Promoter recognition and discrimination by Eσ<sup>S</sup> RNA polymerase

Gaál, Tamás; Ross, Wilma; Estrem, Shawn T.; Nguyen, Lam H.; Burgess, Richard R.; Gourse, Richard L.

doi:10.1046/j.1365-2958.2001.02703.x

Cited by 161 publications

(172 citation statements)

References 66 publications

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“…DNase I protection assays suggest different interactions between the two forms of RNA polymerase and PaidB(12C3 T) in and immediately upstream of the Ϫ35 region (Fig. 6), confirming previous observations at the synthetic "Ϫ35 con" promoter and at osmY (10,41). This preferential interaction of PaidB(12C3 T) with E S is likely to depend on two promoter elements, i.e.…”

Section: Discussionsupporting

confidence: 88%

“…At the so-called "extended Ϫ10" promoters, this TG dinucleotide can be found two nucleotides upstream from the Ϫ10 sequence, where it allows transcription even in the absence of a conserved Ϫ35 sequence (23)(24)(25)(26)(27)(28). However, in vitro selection experiments (10) suggest that a TG motif might also be recognized by S and have proposed TGTGCTATA(c/a)T as the optimal extended Ϫ10 sequence for S binding.…”

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confidence: 99%

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Nucleotides from –16 to –12 Determine Specific Promoter Recognition by Bacterial σS-RNA Polymerase

Lacour

Kolb

Landini

2003

Journal of Biological Chemistry

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The alternative sigma factor S , mainly active in stationary phase of growth, recognizes in vitro a ؊10 promoter sequence almost identical to the one for the main sigma factor, 70 , thus raising the problem of how specific promoter recognition by S -RNA polymerase (E S ) is achieved in vivo. We investigated the promoter features involved in selective recognition by E S at the strictly S -dependent aidB promoter. We show that the presence of a C nucleotide as first residue of the aidB ؊10 sequence (؊12C), instead of the T nucleotide canonical for 70 -dependent promoters, is the major determinant for selective recognition by E S . The presence of the ؊12C does not allow formation of an open complex fully proficient in transcription initiation by E 70 . The role of ؊12C as specific determinant for promoter recognition by E S was confirmed by sequence analysis of known E S -dependent promoters as well as site-directed mutagenesis at the promoters of the csgB and sprE genes. We propose that E S , unlike E 70 , can recognize both C and T as the first nucleotide in the ؊10 sequence. Additional promoter features such as the presence of a C nucleotide at position ؊13, contributing to open complex formation by E S , and a TG motif found at the unusual ؊16/؊15 location, possibly contributing to initial binding to the promoter, also represent important factors for S -dependent transcription. We propose a new sequence, TG(N) 0 -2 CCATA(c/a)T, as consensus ؊10 sequence for promoters exclusively recognized by E S .Bacterial cells adapt to changing environmental and physiological conditions by modulating gene expression. Sigma ( ) factors of RNA polymerase, as the subunits responsible for promoter recognition, play a major role in programming gene expression. At least seven different subunits have been identified in Escherichia coli; 70 is the main subunit and can carry out transcription from the majority of E. coli promoters. The alternative subunits can direct transcription toward specific sets of genes (i.e. heat-shock, extracellular proteins, etc.) whose transcription is directed by -specific promoter sequences. A partial exception to the typical role for alternative subunits is represented by S , the product of the rpoS gene, mainly expressed in the stationary phase of growth (1-3). Unlike the other subunits, S -RNA polymerase (E S ) 1 can initiate transcription from several promoters also recognized by E 70 , suggesting that they recognize similar promoter sequences (4). The recognition of similar promoter sequences by S and 70 is reflected by their strong similarity in the DNA binding domains (5). The alignment of E S -dependent promoters and the search for an optimal promoter for E S in vitro using the systematic evolution of ligands by exponential enrichment (SELEX) procedure have pointed to a Ϫ10 consensus sequence for E S , CTATA(c/a)T that is very similar to the canonical TATAAT sequence for 70 (6 -10). These results suggest that promoter selectivity between 70 and S might be determined by factors other than promote...

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Section: Discussionsupporting

confidence: 88%

mentioning

confidence: 99%

Nucleotides from –16 to –12 Determine Specific Promoter Recognition by Bacterial σS-RNA Polymerase

Lacour

Kolb

Landini

2003

Journal of Biological Chemistry

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“…4a). Consistent with these results, the sequence upstream of the mscL transcription start site contained a good match to the consensus sequence recognized by both E S and E 70 (TGtTAGAA͞CT) (20). There was no discernible Ϫ35 hexamer, but it is known that TG immediately upstream of the Ϫ10 can obviate the need for a strong Ϫ35 sequence (32) (Fig.…”

Section: Rpos Is Required For Osmotic Induction Of Ms Channel Genessupporting

confidence: 68%

“…For in vivo mapping, strains containing pRLG6984 (mscL) and pRLG6985 (mscS) were grown at 37°C in LB to early stationary phase (A 600 Ϸ 1.0), RNA was extracted by using a boiling lysis procedure, and reverse transcription was performed as described (18,19). For in vitro mapping, transcripts were synthesized from pRLG6984 (mscL) and pRLG6985 (mscS) by using purified RNA polymerase containing either 70 or S , (E 70 and E S , respectively), which were prepared as described (20). Because saturating amounts of (either S or 70 ) were added to the same amount of core enzyme, the two holoenzymes have the same specific activity.…”

Section: Methodsmentioning

confidence: 99%

A role for mechanosensitive channels in survival of stationary phase: Regulation of channel expression by RpoS

Stokes

Murray

Subramaniam

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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The mechanosensitive (MS) channels MscS and MscL are essential for the survival of hypoosmotic shock by Escherichia coli cells. We demonstrate that MscS and MscL are induced by osmotic stress and by entry into stationary phase. Reduced levels of MS proteins and reduced expression of mscL-and mscS-LacZ fusions in an rpoS mutant strain suggested that the RNA polymerase holoenzyme containing S is responsible, at least in part, for regulating production of MS channel proteins. Consistent with the model that the effect of S is direct, the MscS and MscL promoters both use RNA polymerase containing S in vitro. Conversely, clpP or rssB mutations, which cause enhanced levels of S , show increased MS channel protein synthesis. RpoS null mutants are sensitive to hypoosmotic shock upon entry into stationary phase. These data suggest that MscS and MscL are components of the RpoS regulon and play an important role in ensuring structural integrity in stationary phase bacteria.

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“…3B). We next tested whether 38 , which shares the Ϫ10 recognition determinants (19) with 70 , will recognize the Ϫ10 element as a pause signal. In agreement with ref.…”

Section: Figmentioning

confidence: 99%

The elongation factor RfaH and the initiation factor σ bind to the same site on the transcription elongation complex

Sevostyanova

Svetlov

Vassylyev

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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RNA polymerase is a target for numerous regulatory events in all living cells. Recent studies identified a few ''hot spots'' on the surface of bacterial RNA polymerase that mediate its interactions with diverse accessory proteins. Prominent among these hot spots, the ␤ subunit clamp helices serve as a major binding site for the initiation factor and for the elongation factor RfaH. Furthermore, the two proteins interact with the nontemplate DNA strand in transcription complexes and thus may interfere with each other's activity. We show that RfaH does not inhibit transcription initiation but, once recruited to RNA polymerase, abolishes -dependent pausing. We argue that this apparent competition is due to a steric exclusion of by RfaH that is stably bound to the nontemplate DNA and clamp helices, both of which are necessary for the recruitment to the transcription complex. Our findings highlight the key regulatory role played by the clamp helices during both initiation and elongation stages of transcription.clamp helices ͉ RNA polymerase ͉ transcription factor ͉ nontemplate DNA B acterial RNA polymerase (RNAP) is a principal target for numerous accessory proteins and small ligands that finetune gene expression profiles to match the cell needs. Competition (or cooperation) among these regulators for the finite number of targets on the RNAP surface determines the patterns of gene expression. The classical paradigm for the partitioning of the regulatory space is competition (1) with different initiation factors competing for binding to the core enzyme (subunit composition ␣ 2 ␤␤Ј ) and, when successful, directing it to a subset of -specific promoters. The -subunit makes many contacts to the core RNAP among which the ␤Ј subunit clamp helices (␤Ј CH, a coiled-coil motif comprising residues 260-309 in the Escherichia coli enzyme) are thought to constitute the major binding site in the free RNAP (2, 3) as well as in the transcription elongation complex (TEC) (4). Our recent finding that the ␤Ј CH is also required for recruitment of the elongation factor RfaH (5) suggested that competition for this site may regulate gene expression far beyond -specific promoter recognition.RfaH reduces pausing and termination thereby suppressing transcriptional polarity in long operons encoding virulence and fertility determinants (6, 7). RfaH action depends on the ops DNA sequence (GGCGGTAGnnTG) elements located in the transcribed regions of RfaH-controlled operons (7). In vitro, the ops element indeed mediates RfaH binding to the TEC but only if it is placed in the nontemplate (NT) DNA strand exposed on the surface of RNAP (7). RfaH recruitment is thought to occur in two steps: (i) sequence-specific binding of the N-terminal domain to DNA triggers displacement of the stably bound C-terminal domain to expose the RNAP binding site on the N-domain, and (ii) interactions of the N-domain with ␤Ј CH on one side and (nonspecific) interactions with the NT strand on the other allow for the stable retention of RfaH on the TEC throughout elongation...

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Promoter recognition and discrimination by Eσ^S RNA polymerase

Abstract: thus presents an alternative solution to the problem of promoter selectivity.

Cited by 161 publications

References 66 publications

Nucleotides from –16 to –12 Determine Specific Promoter Recognition by Bacterial σS-RNA Polymerase

Nucleotides from –16 to –12 Determine Specific Promoter Recognition by Bacterial σS-RNA Polymerase

A role for mechanosensitive channels in survival of stationary phase: Regulation of channel expression by RpoS

The elongation factor RfaH and the initiation factor σ bind to the same site on the transcription elongation complex

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Promoter recognition and discrimination by EσS RNA polymerase

Abstract: thus presents an alternative solution to the problem of promoter selectivity.

Cited by 161 publications

References 66 publications

Nucleotides from –16 to –12 Determine Specific Promoter Recognition by Bacterial σS-RNA Polymerase

Nucleotides from –16 to –12 Determine Specific Promoter Recognition by Bacterial σS-RNA Polymerase

A role for mechanosensitive channels in survival of stationary phase: Regulation of channel expression by RpoS

The elongation factor RfaH and the initiation factor σ bind to the same site on the transcription elongation complex

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Promoter recognition and discrimination by Eσ^S RNA polymerase