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Transcriptional provirus silencing as a crosstalk of de novo DNA methylation and epigenomic features at the integration site.

ABSTRACT: The autonomous transcription of integrated retroviruses strongly depends on genetic and epigenetic effects of the chromatin at the site of integration. These effects are mostly suppressive and proviral activity can be finally silenced by mechanisms, such as DNA methylation and histone modifications. To address the role of the integration site at the whole-genome-scale, we performed clonal analysis of provirus silencing with an avian leucosis/sarcoma virus-based reporter vector and correlated the transcriptional silencing with the epigenomic landscape of respective integrations. We demonstrate efficient provirus silencing in human HCT116 cell line, which is strongly but not absolutely dependent on the de novo DNA methyltransferase activity, particularly of Dnmt3b. Proviruses integrated close to the transcription start sites of active genes into the regions enriched in H3K4 trimethylation display long-term stability of expression and are resistant to the transcriptional silencing after over-expression of Dnmt3a or Dnmt3b. In contrast, proviruses in the intergenic regions tend to spontaneous transcriptional silencing even in Dnmt3a(-/-) Dnmt3b(-/-) cells. The silencing of proviruses within genes is accompanied with DNA methylation of long terminal repeats, whereas silencing in intergenic regions is DNA methylation-independent. These findings indicate that the epigenomic features of integration sites are crucial for their permissivity to the proviral expression.

SUBMITTER: Senigl F

PROVIDER: S-EPMC3384319 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

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Transcriptional provirus silencing as a crosstalk of de novo DNA methylation and epigenomic features at the integration site.

Senigl Filip F Auxt Miroslav M Hejnar Jirí J

Nucleic acids research 20120229 12

The autonomous transcription of integrated retroviruses strongly depends on genetic and epigenetic effects of the chromatin at the site of integration. These effects are mostly suppressive and proviral activity can be finally silenced by mechanisms, such as DNA methylation and histone modifications. To address the role of the integration site at the whole-genome-scale, we performed clonal analysis of provirus silencing with an avian leucosis/sarcoma virus-based reporter vector and correlated the ...[more]

PMID: 22379139

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Project description:UnlabelledUnderstanding the mechanisms of HIV proviral latency is essential for development of a means to eradicate infection and achieve a cure. We have previously described an in vitro latency model that reliably identifies HIV expression phenotypes of infected cells using a dual-fluorescence reporter virus. Our results have demonstrated that ?50% of infected cells establish latency immediately upon integration of provirus, a phenomenon termed early latency, which appears to occur by mechanisms that are distinct from epigenetic silencing observed with HIV provirus that establishes productive infections. In this study, we have used a mini-dual HIV reporter virus (mdHIV) to compare the long-term stability of provirus produced as early latent or productive infections using Jurkat-Tat T cell clones. Cloned lines bearing mdHIV provirus integrated at different chromosomal locations display unique differences in responsiveness to signaling agonists and chromatin-modifying compounds, and they also produce characteristic expression patterns from the 5' long terminal repeat (LTR) dsRed and internal EIF1?-enhanced green fluorescent protein (EIF1?-eGFP) reporters. Furthermore, reporter expression profiles of single cell sorted subcultures faithfully reproduce expression profiles identical to that of their original parental population, following prolonged growth in culture, without shifting toward expression patterns resembling that of cell subclones at the time of sorting. Comparison of population dispersion coefficient (CV) and mean fluorescence intensity (MFI) of the subcloned lines showed that both untreated and phorbol myristate acetate (PMA)-ionomycin-stimulated cultures produce expression patterns identical to those of their parental lines. These results indicate that HIV provirus expression characteristics are strongly influenced by the epigenetic landscape at the site of chromosomal integration.ImportanceThere is currently considerable interest in development of therapies to eliminate latently infected cells from HIV-infected patients on antiretroviral therapy. One proposed strategy, known as "shock and kill," would involve treatment with therapies capable of inducing expression of latent provirus, with the expectation that the latently infected cells could be killed by a host immune response or virus-induced apoptosis. In clinical trials, histone deacetylase (HDAC) inhibitors were shown to cause reactivation of latent provirus but did not produce a significant effect toward eliminating the latently infected population. Results shown here indicate that integration of HIV provirus at different chromosomal locations produces significant effects on the responsiveness of virus expression to T cell signaling agonists and chromatin-modifying compounds. Given the variety of phenotypes produced by integrated provirus, it is unlikely that any single potential shock-and-kill therapy will be effective toward purging the latently infected population.

Dataset Information

Transcriptional provirus silencing as a crosstalk of de novo DNA methylation and epigenomic features at the integration site.

Publications

Transcriptional provirus silencing as a crosstalk of de novo DNA methylation and epigenomic features at the integration site.

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