Figure 2.

ET-743 does not affect TSA-induced global histone hyperacetylation or accumulation of MDR1 promoter-associated acetylated histone H4. Stably transfected SW620 cells were incubated for 24 h in the presence of TSA alone (100 ng), ET-743 alone (50 nM), or a combination of both. (A) After incubation, total histones were isolated and subjected to Western analysis with an anti-acetylated histone H4 antibody. The two bands observed represent different acetylated forms of histone H4. (B and C) For chromatin immunoprecipitation assays, cells were treated as above, and then chromatin was immunoprecipitated with mouse IgG, anti-acetylated H4 antibody, or no antibody (negative control). MDR1 promoter DNA associated with hyperacetylated H4 was visualized after PCR amplification. The arrow indicates the specific MDR1 promoter PCR product, and the asterisk denotes the input primers. (B) Input DNA. (C) Immunoprecipitated DNA. ChIP, chromatin immunoprecipitation; NTC, negative control; UT, untreated; M, marker.