Initiation of reverse transcription during cell-to-cell transmission of human immunodeficiency virus infection uses pre-existing reverse transcriptase

Stephenson, Alice J.; Brennan, Peter; Karageorgos, Litsa; Kok, Tuckweng; Kuiper, Lara J.; Swift, J.G.; Burrell, Christopher J.

doi:10.1099/0022-1317-75-8-1917

Cited by 8 publications

(7 citation statements)

References 38 publications

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“…as judged by supernatant reverse transcriptase activity. This lag phase is equivalent to the early observations with the one-step growth experiment using phages (Burnet, 1929;Burnet, 1960;d'Herelle, 1926;Ellis and Delbruck, 1939) and suggested a highly efficient initiation of infection with this cell-to-cell transmission route (Li et al, 1992a;Li et al, 1994). This was followed by the release of progeny viral particles within 24-48h (Barbosa et al, 1994;Li and Burrell, 1992;Sato et al, 1992).…”

supporting

confidence: 78%

Importance of HIV cell-to-cell transmission - implications for neutralizing antibody

T¹,

P²

2018

Preprint

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Viruses can infect a cell via one or both routes viz. cell-to-cell (c-c) or cell-free (c-f) . Pathogenesis studies of various viruses, including HIV, have shown that c-c transmission yields a significantly higher infection magnitude than the c-f route. Expectedly, potent antibodies inhibited c-f infection more efficiently than with c-cell transmission. To achieve a one-step, synchronous infection cycle that provides amplified infection, we have studied a consistent and efficient c-c HIV infection model since 1992. H9 cells persistently infected with HTLV-IIIB (H3B cells) and uninfected target CD4+ lymphocyte line (HuT78) were mixed in a ratio of 1:4 respectively. We have recently used this model to produce HIV designer antigens that have been shown to elicit monoclonal as well as polyclonal specific antibodies against novel epitopes that are formed post virus-cell engagement, but prior to fusion. The model can be extended for HIV neutralizing antibody assays or drug inhibitors against high multiplicity of infection.

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supporting

confidence: 78%

Importance of HIV cell-to-cell transmission - implications for neutralizing antibody

T¹,

P²

2018

Preprint

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“…Different efficiencies arising from using different primer pairs and other PCR conditions were normalized by conversion of each PCR product signal to copy numbers by comparison with known standards. Our results demonstrate that even following cell-to-cell transmission of HIV infection, during which the production of mature virions and the early steps of virus attachment and penetration into the susceptible cells may be bypassed Sato et al, 1992;Li et al, 1994), there still is a considerable time delay in the initiation of minus-strand HIV DNA synthesis. A time delay was also found to be associated with the initiation of the plus-strand DNA synthesis and the first but not the second template transfer.…”

Section: Introductionmentioning

confidence: 85%

“…In contrast, our results identify an initial lag period of ~ 1-5 h required for initiation of the reverse transcription process and the first detection of ~ 0.3 copies of minus-strand strong-stop DNA per cell. This initiation may include fusion of the viral donor and recipient cells, conformational or structural alterations to the viral nucleocapsid, a correctly orientated RNA secondary structure and the interaction of this structure with viral reverse transcriptase and the tRNA primer and possibly other viral and cellular proteins, leading to the activation of the reverse transcription process (Aiyar et al, 1994;Li et al, 1994).…”

Section: Discussionmentioning

confidence: 99%

“…We have previously developed a synchronized onestep cell-to-cell HIV infection model and we have used this model to investigate, in particular, viral reverse transcription in T cells following cell-to-cell HIV infection (Li et al, , 1994Karageorgos et al, 1993). In the present study, we report a quantitative PCR analysis of the kinetics of HIV reverse transcription in this model.…”

Section: Introductionmentioning

confidence: 98%

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Stepwise analysis of reverse transcription in a cell-to-cell human immunodeficiency virus infection model: kinetics and implications

Karageorgos

Burrell

1995

Journal of General Virology

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We have investigated the kinetics of human immunodeficiency virus (HIV) reverse transcription in infected T cells, using a synchronized, one-step, cell-to-cell infection model and quantitative PCR assays for the different DNA intermediate structures that are found sequentially during reverse transcription. Different efficiencies that might arise from the use of different primers and other PCR conditions were normalized by conversion of each PCR product signal to copy numbers by comparing with standards. After an initial lag period, the minus-strand strong-stop viral DNA was detected first. This was followed by the post-transfer newly extended minus-strand viral DNA and then by the plusstrand strong-stop DNA and fully extended minusstrand DNA. Kinetic data indicated that, once reverse transcription was initiated, the HIV reverse transcriptase synthesized minus-strand DNA at a rate of 150-180 bases/min, and that the first template transfer and the initiation of the plus-strand DNA synthesis imposed specific time delays. In contrast, minus-strand viral DNA synthesized after the second template transfer appeared at a time point very close to the time of the appearance of the last piece of DNA synthesized just before the second template switch, suggesting that the second switch occurred very rapidly. Taken together, our results define more accurately than was previously possible the rates of several of the steps in HIV reverse transcription in infected T cell lines and indicate different mechanisms for the two distinct template switches during retrovirus reverse transcription.

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“…We next assessed the presence of these RT isoforms in other biological situations: in (i) virus producer cells (Figure 3A ), (ii) intracellularly following HIV infection (Figure 3B ) and (iii) in HIV RTC's (Figure 3C–E ). H3B cells are chronically HIV infected cells that produce infectious virus and although they contain forms of HIV RT that are active in vitro , RT is not active inside the cell and newly synthesised HIV DNA is not formed until stimulation by mixing with uninfected recipient cells [ 2 ]. H3B cells thus represent a system to analyse changes in RT that occur co-incident with intracellular stimulation of reverse transcription and additionally offers the advantage of a synchronous and highly efficient infection model compared with a cell-free infection [ 13 ].…”

Section: Resultsmentioning

confidence: 99%

Human Immunodeficiency Virus type-1 reverse transcriptase exists as post-translationally modified forms in virions and cells

et al. 2008

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Background: HIV-1 reverse transcriptase (RT) is a heterodimer composed of p66 and p51 subunits and is responsible for reverse transcription of the viral RNA genome into DNA. RT can be post-translationally modified in vitro which may be an important mechanism for regulating RT activity. Here we report detection of different p66 and p51 RT isoforms by 2D gel electrophoresis in virions and infected cells.

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Initiation of reverse transcription during cell-to-cell transmission of human immunodeficiency virus infection uses pre-existing reverse transcriptase

Cited by 8 publications

References 38 publications

Importance of HIV cell-to-cell transmission - implications for neutralizing antibody

Importance of HIV cell-to-cell transmission - implications for neutralizing antibody

Stepwise analysis of reverse transcription in a cell-to-cell human immunodeficiency virus infection model: kinetics and implications

Human Immunodeficiency Virus type-1 reverse transcriptase exists as post-translationally modified forms in virions and cells

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