skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Structural Basis for DNA Recognition by the Two-Component Response Regulator RcsB

Abstract

RcsB is a highly conserved transcription regulator of the Rcs phosphorelay system, a complex two-component signal transduction system (N. Majdalani and S. Gottesman, Annu Rev Microbiol 59:379–405, 2005; A. J. Wolfe, Curr Opin Microbiol 13:204–209, 2010,https://doi.org/10.1016/j.mib.2010.01.002; D. J. Clarke, Future Microbiol 5:1173–1184, 2010,https://doi.org/10.2217/fmb.10.83). RcsB plays an important role in virulence and pathogenicity in human hosts by regulating biofilm formation. RcsB can regulate transcription alone or together with its auxiliary transcription regulators by forming heterodimers. This complexity allows RcsB to regulate transcription of more than 600 bacterial genes in response to different stresses (D. Wang et al., Mol Plant Microbe Interact 25:6–17, 2012,https://doi.org/10.1094/MPMI-08-11-0207). Despite increasing knowledge of RcsB importance, molecular mechanisms that drive the ability of RcsB to control transcription of a large number of genes remain unclear. Here, we present crystal structures of unphosphorylated RcsB in complex with the consensus DNA-binding sequence of 22-mer (DNA22) and 18-mer (DNA18) of the flhDC operon from Escherichia coli determined at 3.15- and 3.37-Å resolution, respectively. The results of our structural analysis combined with the results of in vitro binding assays provide valuable insights to the protein regulatory mechanism, demonstrate how RcsB recognizes target DNA sequences, and reveal a unique oligomeric state that allowsmore » RcsB to form homo- and heterodimers. This information will help us understand the complex mechanisms of transcriptional regulation by RcsB in bacteria.« less

Authors:
 [1];  [2];  [3];  [2];  [1];
  1. Center for Structural Genomics of Infectious Diseases, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA, Department of Biochemistry and Molecular Genetics, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA
  2. Department of Microbiology and Immunology, Loyola University Chicago, Health Sciences Division, Stritch School of Medicine, Maywood, Illinois, USA
  3. Keck Biophysics Facility, Northwestern University, Evanston, Illinois, USA
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States); Univ. of Illinois at Urbana-Champaign, IL (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER); NIAID; National Institutes of Health (NIH); HHS
OSTI Identifier:
1493948
Alternate Identifier(s):
OSTI ID: 1499720
Grant/Contract Number:  
SC0012443; NIH R01AI083640; NIH 2R01AI083640-06A1; R01AI083640; 2R01AI083640-06A1; AC02-06CH11357
Resource Type:
Journal Article: Published Article
Journal Name:
mBio
Additional Journal Information:
Journal Name: mBio Journal Volume: 9 Journal Issue: 1; Journal ID: ISSN 2161-2129
Publisher:
American Society for Microbiology
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Filippova, Ekaterina V., Zemaitaitis, Bozena, Aung, Theint, Wolfe, Alan J., Anderson, Wayne F., and Hendrickson, ed., Wayne A. Structural Basis for DNA Recognition by the Two-Component Response Regulator RcsB. United States: N. p., 2018. Web. doi:10.1128/mBio.01993-17.
Filippova, Ekaterina V., Zemaitaitis, Bozena, Aung, Theint, Wolfe, Alan J., Anderson, Wayne F., & Hendrickson, ed., Wayne A. Structural Basis for DNA Recognition by the Two-Component Response Regulator RcsB. United States. https://doi.org/10.1128/mBio.01993-17
Filippova, Ekaterina V., Zemaitaitis, Bozena, Aung, Theint, Wolfe, Alan J., Anderson, Wayne F., and Hendrickson, ed., Wayne A. 2018. "Structural Basis for DNA Recognition by the Two-Component Response Regulator RcsB". United States. https://doi.org/10.1128/mBio.01993-17.
@article{osti_1493948,
title = {Structural Basis for DNA Recognition by the Two-Component Response Regulator RcsB},
author = {Filippova, Ekaterina V. and Zemaitaitis, Bozena and Aung, Theint and Wolfe, Alan J. and Anderson, Wayne F. and Hendrickson, ed., Wayne A.},
abstractNote = {RcsB is a highly conserved transcription regulator of the Rcs phosphorelay system, a complex two-component signal transduction system (N. Majdalani and S. Gottesman, Annu Rev Microbiol 59:379–405, 2005; A. J. Wolfe, Curr Opin Microbiol 13:204–209, 2010,https://doi.org/10.1016/j.mib.2010.01.002; D. J. Clarke, Future Microbiol 5:1173–1184, 2010,https://doi.org/10.2217/fmb.10.83). RcsB plays an important role in virulence and pathogenicity in human hosts by regulating biofilm formation. RcsB can regulate transcription alone or together with its auxiliary transcription regulators by forming heterodimers. This complexity allows RcsB to regulate transcription of more than 600 bacterial genes in response to different stresses (D. Wang et al., Mol Plant Microbe Interact 25:6–17, 2012,https://doi.org/10.1094/MPMI-08-11-0207). Despite increasing knowledge of RcsB importance, molecular mechanisms that drive the ability of RcsB to control transcription of a large number of genes remain unclear. Here, we present crystal structures of unphosphorylated RcsB in complex with the consensus DNA-binding sequence of 22-mer (DNA22) and 18-mer (DNA18) of the flhDC operon from Escherichia coli determined at 3.15- and 3.37-Å resolution, respectively. The results of our structural analysis combined with the results of in vitro binding assays provide valuable insights to the protein regulatory mechanism, demonstrate how RcsB recognizes target DNA sequences, and reveal a unique oligomeric state that allows RcsB to form homo- and heterodimers. This information will help us understand the complex mechanisms of transcriptional regulation by RcsB in bacteria.},
doi = {10.1128/mBio.01993-17},
url = {https://www.osti.gov/biblio/1493948}, journal = {mBio},
issn = {2161-2129},
number = 1,
volume = 9,
place = {United States},
year = {Wed Mar 07 00:00:00 EST 2018},
month = {Wed Mar 07 00:00:00 EST 2018}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record at https://doi.org/10.1128/mBio.01993-17

Citation Metrics:
Cited by: 10 works
Citation information provided by
Web of Science

Figures / Tables:

FIG 1 FIG 1: Oligomeric state and DNA binding activity of RcsB. (A) EMSA of RcsB against DNA22. Lane 1, free DNA; lane 2, DNA in the presence of RcsB; lane 3, DNA in the presence of RcsB and carbamoyl phosphorylate (CPh); lane 4, DNA in the presence of RcsB and phosphoramidemore » (PA); lane 5, DNA in the presence of RcsB, CPh, and unlabeled mutated (mut) DNA22; lane 6, DNA in the presence of RcsB, CPh, and unlabeled (unlab) wild-type DNA22. HMM complex, higher-molecularmass complex. (B) The SEC-MALS elution profiles of RcsB and RcsB in the presence of phosphodonor (RcsB*). The horizontal bold lines show the determined molecular masses (MM). The theoretical MM of the RcsB monomer is 24 kDa. (C) The SEC-MALS elution profiles of RcsB and RcsB* (with phosphodonor) in the presence of DNA22. The horizontal bold lines show the calculated MM of the RcsB-DNA complex. The theoretical MM of the RcsB dimer bound to the 22-bp DNA duplex is 63 kDa. The second peak eluted around 31 min corresponds to unbound DNA. (D and E) SPR sensograms (colored curves) of unphosphorylated and carbamoyl-phosphorylated RcsB and DNA22, respectively. KD values were calculated based on a 1:1 kinetic model, in which one RcsB dimer interacts with one double-stranded DNA. The fitting curves are shown in black. The corresponding concentrations of RcsB are indicated above the SPR curves. (F) RcsB-binding site from the flhDC promoter and DNA22 and DNA18 sequences.« less

Save / Share:

Works referenced in this record:

Salmonella Biofilm Development Depends on the Phosphorylation Status of RcsB
journal, May 2012


Regulation and mode of action of the second small RNA activator of RpoS translation, RprA: RprA regulation and mode of action
journal, October 2002


Structure of the response regulator ChrA in the haem-sensing two-component system of Corynebacterium diphtheriae
journal, July 2015


RcsCDB His-Asp phosphorelay system negatively regulates the flhDC operon in Escherichia coli: Regulation of flhDC by RcsB
journal, August 2003


Nε−Lysine Acetylation of a Bacterial Transcription Factor Inhibits Its DNA-Binding Activity
journal, December 2010


Dali server: conservation mapping in 3D
journal, May 2010


Transcriptional regulation by BglJ–RcsB, a pleiotropic heteromeric activator in Escherichia coli
journal, December 2013


DNA-binding proteins
journal, September 1983


Structure of the Escherichia coli Response Regulator NarL ,
journal, January 1996


An asymmetric heterodomain interface stabilizes a response regulator–DNA complex
journal, February 2014


Phosphorylation-dependent conformational changes and domain rearrangements in Staphylococcus aureus VraR activation
journal, May 2013


Acetyl phosphate-sensitive regulation of flagellar biogenesis and capsular biosynthesis depends on the Rcs phosphorelay
journal, August 2006


Fine-structure mapping and identification of two regulators of capsule synthesis in Escherichia coli K-12.
journal, June 1988


The importance of the Rcs phosphorelay in the survival and pathogenesis of the enteropathogenic yersiniae
journal, April 2008


Genome Expression Analyses Revealing the Modulation of the Salmonella Rcs Regulon by the Attenuator IgaA
journal, January 2009


Signal transduction in bacteria: molecular mechanisms of stimulus—response coupling
journal, April 1998


Detecting Envelope Stress by Monitoring β-Barrel Assembly
journal, December 2014


Dimerization allows DNA target site recognition by the NarL response regulator
journal, September 2002


Acid stress response in Escherichia coli: mechanism of regulation of gadA transcription by RcsB and GadE
journal, February 2010


Non-canonical activation of OmpR drives acid and osmotic stress responses in single bacterial cells
journal, November 2017


Structure of a bacterial quorum-sensing transcription factor complexed with pheromone and DNA
journal, June 2002


Structural basis of DNA sequence recognition by the response regulator PhoP in Mycobacterium tuberculosis
journal, April 2016


Interaction of the RcsB Response Regulator with Auxiliary Transcription Regulators in Escherichia coli
journal, December 2015


Structural Analysis of the DNA-binding Domain of the Erwinia amylovora RcsB Protein and Its Interaction with the RcsAB Box
journal, March 2003


Acetylation of the Response Regulator RcsB Controls Transcription from a Small RNA Promoter
journal, July 2013


Figures / Tables found in this record:

Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.