The Majority of Simian Immunodeficiency Virus/Mne Circle Junctions Result from Ligation of Unintegrated Viral DNA Ends That Are Aberrant for Integration

Randolph, C; Champoux, James J.

doi:10.1006/viro.1993.1329

Cited by 15 publications

(12 citation statements)

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“…LTR-LTR circle junctions, once thought to be the substrates for integration, are now known to be dead-end products of reverse transcription (60 -62). Sequence analysis of circle junctions has revealed that many contain deletions within U3, some of which could be explained by plus strand priming downstream from the PPT (63)(64)(65).…”

Section: Discussionmentioning

confidence: 99%

RNA Degradation and Primer Selection by Moloney Murine Leukemia Virus Reverse Transcriptase Contribute to the Accuracy of Plus Strand Initiation

Kelleher

Champoux

2000

Journal of Biological Chemistry

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During reverse transcription, plus strand DNA synthesis is initiated at a purine-rich RNA primer generated by the RNase H activity of reverse transcriptase (RT). Specific initiation of plus strand synthesis from this polypurine tract (PPT) RNA is essential for the subsequent integration of the linear viral DNA product. Based on current models, it is predicted that priming from sites upstream of the PPT may be tolerated by the virus, whereas efficient extension from RNA primers located downstream from the PPT is predicted to generate dead-end products. By using hybrid duplex substrates derived from the Moloney murine leukemia virus long terminal repeat, we investigated the extent to which RNase H degrades the viral RNA during time course cleavage assays, and we tested the capacity of the polymerase activity of RT to use the resulting cleavage products as primers. We find that the majority of the RNA fragments generated by RNase H are 2-25 nucleotides in length, and only following extensive degradation are most fragments reduced to 10 nucleotides or smaller. Although extensive RNA degradation by RNase H likely eliminates many potential RNA primers, based on thermostability predictions it appears that some RNA fragments remain stably annealed to the DNA template. RNA primers generated by RNase H within the long terminal repeat sequence are found to have the capacity to initiate DNA synthesis by RT; however, the priming efficiency is significantly less than that observed with the PPT primer. We find that Moloney murine leukemia virus nucleocapsid protein reduces RNase H degradation and slightly alters the cleavage specificity of RT; however, nucleocapsid protein does not appear to enhance PPT primer utilization or suppress extension from non-PPT RNA primers.

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Section: Discussionmentioning

confidence: 99%

RNA Degradation and Primer Selection by Moloney Murine Leukemia Virus Reverse Transcriptase Contribute to the Accuracy of Plus Strand Initiation

Kelleher

Champoux

2000

Journal of Biological Chemistry

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“…Importantly, this overaccumulation of 2-LTR species has also been associated with IN-defective HIV viruses [50,80-82]. To explain this observation, it is currently assumed that linear HIV DNA, representing the precursor of integration [3,5], accumulates because it cannot be integrated and is rerouted into the circularization pathway producing 2-LTR molecules in the nucleus [29,83-85]. However, 2-LTR circles are also detected in WT infected cells.…”

Section: Resultsmentioning

confidence: 99%

“…However, 2-LTR circles are also detected in WT infected cells. In this case, 2-LTR formation was suggested to result from aberrant att sequences preventing their recognition by IN [83]. Moreover, since 2-LTR molecules have been detected both in the cytoplasm and the nucleus of PFV WT infected cells [40], as well as at very early time-points in cytoplasm of MLV infected cells [41], overproduction of 2-LTR DNA cannot simply be explained by such a rerouting of non-integrated viral DNA.…”

Section: Resultsmentioning

confidence: 99%

A novel function for spumaretrovirus integrase: an early requirement for integrase-mediated cleavage of 2 LTR circles

Delelis

Petit

Leh³

et al. 2005

Retrovirology

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Retroviral integration is central to viral persistence and pathogenesis, cancer as well as host genome evolution. However, it is unclear why integration appears essential for retrovirus production, especially given the abundance and transcriptional potential of non-integrated viral genomes. The involvement of retroviral endonuclease, also called integrase (IN), in replication steps apart from integration has been proposed, but is usually considered to be accessory. We observe here that integration of a retrovirus from the spumavirus family depends mainly on the quantity of viral DNA produced. Moreover, we found that IN directly participates to linear DNA production from 2-LTR circles by specifically cleaving the conserved palindromic sequence found at LTR-LTR junctions. These results challenge the prevailing view that integrase essential function is to catalyze retroviral DNA integration. Integrase activity upstream of this step, by controlling linear DNA production, is sufficient to explain the absolute requirement for this enzyme.The novel role of IN over 2-LTR circle junctions accounts for the pleiotropic effects observed in cells infected with IN mutants. It may explain why 1) 2-LTR circles accumulate in vivo in mutants carrying a defective IN while their linear and integrated DNA pools decrease; 2) why both LTRs are processed in a concerted manner. It also resolves the original puzzle concerning the integration of spumaretroviruses. More generally, it suggests to reassess 2-LTR circles as functional intermediates in the retrovirus cycle and to reconsider the idea that formation of the integrated provirus is an essential step of retrovirus production.

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“…The 2-LTR junctions created by host processes result mostly from fusion of unprocessed or improperly processed viral cDNA (49). The junctions created by autointegration, on the other hand, should look identical to those created by bona fide proviral formation; i.e., a processed "CA" dinucleotide should be joined to a random viral sequence.…”

Section: ) (B)mentioning

confidence: 99%

Enhanced Autointegration in Hyperstable Simian Immunodeficiency Virus Capsid Mutants Blocked after Reverse Transcription

Tipper

Sodroski

2013

J Virol

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cAfter entering a host cell, retroviruses such as simian immunodeficiency virus (SIV) uncoat, disassembling the viral capsid. Rates of uncoating that are too high and too low can be detrimental to the efficiency of infection. Rapid uncoating typically leads to blocks in reverse transcription, but the basis for replication defects associated with slow uncoating is less clear. Here we characterize the phenotypes of two SIVmac239 mutants with changes, A87E and A87D, in the helix 4/5 loop of the capsid protein. These mutant viruses exhibited normal capsid morphology but were significantly attenuated for infectivity. The infectivity of wild-type and mutant SIVmac239 was not decreased by aphidicolin-induced growth arrest of the target cells. In the cytosol of infected cells, the A87E and A87D capsids remained in particulate form longer than the wild-type SIVmac239 capsid, suggesting that the mutants uncoat more slowly than the wild-type capsid. Both mutants exhibited much higher levels of autointegrated DNA forms than wild-type SIVmac239. Thus, some changes in the helix 4/5 loop of the SIVmac239 capsid protein result in capsid hyperstability and an increase in autointegration.

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The Majority of Simian Immunodeficiency Virus/Mne Circle Junctions Result from Ligation of Unintegrated Viral DNA Ends That Are Aberrant for Integration

Cited by 15 publications

References 0 publications

RNA Degradation and Primer Selection by Moloney Murine Leukemia Virus Reverse Transcriptase Contribute to the Accuracy of Plus Strand Initiation

RNA Degradation and Primer Selection by Moloney Murine Leukemia Virus Reverse Transcriptase Contribute to the Accuracy of Plus Strand Initiation

A novel function for spumaretrovirus integrase: an early requirement for integrase-mediated cleavage of 2 LTR circles

Enhanced Autointegration in Hyperstable Simian Immunodeficiency Virus Capsid Mutants Blocked after Reverse Transcription

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