Figure 6.

Histone acetyltransferase p300, but not PCAF, stimulates rDNA transcription in vitro. Truncated template assays were carried out using extracts from N1S1 cells, the rat rDNA promoter (–286 +624) and increasing amounts of p300 and PCAF as described in Materials and Methods. After the transcription reaction, an internal standard was added to correct for the recovery of transcripts and the transcripts were purified by phenol extraction and ethanol precipitation. The transcripts were resolved by electrophoresis on urea–acrylamide gels, the gels were dried and and transcription quantitated with a Molecular Dynamics PhosphorImager. The results in each reaction were normalized for recovery of the internal standard and stimulation was determined by comparison of the ratios of the authentic transcript to the internal standard. (A) The in vitro transcription assay shows stimulation of rDNA transcription by p300. The assays included 1.5 and 2.0 µl of affinity-purified p300 (concentration ∼1 mg/ml). (B) Transcription in the presence of 1 and 2 µl (concentration ∼1mg/ml) of affinity-purified PCAF.