A connecter-like factor, CacA, links RssB/RpoS and the CpxR/CpxA two-component system in Salmonella

BMC Microbiology

Ryutaro Utsumi¹  Kei Hagihara¹  Haruka Emori¹  Wataru Nomura¹  Hironori Hayashi¹  Akinori Kato¹ 

[1] Department of Advanced Bioscience, Graduate School of Agriculture, Kinki University, Nakamachi, Nara 631–8505, 3327-204, Japan

关键词: CpxR/CpxA;    CacA;    RpoS;    RssB;    Network;    Connector;    Two-component system;   

Others  :  1221729
DOI  :  10.1186/1471-2180-12-224

 received in 2012-07-11, accepted in 2012-09-21,  发布年份 2012

【 摘 要 】

Background

Bacteria integrate numerous environmental stimuli when generating cellular responses. Increasing numbers of examples describe how one two-component system (TCS) responds to signals detected by the sensor of another TCS. However, the molecular mechanisms underlying this phenomenon remain poorly defined.

Results

Here, we report a connector-like factor that affects the activity of the CpxR/CpxA two-component system in Salmonella enterica serovar Typhimurium. We isolated a clone that induced the expression of a cpxP-lac gene fusion from a high-copy-number plasmid pool of random Salmonella genomic fragments. A 63-amino acid protein, CacA, was responsible for the CpxA/CpxR-dependent activation of the cpxP gene. The CpxR-activated genes cpxP and spy exhibited approximately 30% and 50% reductions in transcription, respectively, in a clean cacA deletion mutant strain in comparison to wild-type. From 33 response regulator (RR) deletion mutants, we identified that the RssB regulator represses cacA transcription. Substitution mutations in a conserved -10 region harboring the RNA polymerase recognition sequence, which is well conserved with a known RpoS -10 region consensus sequence, rendered the cacA promoter RpoS-independent. The CacA-mediated induction of cpxP transcription was affected in a trxA deletion mutant, which encodes thioredoxin 1, suggesting a role for cysteine thiol-disulfide exchange(s) in CacA-dependent Cpx activation.

Conclusions

We identified CacA as an activator of the CpxR/CpxA system in the plasmid clone. We propose that CacA may integrate the regulatory status of RssB/RpoS into the CpxR/CpxA system. Future investigations are necessary to thoroughly elucidate how CacA activates the CpxR/CpxA system.

【 授权许可】

   
2012 Kato et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

【 参考文献 】

• [1]Ulrich LE, Zhulin IB: The MiST2 database: a comprehensive genomics resource on microbial signal transduction. Nucleic Acids Res 2010, 38(Database issue):D401-407.
• [2]Laub MT, Goulian M: Specificity in two-component signal transduction pathways. Annu Rev Genet 2007, 41:121-145.
• [3]Bijlsma JJ, Groisman EA: Making informed decisions: regulatory interactions between two-component systems. Trends Microbiol 2003, 11(8):359-366.
• [4]Mitrophanov AY, Groisman EA: Signal integration in bacterial two-component regulatory systems. Genes Dev 2008, 22(19):2601-2611.
• [5]Kato A, Groisman EA: The PhoQ/PhoP regulatory network of Salmonella enterica. Adv Exp Med Biol 2008, 631:7-21.
• [6]Kato A, Groisman EA: Connecting two-component regulatory systems by a protein that protects a response regulator from dephosphorylation by its cognate sensor. Genes Dev 2004, 18(18):2302-2313.
• [7]Kox LF, Wosten MM, Groisman EA: A small protein that mediates the activation of a two-component system by another two-component system. EMBO J 2000, 19(8):1861-1872.
• [8]Tu X, Latifi T, Bougdour A, Gottesman S, Groisman EA: The PhoP/PhoQ two-component system stabilizes the alternative sigma factor RpoS in Salmonella enterica. Proc Natl Acad Sci USA 2006, 103(36):13503-13508.
• [9]Bougdour A, Cunning C, Baptiste PJ, Elliott T, Gottesman S: Multiple pathways for regulation of sigmaS (RpoS) stability in Escherichia coli via the action of multiple anti-adaptors. Mol Microbiol 2008, 68(2):298-313.
• [10]Eguchi Y, Itou J, Yamane M, Demizu R, Yamato F, Okada A, Mori H, Kato A, Utsumi R: B1500, a small membrane protein, connects the two-component systems EvgS/EvgA and PhoQ/PhoP in Escherichia coli. Proc Natl Acad Sci USA 2007, 104(47):18712-18717.
• [11]Gerken H, Charlson ES, Cicirelli EM, Kenney LJ, Misra R: MzrA: a novel modulator of the EnvZ/OmpR two-component regulon. Mol Microbiol 2009, 72(6):1408-1422.
• [12]Kato A, Ohnishi H, Yamamoto K, Furuta E, Tanabe H, Utsumi R: Transcription of emrKY is regulated by the EvgA-EvgS two-component system in Escherichia coli K-12. Biosci Biotechnol Biochem 2000, 64(6):1203-1209.
• [13]Cosma CL, Danese PN, Carlson JH, Silhavy TJ, Snyder WB: Mutational activation of the Cpx signal transduction pathway of Escherichia coli suppresses the toxicity conferred by certain envelope-associated stresses. Mol Microbiol 1995, 18(3):491-505.
• [14]Kato A, Tanabe H, Utsumi R: Molecular characterization of the PhoP-PhoQ two-component system in Escherichia coli K-12: identification of extracellular Mg2+-responsive promoters. J Bacteriol 1999, 181(17):5516-5520.
• [15]Lippa AM, Goulian M: Feedback inhibition in the PhoQ/PhoP signaling system by a membrane peptide. PLoS Genet 2009, 5(12):e1000788.
• [16]Kato A, Chen HD, Latify T, Groisman EA: Reciprocal Control Between a Bacterium's Regulatory System and the Modification Status of its Lipopolysaccharide. Mol Cell 2012, 47(6):897-908.
• [17]Vogt SL, Raivio TL: Just scratching the surface: an expanding view of the Cpx envelope stress response. FEMS Microbiol Lett 2012, 326(1):2-11.
• [18]Buelow DR, Raivio TL: Cpx signal transduction is influenced by a conserved N-terminal domain in the novel inhibitor CpxP and the periplasmic protease DegP. J Bacteriol 2005, 187(19):6622-6630.
• [19]DiGiuseppe PA, Silhavy TJ: Signal detection and target gene induction by the CpxRA two-component system. J Bacteriol 2003, 185(8):2432-2440.
• [20]Isaac DD, Pinkner JS, Hultgren SJ, Silhavy TJ: The extracytoplasmic adaptor protein CpxP is degraded with substrate by DegP. Proc Natl Acad Sci USA 2005, 102(49):17775-17779.
• [21]Snyder WB, Davis LJ, Danese PN, Cosma CL, Silhavy TJ: Overproduction of NlpE, a new outer membrane lipoprotein, suppresses the toxicity of periplasmic LacZ by activation of the Cpx signal transduction pathway. J Bacteriol 1995, 177(15):4216-4223.
• [22]Otto K, Silhavy TJ: Surface sensing and adhesion of Escherichia coli controlled by the Cpx-signaling pathway. Proc Natl Acad Sci USA 2002, 99(4):2287-2292.
• [23]Raivio TL, Laird MW, Joly JC, Silhavy TJ: Tethering of CpxP to the inner membrane prevents spheroplast induction of the Cpx envelope stress response. Mol Microbiol 2000, 37(5):1186-1197.
• [24]Yamamoto K, Ishihama A: Characterization of copper-inducible promoters regulated by CpxA/CpxR in Escherichia coli. Biosci Biotechnol Biochem 2006, 70(7):1688-1695.
• [25]McClelland M, Sanderson KE, Spieth J, Clifton SW, Latreille P, Courtney L, Porwollik S, Ali J, Dante M, Du F, et al.: Complete genome sequence of Salmonella enterica serovar Typhimurium LT2. Nature 2001, 413(6858):852-856.
• [26]Raivio TL, Silhavy TJ: The sigmaE and Cpx regulatory pathways:overlapping but distinct envelope stress responses. Curr Opin Microbiol 1999, 2(2):159-165.
• [27]Raffa RG, Raivio TL: A third envelope stress signal transduction pathway in Escherichia coli. Mol Microbiol 2002, 45(6):1599-1611.
• [28]Hagiwara D, Sugiura M, Oshima T, Mori H, Aiba H, Yamashino T, Mizuno T: Genome-wide analyses revealing a signaling network of the RcsC-YojN-RcsB phosphorelay system in Escherichia coli. J Bacteriol 2003, 185(19):5735-5746.
• [29]Lee SJ, Gralla JD: Sigma38 (rpoS) RNA polymerase promoter engagement via -10 region nucleotides. J Biol Chem 2001, 276(32):30064-30071.
• [30]Ramachandran VK, Shearer N, Jacob JJ, Sharma CM, Thompson A: The architecture and ppGpp-dependent expression of the primary transcriptome of Salmonella Typhimurium during invasion gene expression. BMC Genomics 2012, 13:25. BioMed Central Full Text
• [31]Ritz D, Beckwith J: Roles of thiol-redox pathways in bacteria. Annu Rev Microbiol 2001, 55:21-48.
• [32]Slamti L, Waldor MK: Genetic analysis of activation of the Vibrio cholerae Cpx pathway. J Bacteriol 2009, 191(16):5044-5056.
• [33]Stewart EJ, Katzen F, Beckwith J: Six conserved cysteines of the membrane protein DsbD are required for the transfer of electrons from the cytoplasm to the periplasm of Escherichia coli. EMBO J 1999, 18(21):5963-5971.
• [34]Hirano Y, Hossain MM, Takeda K, Tokuda H, Miki K: Structural studies of the Cpx pathway activator NlpE on the outer membrane of Escherichia coli. Structure 2007, 15(8):963-976.
• [35]Tao K, Watanabe S, Narita S, Tokuda H: A periplasmic LolA derivative with a lethal disulfide bond activates the Cpx stress response system. J Bacteriol 2010, 192(21):5657-5662.
• [36]Lippa AM, Goulian M: Perturbation of the oxidizing environment of the periplasm stimulates the PhoQ/PhoP system in Escherichia coli. J Bacteriol 2012, 194(6):1457-1463.
• [37]Kumar JK, Tabor S, Richardson CC: Proteomic analysis of thioredoxin-targeted proteins in Escherichia coli. Proc Natl Acad Sci USA 2004, 101(11):3759-3764.
• [38]Hemm MR, Paul BJ, Miranda-Rios J, Zhang A, Soltanzad N, Storz G: Small stress response proteins in Escherichia coli: proteins missed by classical proteomic studies. J Bacteriol 2010, 192(1):46-58.
• [39]Kato A, Mitrophanov AY, Groisman EA: A connector of two-component regulatory systems promotes signal amplification and persistence of expression. Proc Natl Acad Sci USA 2007, 104(29):12063-12068.
• [40]Davis RW, Bolstein D, Roth JR: Advanced bacterial genetics. Cold Spring Harbor Lab, Cold Spring Harbor, N.Y.; 1980.
• [41]Snavely MD, Gravina SA, Cheung T-BT, Miller CG, Maguire ME: Magnesium transport in Salmonella typhimurium: regulation of mgtA and mgtB expression. J Biol Chem 1991, 266(2):824-829.
• [42]Camp AH, Losick R: A feeding tube model for activation of a cell-specific transcription factor during sporulation in Bacillus subtilis. Genes Dev 2009, 23(8):1014-1024.
• [43]Miller JH: Experiments in molecular genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; 1972.
• [44]Ellermeier CD, Janakiraman A, Slauch JM: Construction of targeted single copy lac fusions using lambda Red and FLP-mediated site-specific recombination in bacteria. Gene 2002, 290(1–2):153-161.
• [45]Datsenko KA, Wanner BL: One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci USA 2000, 97(12):6640-6645.
• [46]Pan W, Ravot E, Tolle R, Frank R, Mosbach R, Turbachova I, Bujard H: Vaccine candidate MSP-1 from Plasmodium falciparum: a redesigned 4917 bp polynucleotide enables synthesis and isolation of full-length protein from Escherichia coli and mammalian cells. Nucleic Acids Res 1999, 27(4):1094-1103.
• [47]Zhou MY, Gomez-Sanchez CE: Universal TA cloning. Curr Issues Mol Biol 2000, 2(1):1-7.
• [48]Fields PI, Groisman EA, Heffron F: A Salmonella locus that controls resistance to microbicidal proteins from phagocytic cells. Science 1989, 243(4894 Pt 1):1059-1062.
• [49]Hanahan D: Studies on transformation of Escherichia coli with plasmids. J Mol Biol 1983, 166(4):557-580.
• [50]Tabor S, Richardson CC: A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci USA 1985, 82:1074-1078.
• [51]Cherepanov PP, Wackernagel W: Gene disruption in Escherichia coli: TcR and KmR cassettes with the option of Flp-catalyzed excision of the antibiotic-resistance determinant. Gene 1995, 158(1):9-14.
• [52]Guzman L-M, Belin D, Carson MJ, Beckwith J: Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol 1995, 177(14):4121-4130.