IgaA negatively regulates the Rcs Phosphorelay via contact with the RcsD Phosphotransfer Protein

Autoři: Erin A. Wall aff001;  Nadim Majdalani aff001;  Susan Gottesman aff001
Působiště autorů: National Cancer Institute, Bethesda, Maryland, United States of America aff001
Vyšlo v časopise: IgaA negatively regulates the Rcs Phosphorelay via contact with the RcsD Phosphotransfer Protein. PLoS Genet 16(7): e32767. doi:10.1371/journal.pgen.1008610
Kategorie: Research Article
doi: 10.1371/journal.pgen.1008610


Two-component systems and phosphorelays play central roles in the ability of bacteria to rapidly respond to changing environments. In E. coli and related enterobacteria, the complex Rcs phosphorelay is a critical player in the bacterial response to antimicrobial peptides, beta-lactam antibiotics, and other disruptions at the cell surface. The Rcs system is unusual in that an inner membrane protein, IgaA, is essential due to its negative regulation of the RcsC/RcsD/RcsB phosphorelay. While it is known that IgaA transduces signals from the outer membrane lipoprotein RcsF, how it interacts with the phosphorelay has remained unknown. Here we performed in vivo interaction assays and genetic dissection of the critical proteins and found that IgaA interacts with the phosphorelay protein RcsD, and that this interaction is necessary for regulation. Interactions between IgaA and RcsD within their respective periplasmic domains of these two proteins anchor repression of signaling. However, the signaling response depends on a second interaction between cytoplasmic loop 1 of IgaA and a truncated Per-Arndt-Sim (PAS-like) domain in RcsD. A single point mutation in the PAS-like domain increased interactions between the two proteins and blocked induction of the phosphorelay. IgaA may regulate RcsC, the histidine kinase that initiates phosphotransfer through the phosphorelay, indirectly, via its contacts with RcsD. Unlike RcsD, and unlike many other histidine kinases, the periplasmic domain of RcsC is dispensable for the response to signals that induce the Rcs phosphorelay system. The multiple contacts between IgaA and RcsD constitute a poised sensing system, preventing potentially toxic over-activation of this phosphorelay while enabling it to rapidly and quantitatively respond to signals.

Klíčová slova:

Arabinose – Cell fusion – Glucose – Outer membrane proteins – Phosphates – Plasmid construction – Point mutation – Protein interactions


1. Wall E, Majdalani N, Gottesman S. The Complex Rcs Regulatory Cascade. Annu Rev Microbiol. 2018;72:111–39. doi: 10.1146/annurev-micro-090817-062640 29897834

2. Guo XP, Sun Y-C. New insights in the non-orthodox two component Rcs Phosphorelay system. Frontiers in Microbiology. 2017;8. doi: 10.3389/fmicb.2017.02014 29089936

3. Cano DA, Martinez-Moya M, Pucciarelli MG, Groisman EA, Casadesus J, Portillo FG-D. Salmonella enterica Serovar Typhimurium response involved in attenuation of pathogen intracellular proliferation. Infect Immun. 2001;69:6463–74. doi: 10.1128/IAI.69.10.6463-6474.2001 11553591

4. Cano DA, Dominguez-Bernal G, Tierrez A, Garcia-Del Portillo F, Casadesus J. Regulation of capsule synthesis and cell motility in Salmonella enterica by the essential gene igaA. Genetics. 2002;162(4):1513–23. 12524328.

5. Cho S-H, Szewczyk J, Pesavento C, Zietek M, Banzhaf M, Roszczenko P, et al. Detecting envelope stress by monitoring B-barrel assembly. Cell. 2014;159:1652–64. doi: 10.1016/j.cell.2014.11.045 25525882

6. Konovalova A, Mitchell AM, Silhavy TJ. A lipoprotein/b-barrel complex monitors lipopolysaccharide integrity transducing information across the outer membrane. Elife. 2016. doi: 10.7554/eLife.15276 [27282389

7. Konovalova A, Perlman DH, Cowles CE, Silhavy TJ. Transmembrane domain of surface-exposed outer membrane lipoprotein RcsF is threaded through the lumen of B-barrel proteins. Proc Natl Acad Sci USA. 2014;111:E4350–E8. doi: 10.1073/pnas.1417138111 25267629

8. Laloux G, Collet J-F. Major Tom to Ground Control: How Lipoproteins Communicate Extracytoplasmic Stress to the Decision Center of the Cell. Journal of Bacteriology. 2017;199(21):1–13. doi: 10.1128/JB

9. Asmar AT, Ferreira JL, Cohen EJ, Cho S-H, Beeby M, Hughes KT, et al. Communication across the bacterial cell envelope depends on the size of the periplasm. PLoS Biology. 2017. doi: 10.1371/journal.pbio.2004303

10. Hussein NA, Cho S-H, Laloux G, Siam R, Collet J-F. Distinct domains of Escherichia coli IgaA connect envelope stress sensing and down-regulation of the Rcs phosphorelay across subcellular compartments. PLoS Genet. 2018;14:e1007398. doi: 10.1371/journal.pgen.1007398 29852010

11. Majdalani N, Hernandez D, Gottesman S. Regulation and mode of action of the second small RNA activator of RpoS translation, RprA. Mol Microbiol. 2002;46(3):813–26. doi: 10.1046/j.1365-2958.2002.03203.x 12410838

12. Castanie-Cornet M-P, Cam K, Jacq A. RcsF is an outer membrane lipoprotein involved in the RcsCDB phosphorelay signaling pathway in Escherichia coli. J Bacteriol. 2006;188:4264–70. doi: 10.1128/JB.00004-06 16740933

13. Majdalani N, Heck M, Stout V, Gottesman S. Role of RcsF in Signaling to the Rcs Phosphorelay Pathway in Escherichia coli. Journal of Bacteriology. 2005;187(19):6770–8. doi: 10.1128/JB.187.19.6770-6778.2005 16166540

14. Shiba Y, Miyagawa H, Nagahama H, Matsumoto K, Kondo D, Matsuoka S, et al. Exploring the relationship between lipoprotein mislocalization and activation of the Rcs signal transduction system in Escherichia coli. Microbiology. 2012;158(5):1238–48. doi: 10.1099/mic.0.056945–0

15. Shiba Y, Matsumoto K, Hara H. DjlA negatively regulates the Rcs signal transduction system in Escherichia coli. Genes Genet Syst. 2006;81:51–6. doi: 10.1266/ggs.81.51 16607041

16. Ferrieres L, Thompson A, Clarke DJ. Elevated levels of sigma S inhibit biofilm formation in Escherichia coli: a role for the Rcs phosphorelay. Microbiology. 2009;155(Pt 11):3544–53. doi: 10.1099/mic.0.032722-0 19696107.

17. Fredericks CE, Shibata S, Aizawa S-I, Reimann SA, Wolfe AJ. Acetyl phosphate-sensitive regulation of flagellar biogenesis and capsular biosynthesis depends on the Rcs phosphorelay. Molecular Microbiology. 2006;61(3):734–47. doi: 10.1111/j.1365-2958.2006.05260.x 16776655

18. Clarke DJ, Joyce SA, Toutain CM, Jacq A, Holland IB. Genetic analysis of the RcsC sensor kinase from Escherichia coli K-12. J Bacteriol. 2002;184:1204–8. doi: 10.1128/jb.184.4.1204-1208.2002 11807084

19. De Mets F, Van Melderen L, Gottesman S. Regulation of acetate metabolism and coordination with the TCA cycle via a processed small RNA. Proc Natl Acad Sci USA. 2019;116:1043–52. doi: 10.1073/pnas.1815288116 30591570

20. Karimova G, Pidoux J, Ullmann A, Ladant D. A bacterial two-hybrid system based on a reconstituted signal transduction pathway. Proc Natl Acad Sci USA. 1998;95(10):5752–6. doi: 10.1073/pnas.95.10.5752 9576956.

21. Battesti A, Bouveret E. The bacterial two-hybrid system based on adenylate cyclase reconstitution in Escherichia coli. Methods. 2012;58(4):325–34. doi: 10.1016/j.ymeth.2012.07.018 22841567

22. Zschiedrich C, Keidel V, Szurmant H. Molecular mechanisms of two-component signal transduction. Journal of Molecular Biology. 2016;428(19):3752–75. doi: 10.1016/j.jmb.2016.08.003 27519796

23. Schmöe K, Rogov VV, Rogova NY, Löhr F, Güntert P, Bernhard F, et al. Structural insights into Rcs phosphotransfer: The newly identified RcsD-ABL domain enhances interaction with the response regulator RcsB. Structure. 2011;19(4):577–87. doi: 10.1016/j.str.2011.01.012 21481780

24. Takeda S, Fujisawa Y, Matsubara M, Aiba H, Mizuno T. A novel feature of the multistep phosphorelay in Escherichia coli: a revised model of the RcsC—> YojN—>RcsB signalling pathway implicated in capsular synthesis and swarming behavior. Mol Microbiol. 2001;40:440–50. doi: 10.1046/j.1365-2958.2001.02393.x 11309126

25. Laubacher ME, Ades SE. The Rcs Phosphorelay Is a Cell Envelope Stress Response Activated by Peptidoglycan Stress and Contributes to Intrinsic Antibiotic Resistance. Journal of Bacteriology. 2008;190(6):2065–74. doi: 10.1128/JB.01740-07 18192383

26. Domínguez-Bernal G, Pucciarelli MG, Ramos-Morales F, García-Quintanilla M, Cano DA, Casadesús J, et al. Repression of the RcsC-YojN-RcsB phosphorelay by the IgaA protein is a requisite for Salmonella virulence. Molecular Microbiology. 2004;53(5):1437–49. doi: 10.1111/j.1365-2958.2004.04213.x 15387821

27. Clarke DJ, Holland IB, Jacq A. Point mutations in the transmembrane domain of DjlA, a membrane-linked DnaJ-like protein, abolish its function in promoting colanic acid production via the Rcs signal transduction pathway. Mol Microbiol. 1997;25:933–44. doi: 10.1111/j.1365-2958.1997.mmi528.x 9364918

28. Yu D, Ellis HM, Lee EC, Jenkins NA, Copeland NG, Court DL. An efficient recombination system for chromosome engineering in Escherichia coli. Proc Natl Acad Sci USA. 2000;97(11):5978–83. doi: 10.1073/pnas.100127597 10811905.

29. Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, et al. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol Syst Biol [Internet]. 2006; 2:[2006.0008 p.].

30. Khlebnikov A, Datsenko KA, Skaug T, Wanner BL, Keasling JD. Homogeneous expression of the P(BAD) promoter in Escherichia coli by constitutive expression of the low-affinity high-capacity AraE transporter. Microbiology. 2001;147(Pt 12):3241–7. doi: 10.1099/00221287-147-12-3241 11739756.

31. Krin E, Danchin A, Soutourina O. RcsB plays a central role in H-NS-dependent regulation of motility and acid stress resistance in Escherichia coli. Res Microbiol. 2010;161:363–71. doi: 10.1016/j.resmic.2010.04.002 20435136

32. Pescaretti MdlM, Farizano JV, Morero R, Delgado MA. A Novel Insight on Signal Transduction Mechanism of RcsCDB System in Salmonella enterica Serovar Typhimurium. PLoS ONE. 2013;8(9):1–10. doi: 10.1371/journal.pone.0072527 24023746

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PLOS Genetics

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