Histone acetyltransferase and protein kinase activities copurify with a putative Xenopus RNA polymerase I holoenzyme self-sufficient for promoter-dependent transcription

Mounting evidence suggests that eukaryotic RNA polymerases preassociate wit h multiple transcription factors in the absence of DNA, forming RNA polymer ase holoenzyme complexes. We have purified an apparent RNA polymerase I (Po l I) holoenzyme from Xenopus laevis cells by sequential chromatography on f ive columns: DEAE-Sepharose, Biorex 70, Sephacryl S300, Mono Q, and DNA-cel lulose. Single fractions from every column programmed accurate promoter dep endent transcription. Upon gel filtration chromatography, the Pol I holoenz yme elutes at a position overlapping the peak of Blue Dextran, suggesting a molecular mass in the range of similar to 2 MDa. Consistent with its large mass, Coomassie blue-stained sodium dodecyl sulfate-polyacrylamide gels re veal approximately 55 proteins in fractions purified to near homogeneity. W estern blotting shows that TATA-binding protein precisely copurifies with h oloenzyme activity, whereas the abundant Pol I transactivator upstream bind ing factor does not. Also copurifying with the holoenzyme are casein kinase II and a histone acetyltransferase activity with a substrate preference fo r histone H3. These results extend to Pol I the suggestion that signal tran sduction and chromatin-modifying activities are associated with eukaryotic RNA polymerases.