BMC Microbiology | |
Two RND proteins involved in heavy metal efflux in Caulobacter crescentus belong to separate clusters within proteobacteria | |
Marilis V Marques1  Cristiane Guzzo1  Vânia S Braz1  Estela Y Valencia1  | |
[1] Departamento de Microbiologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, Av. Prof. Lineu Prestes 1374, São Paulo, SP, 05508-900, Brazil | |
关键词: Gene expression; Caulobacter crescentus; RND systems; Heavy metal efflux; | |
Others : 1143953 DOI : 10.1186/1471-2180-13-79 |
|
received in 2012-10-16, accepted in 2013-03-21, 发布年份 2013 | |
【 摘 要 】
Background
Heavy metal Resistance-Nodulation-Division (HME-RND) efflux systems help Gram-negative bacteria to keep the intracellular homeostasis under high metal concentrations. These proteins constitute the cytoplasmic membrane channel of the tripartite RND transport systems. Caulobacter crescentus NA1000 possess two HME-RND proteins, and the aim of this work was to determine their involvement in the response to cadmium, zinc, cobalt and nickel, and to analyze the phylogenetic distribution and characteristic signatures of orthologs of these two proteins.
Results
Expression assays of the czrCBA operon showed significant induction in the presence of cadmium and zinc, and moderate induction by cobalt and nickel. The nczCBA operon is highly induced in the presence of nickel and cobalt, moderately induced by zinc and not induced by cadmium. Analysis of the resistance phenotype of mutant strains showed that the ΔczrA strain is highly sensitive to cadmium, zinc and cobalt, but resistant to nickel. The ΔnczA strain and the double mutant strain showed reduced growth in the presence of all metals tested. Phylogenetic analysis of the C. crescentus HME-RND proteins showed that CzrA-like proteins, in contrast to those similar to NczA, are almost exclusively found in the Alphaproteobacteria group, and the characteristic protein signatures of each group were highlighted.
Conclusions
The czrCBA efflux system is involved mainly in response to cadmium and zinc with a secondary role in response to cobalt. The nczCBA efflux system is involved mainly in response to nickel and cobalt, with a secondary role in response to cadmium and zinc. CzrA belongs to the HME2 subfamily, which is almost exclusively found in the Alphaproteobacteria group, as shown by phylogenetic analysis. NczA belongs to the HME1 subfamily which is more widespread among diverse Proteobacteria groups. Each of these subfamilies present distinctive amino acid signatures.
【 授权许可】
2013 Valencia et al.; licensee BioMed Central Ltd.
【 预 览 】
Files | Size | Format | View |
---|---|---|---|
20150330045813461.pdf | 2032KB | download | |
Figure 6. | 182KB | Image | download |
Figure 5. | 117KB | Image | download |
Figure 4. | 44KB | Image | download |
Figure 3. | 58KB | Image | download |
Figure 2. | 20KB | Image | download |
Figure 1. | 16KB | Image | download |
【 图 表 】
Figure 1.
Figure 2.
Figure 3.
Figure 4.
Figure 5.
Figure 6.
【 参考文献 】
- [1]Valls M, de Lorenzo V: Exploiting the genetic and biochemical capacities of bacteria for the remediation of heavy metal pollution. FEMS Microbiol Rev 2002, 26:327-338.
- [2]Mitra RS, Bernstein IA: Single-strand breakage in DNA of Escherichia coli exposed to Cd2+. J Bacteriol 1978, 133:75-80.
- [3]Bruins MR, Kapil S, Oehme FW: Microbial resistance to metals in the environment. Ecotoxicol Environ Saf 2000, 45:198-207.
- [4]Nzengue Y, Candeias SM, Sauvaigo S, Douki T, Favier A, Rachidi W, Guiraud P: The toxicity redox mechanisms of cadmium alone or together with copper and zinc homeostasis alteration: its redox biomarkers. J Trace Elem Med Biol 2011, 25:171-180.
- [5]Ma Z, Jacobsen FE, Giedroc DP: Metal Transporters and Metal Sensors: How Coordination Chemistry Controls Bacterial Metal Homeostasis. Chem Rev 2009, 109:4644-4681.
- [6]Monchy S, Benotmane MA, Janssen P, Vallaeys T, Taghavi S, van der Lelie D, Mergeay M: Plasmids pMOL28 and pMOL30 of Cupriavidus metallidurans are specialized in the maximal viable response to heavy metals. J Bacteriol 2007, 189:7417-7425.
- [7]Haritha A, Sagar KP, Tiwari A, Kiranmayi P, Rodrigue A, Mohan PM, Singh SS: MrdH, a novel metal resistance determinant of Pseudomonas putida KT2440, is flanked by metal-inducible mobile genetic elements. J Bacteriol 2009, 191:5976-5987.
- [8]von Rozycki T, Nies DH: Cupriavidus metallidurans: evolution of a metal-resistant bacterium. Antonie Van Leeuwenhoek 2009, 96:115-139.
- [9]Xiong J, Li D, Li H, He M, Miller SJ, Yu L, Rensing C, Wang G: Genome analysis and characterization of zinc efflux systems of a highly zinc-resistant bacterium, Comamonas testosteroni S44. Res Microbiol 2011, 162:671-679.
- [10]Saier MH Jr: A Functional-Phylogenetic System for the Classification of Transport Proteins. J Cell Biochem Suppl 1999, 32/33:84-94.
- [11]Silver S, Phung T: A bacterial view of the periodic table: genes and proteins for toxic inorganic ions. J Ind Microbiol Biotechnol 2005, 32:587-605.
- [12]Chan H, Babayan V, Blyumin E, Gandhi C, Hak K, Harake D, Kumar K, Lee P, Li TT, Liu HY, et al.: The P-type ATPase superfamily. J Mol Microbiol Biotechnol 2010, 19:5-104.
- [13]Arguello JM, Gonzalez-Guerrero M, Raimunda D: Bacterial transition metal P(1B)-ATPases: transport mechanism and roles in virulence. Biochemistry 2011, 50:9940-9949.
- [14]Nies DH: Efflux-mediated heavy metal resistance in prokaryotes. FEMS Microbiol Rev 2003, 27:313-339.
- [15]Higuchi T, Hattori M, Tanaka Y, Ishitani R, Nureki O: Crystal structure of the cytosolic domain of the cation diffusion facilitator family protein. Ptoteins 2009, 76:768-771.
- [16]Saier MH Jr, Tam R, Reizer A, Reizer J: Two novel families of bacterial membrane proteins concerned with nodulation, cell division and transport. Mol Microbiol 1994, 11:841-847.
- [17]Tseng TT, Gratwick KS, Kollman J, Park D, Nies DH, Goffeau A, Saier MH Jr: The RND permease superfamily: an ancient, ubiquitous and diverse family that includes human disease and development proteins. J Mol Microbiol Biotechnol 1999, 1:107-125.
- [18]Murakami S, Nakashima R, Yamashita E, Yamaguchi A: Crystal structure of bacterial multidrug efflux transporter AcrB. Nature 2002, 419:587-593.
- [19]Koronakis V, Sharff A, Koronakis E, Luisi B, Hughes C: Crystal structure of the bacterial membrane protein TolC central to multidrug efflux and protein export. Nature 2000, 405:914-919.
- [20]Akama H, Kanemaki M, Yoshimura M, Tsukihara T, Kashiwagi T, Yoneyama H, Narita S, Nakagawa A, Nakae T: Crystal structure of the drug discharge outer membrane protein, OprM, of Pseudomonas aeruginosa: dual modes of membrane anchoring and occluded cavity end. J Biol Chem 2004, 279:52816-52819.
- [21]Akama H, Matsuura T, Kashiwagi S, Yoneyama H, Narita S, Tsukihara T, Nakagawa A, Nakae T: Crystal structure of the membrane fusion protein, MexA, of the multidrug transporter in Pseudomonas aeruginosa. J Biol Chem 2004, 279:25939-25942.
- [22]Saier MH Jr: A functional-phylogenetic classification system for transmembrane solute transporters. Microbiol Mol Biol Rev 2000, 64:354-411.
- [23]Goldberg M, Pribyl T, Juhnke S, Nies DH: Energetics and topology of CzcA, a cation/proton antiporter of the resistance-nodulation-cell division protein family. J Biol Chem 1999, 274:26065-26070.
- [24]Franke S, Grass G, Rensing C, Nies DH: Molecular analysis of the copper-transporting efflux system CusCFBA of Escherichia coli. J Bacteriol 2003, 185:3804-3812.
- [25]Long F, Su CC, Zimmermann MT, Boyken SE, Rajashankar KR, Jernigan RL, Yu EW: Crystal structures of the CusA efflux pump suggest methionine-mediated metal transport. Nature 2010, 467:484-488.
- [26]Nies DH: The cobalt, zinc, and cadmium efflux system CzcABC from Alcaligenes eutrophus functions as a cation-proton antiporter in Escherichia coli. J Bacteriol 1995, 177:2707-2712.
- [27]Grosse C, Grass G, Anton A, Franke S, Santos AN, Lawley B, Brown NL, Nies DH: Transcriptional organization of the czc heavy-metal homeostasis determinant from Alcaligenes eutrophus. J Bacteriol 1999, 181:2385-2393.
- [28]Legatzki A, Franke S, Lucke S, Hoffmann T, Anton A, Neumann D, Nies DH: First step towards a quantitative model describing Czc-mediated heavy metal resistance in Ralstonia metallidurans. Biodegradation 2003, 14:153-168.
- [29]Tibazarwa C, Wuertz S, Mergeay M, Wyns L, van Der Lelie D: Regulation of the cnr cobalt and nickel resistance determinant of Ralstonia eutropha (Alcaligenes eutrophus) CH34. J Bacteriol 2000, 182:1399-1409.
- [30]Grass G, Grosse C, Nies DH: Regulation of the cnr cobalt and nickel resistance determinant from Ralstonia sp. strain CH34. J Bacteriol 2000, 182:1390-1398.
- [31]Schmidt T, Schlegel HG: Combined nickel-cobalt-cadmium resistance encoded by the ncc locus of Alcaligenes xylosoxidans 31A. J Bacteriol 1994, 176:7045-7054.
- [32]Hassan MT, van der Lelie D, Springael D, Romling U, Ahmed N, Mergeay M: Identification of a gene cluster, czr, involved in cadmium and zinc resistance in Pseudomonas aeruginosa. Gene 1999, 238:417-425.
- [33]Stahler FN, Odenbreit S, Haas R, Wilrich J, Van Vliet AH, Kusters JG, Kist M, Bereswill S: The novel Helicobacter pylori CznABC metal efflux pump is required for cadmium, zinc, and nickel resistance, urease modulation, and gastric colonization. Infect Immun 2006, 74:3845-3852.
- [34]Braz VS, Marques MV: Genes involved in cadmium resistance in Caulobacter crescentus. FEMS Microbiol Lett 2005, 251:289-295.
- [35]Hu P, Brodie EL, Suzuki Y, McAdams HH, Andersen GL: Whole-genome transcriptional analysis of heavy metal stresses in Caulobacter crescentus. J Bacteriol 2005, 187:8437-8449.
- [36]Grosse C, Anton A, Hoffmann T, Franke S, Schleuder G, Nies DH: Identification of a regulatory pathway that controls the heavy-metal resistance system Czc via promoter czcNp in Ralstonia metallidurans. Arch Microbiol 2004, 182:109-118.
- [37]McGrath PT, Lee H, Zhang L, Iniesta AA, Hottes AK, Tan MH, Hillson NJ, Hu P, Shapiro L, McAdams HH: High-throughput identification of transcription start sites, conserved promoter motifs and predicted regulons. Nat Biotechnol 2007, 25:584-592.
- [38]Miller JH: Experiments in Molecular Genetics. New York: Cold Spring Harbor, Laboratory Press; 1972. [1]
- [39]Nierman WC, Feldblyum TV, Laub MT, Paulsen IT, Nelson KE, Eisen JA, Heidelberg JF, Alley MR, Ohta N, Maddock JR: Complete genome sequence of Caulobacter crescentus. Proc Natl Acad Sci USA 2001, 98:4136-4141.
- [40]Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, et al.: Clustal W and Clustal X version 2.0. Bioinformatics 2007, 23:2947-2948.
- [41]Liesegang H, Lemke K, Siddiqui RA, Schlegel HG: Characterization of the inducible nickel and cobalt resistance determinant cnr from pMOL28 of Alcaligenes eutrophus CH34. J Bacteriol 1993, 175:767-778.
- [42]Crooks GE, Hon G, Chandonia JM, Brenner SE: WebLogo: A sequence logo generator. Genome Res 2004, 14:1188-1190.
- [43]The PyMOL Molecular Graphics System. Version 1.5.0.4 Schrödinger, LLC.
- [44]Kelley LA, Sternberg MJE: Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 2009, 4:363-371.
- [45]Su CC, Long F, Zimmermann MT, Rajashankar KR, Jernigan RL, Yu EW: Crystal structure of the CusBA heavy-metal efflux complex of Escherichia coli. Nature 2011, 470:558-562.
- [46]Ely B: Genetics of Caulobacter crescentus. Methods Enzymol 1991, 204:372-384.
- [47]Hanahan D: Studies on transformation of Escherichia coli with plasmids. J Mol Biol 1983, 166:557-580.
- [48]Simon R, Prieffer U, Puhler A: A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in Gram-negative bacteria. Nat Biotechnol 1983, 1:784-791.
- [49]Evinger M, Agabian N: Envelope-associated nucleoid from Caulobacter crescentus stalked and swarmer cells. J Bacteriol 1977, 132:294-301.
- [50]Gober JW, Shapiro L: A developmentally regulated Caulobacter flagellar promoter is activated by 3′ enhancer and IHF binding elements. Mol Biol Cell 1992, 3:913-926.
- [51]Jenal U, Fuchs T: An essential protease involved in bacterial cell-cycle control. EMBO J 1998, 17:5658-5669.