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Protein acetylation has emerged as a means of controlling levels of mRNA synthesis in eukaryotic cells. Here we report that acetyl coenzyme A (acetyl-CoA) stimulates RNA polymerase II transcription in vitro in the absence of histones. The effect of acetyl-CoA on basal and activated transcription was studied in a human RNA polymerase II transcription system reconstituted from recombinant and highly purified transcription factors. Both basal and activated transcription were stimulated by the addition of acetyl-CoA to transcription reaction mixtures. By varying the concentrations of general transcription factors in the reaction mixtures, we found that acetyl-CoA decreased the concentration of TFIID required to observe transcription. Electrophoretic mobility shift assays and DNase I footprinting revealed that acetyl-CoA increased the affinity of the general transcription factor TFIID for promoter DNA in a TBP-associated factor (TAF)-dependent manner. Interestingly, acetyl-CoA also caused a conformational change in the TFIID-TFIIA-promoter complex as assessed by DNase I footprinting. These results show that acetyl-CoA alters the DNA binding activity of TFIID and indicate that this biologically important cofactor functions at multiple levels to control gene expression.Transcription from natural RNA polymerase II promoters is tightly controlled by the combined actions of positive and negative regulatory factors, including site-specific activators, repressors, cofactors, and chromatin-associated proteins. For transcription of any given gene, a complex array of signals must ultimately by integrated at the promoter to set the proper level of RNA production. Protein acetylation is one such signal implicated in controlling levels of mRNA synthesis in eukaryotes. It has been shown that specific lysines in the N termini of core histones are acetylated in transcriptionally active regions of chromatin (reviewed in references 6 and 39). The prevailing model suggests that acetylation of histones relieves the repressive effects of chromatin and allows the transcription machinery access to promoters, resulting in a localized derepression of transcription (reviewed in reference 40). Several nuclear histone acetyltransferases have been identified, including Tetrahymena p55 and the human coactivators CREB binding protein (CBP), p300, P/CAF, GCN5, and TAF II 250 (3, 7, 27, 29, 42). Transcriptional activators can recruit acetyltransferases to promoters, where they acetylate histones in nucleosomes (38). Conversely, deacetylases are thought to be recruited to chromatin by transcriptional repressors (reviewed in reference 31).Histones are not the only substrates of nuclear acetyltransferases. The tumor suppressor p53 is a substrate of acetylation by p300 (22). Acetylation at specific lysines stimulates DNA binding by p53 in response to DNA damage (22, 34). TFIIE and TFIIF can be acetylated by P/CAF, p300, and TAF II 250, although the functions of these posttranslational modifications have not been identified (15). HMG I(Y) is acetylated ...

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