Involvement of Histone H3 Lysine 9 (H3K9) Methyltransferase G9a in the Maintenance of HIV-1 Latency and Its Reactivation by BIX01294

Elucidating the mechanism of human immunodeficiency virus, type 1 (HIV-1) provirus transcriptional silencing in latently infected cells is crucial for understanding the pathophysiological process of HIV-1 infection. It is well established that hypoacetylation of histone proteins by histone deacetylases is involved in the maintenance of HIV-1 latency by repressing viral transcription. Although histone methylation is involved in the organization of chromatin domains and plays a central epigenetic role in gene expression, the role of histone methylation in the maintenance of HIV-1 latency has not been clarified. Here we present evidence that histone H3 Lys9 (H3K9) methyltransferase G9a is responsible for transcriptional repression of HIV-1 by promoting repressive dimethylation at H3K9 and for the maintenance of viral latency. We observed that G9a significantly inhibited basal, as well as, the induced HIV-1 gene expression by tumor necrosis factor-alpha or Tat. Mutant G9a, however, lacking the SET domain responsible for the catalytic activity of histone methyltransferase, did not show such an effect. When G9a expression was knocked down by small interfering RNA, HIV-1 replication was augmented from cells transiently transfected with a full-length HIV-1 clone. Moreover, a specific inhibitor of G9a, BIX01294, could reactivate expression of HIV-1 from latently infected cells such as ACH-2 and OM10.1. Furthermore, chromatin immunoprecipitation assays revealed the presence of G9a and H3K9 dimethylation on nucleosome histones in the vicinity of the HIV-1 long terminal repeat promoter. These results suggest that G9a is responsible for the transcriptional quiescence of latent HIV-1 provirus and provide a molecular basis for understanding the mechanism by which HIV-1 latency is maintained.