Lysine methylation of HIV-1 Tat regulates transcriptional activity of the viral LTR

We analyzed the association of Tat with histone methyltransferases of the SUV39-family of SET domain containing proteins in vitro. Tat was found to associate with both SETDB1 and SETDB2, two enzymes which exhibit methyltransferase activity. siRNA against SETDB1 transfected into cell systems with both transient and integrated LTR reporter genes resulted in an increase in transcription of the HIV-LTR in the presence of suboptimal levels of Tat. In vitro methylation assays with Tat peptides containing point mutations at lysines 50 and 51 showed an increased incorporation of methyl groups on lysine 51, however, both residues indicated susceptibility for methylation.The association of Tat with histone methyltransferases and the ability for Tat to be methylated suggests an interesting mechanism of transcriptional regulation through the recruitment of chromatin remodeling proteins to the HIV-1 promoter.The HIV-1 genome incorporates nine viral genes, all of which are expressed from a single promoter located within the viral long terminal repeat (LTR) [1,2]. The activity of the HIV-1 promoter is strongly dependant on the viral transactivator, Tat, the protein responsible for transcriptional activation and elongation [3-8]. The main function of Tat is to activate the HIV-1 LTR by binding to an RNA stem-loop structure, TAR [3,4,6,9-11]. This interaction initiates a binding cascade where cellular transcription factors such as Cdk9 and cyclin T1 are recruited to the HIV-1 promoter to facilitate viral transcription [12-15]. Tat mediates the functional modifications associated with viral transcription primarily by interacting with host cellular kinases, specifically to phosphorylate the large subunit of RNA Pol II CTD resulting in the activation of elongation [12,16,17]. In addition to the recruitment of host cellular proteins and enzymes for transcriptional initiation, such as NF-κB, Sp1, and TFIID, Tat has also been shown to bind a number of other factors which regulate chromati