In vitro studies of transcript initiation by Escherichia coli RNA polymerase. 3. Influences of individual DNA elements within the promoter recognition region on abortive initiation and promoter escape.
Abortive initiation and promoter escape are two principal biochemical reactions occurring in the latter stage of transcript initiation. We have analyzed the influences of individual DNA elements within the promoter recognition region (PRR) on these reactions by measuring the quantitative initiation parameters that describe abortive initiation and promoter escape; these parameters are the abortive rate, the productive rate, the abortive:productive ratio, the abortive probability, and the maximum size of abortive transcripts. Changes in the individual DNA elements within the PRR can have a substantial effect on each of these parameters. The discriminator region and the -10 element primarily influence the abortive probability at positions 2-5 and 6-10, respectively, while the -10 and -35 conserved hexamers and the spacer region affect the abortive probability at positions 11-15. Surprisingly, transcription of a consensus promoter invariably gives a higher abortive yield, a higher abortive probability, a longer abortive ladder, and a lower productive rate than promoter variants carrying even a single deviation in the consensus hexamers. These results suggest that strong RNA polymerase-PRR interactions stall the polymerase at the promoter, thereby reducing the rate of promoter escape and consequently enhancing the extent of abortive initiation.
Vo
Nam V
NV
Division of Biochemistry and Molecular Biology, Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3202, USA.
Hsu
Lilian M
LM
Kane
Caroline M
CM
Chamberlin
Michael J
MJ
D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D003570:Cytidine Triphosphate / Q000737:chemistry / Q000378:metabolism; D018983:DNA Footprinting; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006160:Guanosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000502:physiology*; D012329:RNA, Bacterial / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012689:Sequence Homology, Nucleic Acid; D013604:T-Phages / Q000235:genetics; D024363:Transcription Initiation Site / Q000502:physiology*; D014158:Transcription, Genetic; D014544:Uridine Triphosphate / Q000737:chemistry / Q000378:metabolism
D029968:Escherichia coli Proteins; D009692:Nucleic Acid Heteroduplexes; D012329:RNA, Bacterial; D003570:Cytidine Triphosphate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D012321:DNA-Directed RNA Polymerases; D014544:Uridine Triphosphate