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. 2014 Jul 23;42(14):8954–8969. doi: 10.1093/nar/gku611

Figure 2.

Figure 2.

Effect of depletion of class I HDACs on 7SK snRNA and cellular distribution of N-CoR/HDAC3 and Cdk9 in normoxic and hypoxic HeLa cells. (A) Effect of disruption of class I HDACs on the formation of an inactive endogenous P-TEFb complex in response to hypoxia. HeLa cells were exposed for 1 h to a normoxic (N) or hypoxic (H) gas mixture 48 h after transfection with siRNA. Whole cell lysates were immunoprecipitated with a goat anti-Cyclin T1 antibody and analyzed by western blot using rabbit anti-HEXIM1 and an anti-Cdk9 antibody. Direct western blots of HDAC1, HDAC2 and HDAC3 served as a control for siRNA-mediated knockdown. Western blot with an anti-Cyclin T1 antibody served as a control of total immunoprecipitated protein. (B) Localization of endogenous Cdk9 (red) and HDAC3 (green). Cells were treated with hypoxia, IL-1β or their combination for 2 h and analyzed by immunofluorescence staining. (C) Immunofluorescence showing cellular localization of endogenous HDAC3 (green) and N-CoR (red) in response to 2 h exposure to IL-1β alone or in combination with hypoxia. Nucleic acids were stained using TO-PRO-3 (blue).