Protein Kinase D3 Is Essential for Prostratin-Activated Transcription of Integrated HIV-1 Provirus Promoter via NF-κB Signaling Pathway
Figure 4
Prostratin-induced HIV-1 transcription depends on NF-κB. (a) Schematics of HIV-1 promoter deletion mutations. dSp1 (without Sp1 binding sites), dEnh (without NF-κB enhancer element), dTAR (without TAR RNA sequences), or dSp1/dEnh (without Sp1 or NF-κB enhancer element). (b) Effect of promoter mutation on prostratin-induced HIV-1 transcription. HeLa cells were transfected with HIV-LTR-Luc reporter constructs containing the indicated promoter deletions, followed by prostratin treatment. The luciferase activities were plotted based on 3 independent experiments, with the level of untreated cells set to 1.0. (c) Effect of prostratin treatment on RelA and Sp1 recruitment to promoter. HIV-LTR-Luc cells were treated with prostratin for 1 hr and subjected to chromatin immunoprecipitation (ChIP) analysis with indicated antibody. The levels of DNA isolated by ChIP were analyzed by quantitative PCR (qPCR) with the primers targeting promoter region of HIV-LTR as indicated on the left and plotted based on 2 independent experiments, with the level of untreated cells set to 1.0. (d) Effect of inhibiting NF-κB signaling on prostratin-stimulated HIV-1 transcription. HIV-LTR-Luc cells were pretreated with inhibitor BAY, followed by prostratin treatment. The luciferase activities were plotted based on 3 independent experiments, with the level of untreated cells set to 1.0.
