Reconstitution of chromatin transcription with purified components reveals a chromatin-specific repressive activity of p300

Here we describe an in vitro chromatin transcription system in which chromatin assembly and transcription are carried out with purified and defined factors. With basal ( also known as general) transcription factors and sequence-specific DNA-binding activators, we observed chromatin-specific, activation domain - dependent transcription. We then examined the biochemical function of purified p300 in the absence of the endogenous factor and other related activities and found, unexpectedly, that p300 has a chromatin-specific, transcriptional repression activity that can be relieved by the addition of acetyl-CoA. This p300-mediated repression is reversible, requires the p300 bromodomain but not the acetyltransferase region, and does not involve the formation of a stable, nuclease-resistant nucleoprotein complex. Hence, the mechanism of transcriptional repression by p300 is distinct from that of histone H1, PARP-1 or Sir2. These findings reveal a novel chromatin-specific repressive function of p300.