samples were inoculated into TZM-bl cells (4 x 10 3 cells were seeded into a well of a 96-well plate one day before infection), and on day 1 post-infection, cells were lysed for luciferase assays (Promega Corporation) as described previously (20). Luciferase activity in cell lysates relative to that of NL4 -3 was considered as viral single -cycle infectivity.…”

Section: Assays For Viral Infectivitymentioning

confidence: 99%

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Virological characterization of HIV‐1 CA‐NTD mutants constructed in a virus‐lineage reflected manner

et al. 2018

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: Capsid (CA) protein is a major virion-constituent of all retroviruses including human immunodeficiency virus type 1 (HIV-1), and is essential for early and late phases in viral replication cycle through interaction with numerous cellular factors. In particular, N-terminal domain (NTD) of HIV-1 CA has been frequently and well reported to bind to various host cell proteins that considerably affect viral replication potential. In this study, in order to better define biological bases of the CA-NTD for HIV-1 replication, we performed an extensive mutational analysis in an unprecedented manner. By aligning CA-NTD sequences derived from representative infectious molecular clones of HIV-1, HIV-2, and simian immunodeficiency virus isolated from the rhesus macaque (SIVmac), a number of amino acids specific to HIV-1 were selected, and were replaced with those from SIVmac at the corresponding sites. Mutant viruses thus generated were then examined for multi -cycle infectivity, single-cycle infectivity, and ability to produce progeny virions. While some CA-NTD mutations affected viral replication ability to varying degrees, those in helix 7 abolished viral growth potential without exception. These results highlight functional importance of non -conserved amino acids in helix 7, and give new insights into functionality of HIV-1 CA-NTD.

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Single-amino acid mutation 66SR in Gag-matrix enhances viral single-cycle infectivity of R5-tropic HIV-1rmt

Cited by 2 publications

References 21 publications

Generation and characterization of new CCR5-tropic HIV-1rmt clones

Generation and characterization of new CCR5-tropic HIV-1rmt clones

Virological characterization of HIV‐1 CA‐NTD mutants constructed in a virus‐lineage reflected manner

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