Regulatory sequences in sigma 54 localise near the start of DNA melting.
Transcription initiation by the enhancer-dependent sigma(54) RNA polymerase holoenzyme is positively regulated after promoter binding. The promoter DNA melting process is subject to activation by an enhancer-bound activator protein with nucleoside triphosphate hydrolysis activity. Tethered iron chelate probes attached to amino and carboxyl-terminal domains of sigma(54) were used to map sigma(54)-DNA interaction sites. The two domains localise to form a centre over the -12 promoter region. The use of deletion mutants of sigma(54) suggests that amino-terminal and carboxyl-terminal sequences are both needed for the centre to function. Upon activation, the relationship between the centre and promoter DNA changes. We suggest that the activator re-organises the centre to favour stable open complex formation through adjustments in sigma(54)-DNA contact and sigma(54) conformation. The centre is close to the active site of the RNA polymerase and includes sigma(54) regulatory sequences needed for DNA melting upon activation. This contrasts systems where activators recruit RNA polymerase to promoter DNA, and the protein and DNA determinants required for activation localise away from promoter sequences closely associated with the start of DNA melting.
Department of Biology, Imperial College of Science Technology and Medicine, Imperial College Road, London, SW7 2AZ, UK.
S R
SR
Wigneshweraraj
M K
MK
Chaney
A
A
Ishihama
M
M
Buck
D001205:Ascorbic Acid / Q000494:pharmacology; D020029:Base Pairing; D001483:Base Sequence; D001665:Binding Sites; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004492:Edetic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004742:Enhancer Elements, Genetic / Q000235:genetics; D004795:Enzyme Stability; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D020035:Holoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007502:Iron Chelating Agents / Q000494:pharmacology; D007711:Klebsiella pneumoniae / Q000201:enzymology*; D009154:Mutation; D009691:Nucleic Acid Denaturation; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009942:Organometallic Compounds / Q000494:pharmacology; D010088:Oxidoreductases / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051877:RNA Polymerase Sigma 54; D012808:Sigma Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016962:Sinorhizobium meliloti / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects
D015342:DNA Probes; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D020035:Holoenzymes; D007502:Iron Chelating Agents; D009692:Nucleic Acid Heteroduplexes; D009942:Organometallic Compounds; D012808:Sigma Factor; C042007:1-(4-bromoacetamidobenzyl)EDTA; D004492:Edetic Acid; D006861:Hydrogen Peroxide; D010088:Oxidoreductases; C029130:nitrogenase reductase; D012321:DNA-Directed RNA Polymerases; D051877:RNA Polymerase Sigma 54; D001205:Ascorbic Acid