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dc.contributor.authorXu, Hongzhan
dc.contributor.authorFranks, Tamera
dc.contributor.authorGibson, Gregory
dc.contributor.authorHuber, Kelly
dc.contributor.authorRahm, Nadia
dc.contributor.authorDe Castillia, Caterina Strambio
dc.contributor.authorLuban, Jeremy
dc.contributor.authorAiken, Christopher
dc.contributor.authorWatkins, Simon
dc.contributor.authorSluis-Cremer, Nicolas
dc.contributor.authorAmbrose, Zandrea
dc.date2022-08-11T08:08:29.000
dc.date.accessioned2022-08-23T15:57:01Z
dc.date.available2022-08-23T15:57:01Z
dc.date.issued2013-07-09
dc.date.submitted2014-03-11
dc.identifier.citation<p>Xu H, Franks T, Gibson G, Huber K, Rahm N, De Castillia CS, Luban J, Aiken C, Watkins S, Sluis-Cremer N, Ambrose Z. Evidence for biphasic uncoating during HIV-1 infection from a novel imaging assay. Retrovirology. 2013 Jul 9;10:70. doi: 10.1186/1742-4690-10-70. <a href="http://dx.doi.org/10.1186/1742-4690-10-70">Link to article on publisher's site</a></p>
dc.identifier.issn1742-4690 (Linking)
dc.identifier.doi10.1186/1742-4690-10-70
dc.identifier.pmid23835323
dc.identifier.urihttp://hdl.handle.net/20.500.14038/30097
dc.description.abstractBACKGROUND: Uncoating of the HIV-1 core plays a critical role during early post-fusion stages of infection but is poorly understood. Microscopy-based assays are unable to easily distinguish between intact and partially uncoated viral cores. RESULTS: In this study, we used 5-ethynyl uridine (EU) to label viral-associated RNA during HIV production. At early time points after infection with EU-labeled virions, the viral-associated RNA was stained with an EU-specific dye and was detected by confocal microscopy together with viral proteins. We observed that detection of the viral-associated RNA was specific for EU-labeled virions, was detected only after viral fusion with target cells, and occurred after an initial opening of the core. In vitro staining of cores showed that the opening of the core allowed the small molecule dye, but not RNase A or antibodies, inside. Also, staining of the viral-associated RNA, which is co-localized with nucleocapsid, decays over time after viral infection. The decay rate of RNA staining is dependent on capsid (CA) stability, which was altered by CA mutations or a small molecule inducer of HIV-1 uncoating. While the staining of EU-labeled RNA was not affected by inhibition of reverse transcription, the kinetics of core opening of different CA mutants correlated with initiation of reverse transcription. Analysis of the E45A CA mutant suggests that initial core opening is independent of complete capsid disassembly. CONCLUSIONS: Taken together, our results establish a novel RNA accessibility-based assay that detects an early event in HIV-1 uncoating and can be used to further define this process.
dc.language.isoen_US
dc.relation<p><a href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&list_uids=23835323&dopt=Abstract">Link to Article in PubMed</a></p>
dc.rightsCopyright 2013 Xu et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( <a href="http://creativecommons.org/licenses/by/2.0">http://creativecommons.org/licenses/by/2.0</a>), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
dc.subjectCell Line
dc.subjectHIV-1
dc.subjectHumans
dc.subjectMicroscopy, Confocal
dc.subjectRNA, Viral
dc.subjectStaining and Labeling
dc.subjectUridine
dc.subjectViral Proteins
dc.subjectVirology
dc.subject*Virus Uncoating
dc.subjectVirology
dc.titleEvidence for biphasic uncoating during HIV-1 infection from a novel imaging assay
dc.typeJournal Article
dc.source.journaltitleRetrovirology
dc.source.volume10
dc.identifier.legacyfulltexthttps://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=1331&amp;context=faculty_pubs&amp;unstamped=1
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/faculty_pubs/332
dc.identifier.contextkey5319147
refterms.dateFOA2022-08-23T15:57:01Z
html.description.abstract<p>BACKGROUND: Uncoating of the HIV-1 core plays a critical role during early post-fusion stages of infection but is poorly understood. Microscopy-based assays are unable to easily distinguish between intact and partially uncoated viral cores.</p> <p>RESULTS: In this study, we used 5-ethynyl uridine (EU) to label viral-associated RNA during HIV production. At early time points after infection with EU-labeled virions, the viral-associated RNA was stained with an EU-specific dye and was detected by confocal microscopy together with viral proteins. We observed that detection of the viral-associated RNA was specific for EU-labeled virions, was detected only after viral fusion with target cells, and occurred after an initial opening of the core. In vitro staining of cores showed that the opening of the core allowed the small molecule dye, but not RNase A or antibodies, inside. Also, staining of the viral-associated RNA, which is co-localized with nucleocapsid, decays over time after viral infection. The decay rate of RNA staining is dependent on capsid (CA) stability, which was altered by CA mutations or a small molecule inducer of HIV-1 uncoating. While the staining of EU-labeled RNA was not affected by inhibition of reverse transcription, the kinetics of core opening of different CA mutants correlated with initiation of reverse transcription. Analysis of the E45A CA mutant suggests that initial core opening is independent of complete capsid disassembly.</p> <p>CONCLUSIONS: Taken together, our results establish a novel RNA accessibility-based assay that detects an early event in HIV-1 uncoating and can be used to further define this process.</p>
dc.identifier.submissionpathfaculty_pubs/332
dc.contributor.departmentProgram in Molecular Medicine
dc.source.pages70


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