Contacts between mammalian RNA polymerase II and the template DNA in a ternary elongation complex.
Elongation complexes of RNA polymerase II, RNA-DNA-enzyme ternary complexes, are intermediates in the synthesis of all eukaryotic mRNAs and are potential regulatory targets for factors controlling RNA chain elongation and termination. Analysis of such complexes can provide information concerning the structure of the catalytic core of the RNA polymerase and its interactions with the DNA template and RNA transcript. Knowledge of the structure of such complexes is essential in understanding the catalytic and regulatory properties of RNA polymerase. We have prepared and isolated complexes of purified RNA polymerase II halted at defined positions along a DNA template, and we have used deoxyribonuclease I (DNAse I) to map the interactions of the polymerase with the DNA template. DNAse I footprints of three specific ternary complexes reveal that the enzyme-template interactions of individual elongation complexes are not identical. The size of the protected region is distinct for each complex and varies from 48 to 55 bp between different complexes. Additionally, the positioning of the protected region relative to the active site varies in different complexes. Our results suggest that RNA polymerase II is a dynamic molecule and undergoes continual conformational transitions during elongation. These transitions are likely to be important in the processes of transcript elongation and termination and their regulation.
Rice
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Division of Biochemistry and Molecular Biology, University of California, Berkeley 94720.
Chamberlin
M J
MJ
Kane
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CM
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism*; D003850:Deoxyribonuclease I; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D015394:Molecular Structure; D012319:RNA Polymerase II / Q000737:chemistry / Q000378:metabolism*; D013698:Templates, Genetic; D014158:Transcription, Genetic
D004277:DNA, Single-Stranded; D012319:RNA Polymerase II; D003850:Deoxyribonuclease I