0580715 - ÚMG 2024 RIV CZ eng A - Abstrakt
Kaňka, Jakub - Miklík, Dalibor - Hejnar, Jiří
ENLIGHTENING THE QUIETNESS –THE SEARCH FOR EARLY SILENCED PROVIRUSES.
Czech Chemical Society Symposium Series. Roč. 21, č. 5 (2023), č. článku P-63. ISSN 2336-7202.
[Annual meeting of the National Institute of Virology and Bacteriology (NIVB) /2./. 02.10.2023-05.10.2023, Kutná Hora]
Grant CEP: GA MŠMT(CZ) LX22NPO5103
Institucionální podpora: RVO:68378050
Klíčová slova: retroviral DNA * HIV * detection system
Obor OECD: Virology
http://ccsss.cz/index.php/ccsss/issue/view/41

Integration of retroviral DNA into the genome of an infected cell is a hallmark of retroviral infection. Genomic and epigenomic features at the site of integration may determine the expression of retroviral genes. While some proviruses (integrated retroviral genome) are stably expressed for several months, other proviruses are silenced shortly after integration. In the case of HIV infection, provirus silencing is the first step leading to the establishment of the latent reservoir in vivo, the main hurdle in curing HIV infection. Thus, understanding the mechanisms involved in early proviral silencing is crucial for the functional cure of HIV. Currently, it is possible to distinguish between transcriptionally active and inactive proviruses 2–3 days after transduction. However, whether silenced proviruses were ever transiently active after integration remains to be known.To determine this, we developed a novel detection system. First, sensor cells were generated using the transposon-mediated insertion of a detection cassette. After the sensor cells were established, a replication-defective retroviral vector transducing a recombinase gene was used as an activator. In this system, any cell in which the recombination is detected without the expression of the proviral marker (GFP) contains an early silenced provirus. These cells could then be obtained and further studied.Our data from initial experiments demonstrate that the sensor cells can be prepared within 2–3 weeks using the piggyBactransposon system. Contrary to published data, the use of piggyBactransposon in our system showed that most human embryonic kidney cells (293T) transduced by avian sarcoma leukosis virus (ASLV) contained proviruses that were transiently active after integration. This suggests that early proviral silencing predominantly does not occur immediately after, but within several hours after integration. Our approach can be further applied to virtually any model of retroviral infection, helping to elucidate conditions under which a temporal proviral expression occurs and processes integral to it.
Trvalý link: https://hdl.handle.net/11104/0349470