RNA polymerase II elongation complex. Elongation complexes purified using an anti-RNA antibody do not contain initiation factor alpha.
Gene expression in eukaryotes can be regulated by controlling the efficiency of transcript elongation by RNA polymerase II. The composition of the elongation complex is, however, poorly understood. Previous work has identified DNA sequences which block RNA polymerase II transcription and factors which stimulate RNA chain elongation. Here, I have purified elongation complexes arrested at discrete template locations. Complexes were rapidly and efficiently precipitated from in vitro transcription reactions using a monoclonal antibody that binds RNA. The isolated complexes remained transcriptionally active. This technique enables the facile manipulation of transcription elongation complexes. Using this approach, I show that transcription initiation factor alpha is not associated with a RNA polymerase II elongation complex. Since others have shown that alpha associates stoichiometrically with DNA, RNA polymerase II, and other required factors in an initiation complex, this work suggests that alpha departs from the transcription complex after nucleotides are required but before extensive RNA chain synthesis. In this regard alpha resembles the bacterial promoter-recognition factor sigma.
Reines
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Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001345:Autoradiography; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008099:Liver / Q000201:enzymology; D051379:Mice; D010441:Peptide Chain Elongation, Translational; D010448:Peptide Initiation Factors / Q000302:isolation & purification; D011233:Precipitin Tests; D012313:RNA / Q000276:immunology*; D012319:RNA Polymerase II / Q000302:isolation & purification*; D051381:Rats; D014158:Transcription, Genetic
D000911:Antibodies, Monoclonal; D010448:Peptide Initiation Factors; D012313:RNA; D012319:RNA Polymerase II