HIV-1 infection, and HIV-1 Tat protein, activate K111 provirus expression in part by inducing loss of heterochromatin. HIV-1 infection of cell lines (A) or human peripheral blood lymphocytes (PBLs; B) activates the expression of K111 as detected by qRT-PCR of K111 env RNA using the K111-specific probe K111P. K111 RNA expression was not detected, or was detected only at low titer, in uninfected cells. (C) Activation of K111 in cell lines constitutively expressing Tat, or in human PBLs by addition of exogenous HIV-1-Tat. K111 RNA levels measured by qRT-PCR were essentially undetectable in cells without Tat but were found at titers ranging from 10⁴–10⁶ in Tat-stimulated cells. (D) ChIP assays show that HIV-1-Tat reduces the heterochromatic marks H3K9me3 and H4K20me3 at K111 proviral loci. Histones containing the H3K9me3 or H4K20me3 modification were specifically immunoprecipitated from the nuclear extracts of control HeLa and HeLa Tat expressing cells, and the K111 DNA bound to the histones containing these heterochromatin marks was measured by qPCR using the K111-specific probe K111P. Tat expressing cell lines: Jurkat Tat, Hltat, and HeLaTatIII. (***) P < 0.0001, (**) P < 0.001.