FIG. 6.
Interactions of USF, RNA Pol II, p300, and modified histones with the β-globin gene locus in MEL cells expressing dominant-negative A-USF. (A) pTO/A-USF, pTR (A-USF), and pTO, pTR (control [Ctrl.]) MEL cells were induced with doxycycline. Cells were subjected to ChIP, immunoprecipitating with anti (α)-USF1, α-USF2, or no antibody (No Ab). Semiquantitative PCR was performed on samples, including input, using primers specific for the β-major promoter, HS2, or the ɛγ promoter. (B and C) MEL cells transfected with the A-USF expression vector (pTO/A-USF) or empty vector pTO were subjected to ChIP analysis with antibodies against AcH3 and RNA Pol II, or p300 (B). DNA was purified from the immunoprecipitate and analyzed by qPCR with primers specific for the βmaj-globin promoter and LCR element HS2. The error bars represent the results from two independent experiments. Ctrl represents either no antibody or nonspecific IgG antibody. (C) Results from pTO control cells are shown as white boxes. Results from pTO/A-USF cells are represented by black boxes. Samples described in the legend for panel B were analyzed with primers specific to the GAPDH promoter. All quantitative data were subjected to statistical analysis, and the P values were all below 0.05.