5'-long terminal repeat-selective CpG methylation of latent human T-cell leukemia virus type 1 provirus in vitro and in vivo

CpG methylation of the human T-cell leukemia virus type 1 (HTLV-1) long terminal repeat (LTR) has been implicated in proviral latency, but there is presently little information available regarding the pattern of LTR methylation and its effect on viral gene expression. To gain insight into the mechanisms of HTLV-1 latency, we have studied methylation of individual CpG sites in the U3-R region of the integrated proviral LTR by using bisulfite genomic sequencing methods. Surprisingly, our results reveal selective hypermethylation of the 5' LTR and accompanying hypomethylation of the 3' LTR in both latently infected cell lines and adult T-cell leukemia (ATL) cells having a complete provirus. Moreover, we observed a lack of CpG methylation in the LTRs of 5'-defective proviruses recovered from ATL samples, which is consistent with the selective hypomethylation of the 3' LTR. Thus, the integrated HTLV-1 provirus in these carriers appears to be hypermethylated in the 5' LTR and hypomethylated in the 3' LTR. These results, together with the observation that proviral gene expression is reactivated by 5-azacytidine in latently infected cell lines, indicate that selective hypermethylation of the HTLV-1 5' LTR is common both in vivo and in vitro. Thus, hypermethylation of the 5' LTR appears to be an important mechanism by which HTLV-1 gene expression is repressed during viral latency.     

Histonedeacetylase inhibitor Oxamflatin increase HIV-1 transcription by inducing histone modification in latently infected cells

HIV-1 latency represents a major problem in the eradication of HIV-1 in infected individuals treated with highly active anti-retroviral therapy. Histone deacetylase (HDAC) inhibits HIV-1 gene expression and virus production and may contribute to quiescence of HIV-1 within resting CD4+ T cells. Here, we evaluated the effect of Oxamflatin, a class I HDAC inhibitor, on the epigenetic change at HIV-1 long terminal repeat (LTR) and the induction of the latent viruses in the latency Jurkat T cell line. Flow cytometry assay showed that Oxamflatin activate HIV-1 gene expression in these latently infected cells by 2-17 fold than background levels. Chromatin immunoprecipitation (ChIP) assays further revealed that Oxamflatin increase the acetylation level of histone H3 and histone H4 at the nucleosome 1(nuc-1) site of the HIV-1 LTR compared to mock treatment. We also found that Oxamflatin had a synergization with prostratin, or 5-azacytidine or tumor necrosis factor-α to activate the HIV-1 promoter. Taken together, our results suggest that the histone acetylation plays an important role in regulating HIV-1 LTR gene expression, and Oxamflatin has potential as drug candidates as antilatency therapies.     

Host-cell-determined methylation of specific Epstein-Barr virus promoters regulates the choice between distinct viral latency programs.

Epstein-Barr virus (EBV) is capable of adopting three distinct forms of latency: the type III latency program, in which six EBV-encoded nuclear antigens (EBNAs) are expressed, and the type I and type II latency programs, in which only a single viral nuclear protein, EBNA1, is produced. Several groups have reported heavy CpG methylation of the EBV genome in Burkitt's lymphoma cell lines which maintain type I latency, and loss of viral genome methylation in tumor cell lines has been correlated with a switch to type III latency. Here, evidence that the type III latency program must be inactivated by methylation to allow EBV to enter the type I or type II restricted latency program is provided. The data demonstrates that the EBNA1 gene promoter, Qp, active in types I and II latency, is encompassed by a CpG island which is protected from methylation. CpG methylation inactivates the type III latency program and consequently allows the type I or II latency program to operate by alleviating EBNA1-mediated repression of Qp. Methylation of the type III latency EBNA gene promoter, Cp, appears to be essential to prevent type III latency, since EBNA1 is expressed in all latently infected cells and, as shown here, is the only viral antigen required for activation of Cp. EBV is thus a pathogen which subverts host-cell-determined methylation to regulate distinct genetic programs.     

Paucity of HIV DNA methylation in latently infected, resting CD4+ T cells from infected individuals receiving antiretroviral therapy

Maintenance of HIV latency in vitro has been linked to methylation of HIV DNA. However, examinations of the degree of methylation of HIV DNA in the latently infected, resting CD4(+) T cells of infected individuals receiving antiretroviral therapy have been limited. Here, we show that methylation of the HIV 5' long terminal repeat (LTR) in the latent viral reservoir of HIV-infected aviremic individuals receiving therapy is rare, suggesting that other mechanisms are likely involved in the persistence of viral latency.