Roles for the C-terminal Region of Sigma 54 in Transcriptional Silencing and DNA Binding

Wang, Lei; Gralla, Jay D.

doi:10.1074/jbc.m009587200

Cited by 19 publications

(29 citation statements)

References 40 publications

(51 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Our results here establish that the RpoN box in Region III (which, based on secondary structure predictions, forms an ␣-helical structure) is proximal to the Ϫ24 promoter region. The loss of function mutations in the RpoN box strengthens this view (16,20). The propensity of basic and hydrophobic amino acids in the RpoN box is consistent with it directly interacting with DNA.…”

Section: Discussionmentioning

confidence: 55%

“…The domains and residues involved in core RNAP, activator, and DNA interactions are indicated. In Region III, boxed are residues with potential roles in DNA interactions: the putative helix-turn-helix (HTH) motif (gray box) (19,55), a region that has been shown to cross-link to DNA (black box) (56), and the RpoN box motif (white box) (16,20). The regions that were targeted for introducing a cysteine residue for FeBABE conjugation (residues 39 -48 and 453-476) are expanded and aligned with corresponding regions in 54 BABE, which cleave DNA and polypeptide chains in its proximity (ϳ12 Å plus the 3 Å diffusion distance of the hydroxyl radicals) (28).…”

Section: Creation Of Functional 54 -Febabe Derivativesmentioning

confidence: 99%

“…We have introduced unique cysteines into several positions of the altered K. pneumoniae rpoN. Sites were chosen based on data that indicated that these positions would result in neither complete inactivation nor deregulation of 54 (16,20,34). Unique cysteines were introduced at a single position in Region I (Leu 46 ) and at three positions in the RpoN box (Arg 455 , Lys 460 , and Glu 463 ) (Fig.…”

Section: Creation Of Functional 54 -Febabe Derivativesmentioning

confidence: 99%

“…Recognition of the Ϫ12 GC-region, where DNA melting originates, involves 54 Region I and Region III sequences (16 -19). The RpoN box, a signature sequence found in 54 , has been proposed to interact with the Ϫ24 GG-region, but no physical or genetic suppression evidence has been obtained to directly establish this view (16,20).…”

mentioning

confidence: 99%

See 3 more Smart Citations

Mapping σ54-RNA Polymerase Interactions at the –24 Consensus Promoter Element

Burrows

Severinov

Ishihama

et al. 2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

The 54 promoter specificity factor is distinct from 70 -type factors. The 54 -RNA polymerase binds to promoters with conserved sequence elements at ؊24 and ؊12 and utilizes specialized enhancer-binding activators to convert, through an ATP-dependent process, closed promoter complexes to open promoter complexes. The interface between 54 -RNA polymerase and promoter DNA is poorly characterized, contrasting with 70 . Here, 54 was modified with strategically positioned cleavage reagents to provide physical evidence that the highly conserved RpoN box motif of 54 is close to and may therefore interact with the consensus ؊24 promoter element. We show that the spatial relationship between the 54 -RNA polymerase and the ؊24 promoter element remains unchanged during closed to open complex conversion and transcription initiation but changes during the early elongation phase. In contrast, the spatial relationship between 54 -RNA polymerase and the consensus ؊12 promoter element changes upon conversion of the closed promoter complex to an open one. We provide evidence that some ؊12 promoter region-54 interactions are dependent upon either the core RNA polymerase or a fork junction DNA structure at the ؊12-position, indicating that DNA fork junctions can substitute for core RNAP. We also show the ␤-subunit flap domain contributes to different sets of -promoter DNA interactions at 54 -and 70 -dependent promoters.Transcription of DNA is a fundamental process needed for regulation of cellular adaptation and differentiation and is carried out by DNA-dependent RNA polymerases (RNAPs).

show abstract

Section: Discussionmentioning

confidence: 55%

Section: Creation Of Functional 54 -Febabe Derivativesmentioning

confidence: 99%

Section: Creation Of Functional 54 -Febabe Derivativesmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Mapping σ54-RNA Polymerase Interactions at the –24 Consensus Promoter Element

Burrows

Severinov

Ishihama

et al. 2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…This helix is primarily positively charged 54 C domain (residues Ala-335 to Gly-389). Side chains for residues that were previously shown to be near the Ϫ24 promoter element (50) and/or to decrease significantly DNA binding when mutated (48) are represented as sticks and labeled (except for Tyr-384, which is hidden behind Lys-383). All of these residues are on helix 3 (dark blue) created by the highly conserved RpoN box, except for Met-343 (Arg-421 in E. coli), which is on helix 1.…”

Section: Resultsmentioning

confidence: 99%

The C-terminal RpoN Domain of σ54 Forms an Unpredicted Helix-Turn-Helix Motif Similar to Domains of σ70

Doucleff

Malak

Wemmer

2005

Journal of Biological Chemistry

View full text Add to dashboard Cite

The "" subunit of prokaryotic RNA polymerase allows genespecific transcription initiation. Two families have been identified, 70 and 54 , which use distinct mechanisms to initiate transcription and share no detectable sequence homology. Although the 70 -type factors have been well characterized structurally by x-ray crystallography, no high resolution structural information is available for the 54 -type factors. Here we present the NMR-derived structure of the C-terminal domain of 54 from Aquifex aeolicus. This domain (Thr-323 to Gly-389), which contains the highly conserved RpoN box sequence, consists of a poorly structured N-terminal tail followed by a three-helix bundle, which is surprisingly similar to domains of the 70 -type proteins. Residues of the RpoN box, which have previously been shown to be critical for DNA binding, form the second helix of an unpredicted helix-turn-helix motif. The homology of this structure with other DNA-binding proteins, combined with previous biochemical data, suggests how the C-terminal domain of 54 binds to DNA.Transcription, the synthesis of RNA from double-stranded DNA, is a fundamental process in all forms of life. The primary protein complex catalyzing transcription is composed of five subunits, ␣ 2 ␤Ј␤, called "core RNA polymerase" (RNAP).2 Core RNAP is fully competent to synthesize RNA from DNA. However, the initiation of transcription at specific DNA sequences requires additional protein(s). In bacteria, these additional proteins are called the " factors" (1, 2). The factor facilitates transcription at specific DNA sequences by 1) binding to the core RNAP to form the -RNAP holoenzyme, 2) recognizing and binding to a specific DNA sequence next to the transcription start site, called the promoter element, and 3) opening the double-stranded DNA to initiate transcription.Based on protein sequence homology, factors can be grouped into two classes, 70 and 54 , which were named by the molecular weights of the first members identified (reviewed in Ref. 1 is involved in regulating the kinetics of transcription initiation; 2 binds to the Ϫ10 promoter element and is essential for melting the DNA;3 stabilizes the open complex formation by binding the extended Ϫ10 element; and 4 binds specifically to the Ϫ35 promoter element (reviewed in Ref. 17).In contrast to 70 , 54 is divided into three regions based on function (18 -20). Region 1 (E. coli-(1-55)) interacts with the upstream activator protein to control promoter melting; region 2 (E. coli-(70 -304)) contains the minimal RNAP-binding domain (E. coli-(70 -180) and a part that enhances DNA binding affinity (E. coli-(180 -304)); region 3 (E. coli-(329 -477)) recognizes and binds the consensus promoter elements. This C-terminal part of the protein contains a region that crosslinks to DNA (E. coli-(329 -346)), a predicted helix-turn-helix motif (E. coli-(366 -386); Fig. 1), and a highly conserved sequence (E. coli-* This work was supported by National Institutes of Health Grant GM62163 (to D. E. W.) and by a University of Calif...

show abstract

Inhibition of Bacterial Gene Transcription with an RpoN-Based Stapled Peptide

Payne

Pau

Whiting

et al. 2018

Cell Chemical Biology

View full text Add to dashboard Cite

In response to environmental and other stresses, the σ subunit of bacterial RNA polymerase (RNAP) controls expression of several genes that play a significant role in the virulence of both plant and animal pathogens. Recruitment of σ to RNAP initiates promoter-specific transcription via the double-stranded DNA denaturation mechanism of the cofactor. The RpoN box, a recognition helix found in the C-terminal region of σ, has been identified as the component necessary for major groove insertion at the -24 position of the promoter. We employed the hydrocarbon stapled peptide methodology to design and synthesize stapled σ peptides capable of penetrating Gram-negative bacteria, binding the σ promoter, and blocking the interaction between endogenous σ and its target DNA sequence, thereby reducing transcription and activation of σ response genes.

show abstract

Roles for the C-terminal Region of Sigma 54 in Transcriptional Silencing and DNA Binding

Cited by 19 publications

References 40 publications

Mapping σ54-RNA Polymerase Interactions at the –24 Consensus Promoter Element

Mapping σ54-RNA Polymerase Interactions at the –24 Consensus Promoter Element

The C-terminal RpoN Domain of σ54 Forms an Unpredicted Helix-Turn-Helix Motif Similar to Domains of σ70

Inhibition of Bacterial Gene Transcription with an RpoN-Based Stapled Peptide

Contact Info

Product

Resources

About