Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1995 Apr;177(8):2050–2056. doi: 10.1128/jb.177.8.2050-2056.1995

An Escherichia coli chromosomal ars operon homolog is functional in arsenic detoxification and is conserved in gram-negative bacteria.

C Diorio 1, J Cai 1, J Marmor 1, R Shinder 1, M S DuBow 1
PMCID: PMC176848  PMID: 7721697

Abstract

Arsenic is a known toxic metalloid, whose trivalent and pentavalent ions can inhibit many biochemical processes. Operons which encode arsenic resistance have been found in multicopy plasmids from both gram-positive and gram-negative bacteria. The resistance mechanism is encoded from a single operon which typically consists of an arsenite ion-inducible repressor that regulates expression of an arsenate reductase and inner membrane-associated arsenite export system. Using a lacZ transcriptional gene fusion library, we have identified an Escherichia coli operon whose expression is induced by cellular exposure to sodium arsenite at concentrations as low as 5 micrograms/liter. This chromosomal operon was cloned, sequenced, and found to consist of three cistrons which we named arsR, arsB, and arsC because of their strong homology to plasmid-borne ars operons. Mutants in the chromosomal ars operon were found to be approximately 10- to 100-fold more sensitive to sodium arsenate and arsenite exposure than wild-type E. coli, while wild-type E. coli that contained the operon cloned on a ColE1-based plasmid was found to be at least 2- to 10-fold more resistant to sodium arsenate and arsenite. Moreover, Southern blotting and high-stringency hybridization of this operon with chromosomal DNAs from a number of bacterial species showed homologous sequences among members of the family Enterobacteriaceae, and hybridization was detectable even in Pseudomonas aeruginosa. These results suggest that the chromosomal ars operon may be the evolutionary precursor of the plasmid-borne operon, as a multicopy plasmid location would allow the operon to be amplified and its products to confer increased resistance to this toxic metalloid.

Full Text

The Full Text of this article is available as a PDF (349.3 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Autexier C., DuBow M. S. The Escherichia coli Mu/D108 phage ner homologue gene (nlp) is transcribed and evolutionarily conserved among the Enterobacteriaceae. Gene. 1992 May 1;114(1):13–18. doi: 10.1016/0378-1119(92)90701-p. [DOI] [PubMed] [Google Scholar]
  2. Baba K., Shirai H., Terai A., Kumagai K., Takeda Y., Nishibuchi M. Similarity of the tdh gene-bearing plasmids of Vibrio cholerae non-O1 and Vibrio parahaemolyticus. Microb Pathog. 1991 Jan;10(1):61–70. doi: 10.1016/0882-4010(91)90066-j. [DOI] [PubMed] [Google Scholar]
  3. Bröer S., Ji G., Bröer A., Silver S. Arsenic efflux governed by the arsenic resistance determinant of Staphylococcus aureus plasmid pI258. J Bacteriol. 1993 Jun;175(11):3480–3485. doi: 10.1128/jb.175.11.3480-3485.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carter N. S., Fairlamb A. H. Arsenical-resistant trypanosomes lack an unusual adenosine transporter. Nature. 1993 Jan 14;361(6408):173–176. doi: 10.1038/361173a0. [DOI] [PubMed] [Google Scholar]
  5. Casadaban M. J., Cohen S. N. Lactose genes fused to exogenous promoters in one step using a Mu-lac bacteriophage: in vivo probe for transcriptional control sequences. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4530–4533. doi: 10.1073/pnas.76.9.4530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chen C. M., Misra T. K., Silver S., Rosen B. P. Nucleotide sequence of the structural genes for an anion pump. The plasmid-encoded arsenical resistance operon. J Biol Chem. 1986 Nov 15;261(32):15030–15038. [PubMed] [Google Scholar]
  7. Corbisier P., Ji G., Nuyts G., Mergeay M., Silver S. luxAB gene fusions with the arsenic and cadmium resistance operons of Staphylococcus aureus plasmid pI258. FEMS Microbiol Lett. 1993 Jun 15;110(2):231–238. doi: 10.1111/j.1574-6968.1993.tb06325.x. [DOI] [PubMed] [Google Scholar]
  8. Frey J., Meier R., Gygi D., Nicolet J. Nucleotide sequence of the hemolysin I gene from Actinobacillus pleuropneumoniae. Infect Immun. 1991 Sep;59(9):3026–3032. doi: 10.1128/iai.59.9.3026-3032.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Greer S., Perham R. N. Glutathione reductase from Escherichia coli: cloning and sequence analysis of the gene and relationship to other flavoprotein disulfide oxidoreductases. Biochemistry. 1986 May 6;25(9):2736–2742. doi: 10.1021/bi00357a069. [DOI] [PubMed] [Google Scholar]
  10. Guzzo A., Diorio C., DuBow M. S. Transcription of the Escherichia coli fliC gene is regulated by metal ions. Appl Environ Microbiol. 1991 Aug;57(8):2255–2259. doi: 10.1128/aem.57.8.2255-2259.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Guzzo A., DuBow M. S. A luxAB transcriptional fusion to the cryptic celF gene of Escherichia coli displays increased luminescence in the presence of nickel. Mol Gen Genet. 1994 Feb;242(4):455–460. doi: 10.1007/BF00281796. [DOI] [PubMed] [Google Scholar]
  12. Guzzo A., DuBow M. S. Construction of stable, single-copy luciferase gene fusions in Escherichia coli. Arch Microbiol. 1991;156(6):444–448. doi: 10.1007/BF00245390. [DOI] [PubMed] [Google Scholar]
  13. Huang H., Huang C. F., Wu D. R., Jinn C. M., Jan K. Y. Glutathione as a cellular defence against arsenite toxicity in cultured Chinese hamster ovary cells. Toxicology. 1993 May 24;79(3):195–204. doi: 10.1016/0300-483x(93)90211-a. [DOI] [PubMed] [Google Scholar]
  14. Huckle J. W., Morby A. P., Turner J. S., Robinson N. J. Isolation of a prokaryotic metallothionein locus and analysis of transcriptional control by trace metal ions. Mol Microbiol. 1993 Jan;7(2):177–187. doi: 10.1111/j.1365-2958.1993.tb01109.x. [DOI] [PubMed] [Google Scholar]
  15. Ji G., Silver S. Reduction of arsenate to arsenite by the ArsC protein of the arsenic resistance operon of Staphylococcus aureus plasmid pI258. Proc Natl Acad Sci U S A. 1992 Oct 15;89(20):9474–9478. doi: 10.1073/pnas.89.20.9474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ji G., Silver S. Regulation and expression of the arsenic resistance operon from Staphylococcus aureus plasmid pI258. J Bacteriol. 1992 Jun;174(11):3684–3694. doi: 10.1128/jb.174.11.3684-3694.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kaneko M., Yamaguchi A., Sawai T. Energetics of tetracycline efflux system encoded by Tn10 in Escherichia coli. FEBS Lett. 1985 Dec 2;193(2):194–198. doi: 10.1016/0014-5793(85)80149-6. [DOI] [PubMed] [Google Scholar]
  18. Karkaria C. E., Steiner R. F., Rosen B. P. Ligand interactions in the ArsA protein, the catalytic component of an anion-translocating adenosinetriphosphatase. Biochemistry. 1991 Mar 12;30(10):2625–2628. doi: 10.1021/bi00224a009. [DOI] [PubMed] [Google Scholar]
  19. Kaur P., Rosen B. P. Plasmid-encoded resistance to arsenic and antimony. Plasmid. 1992 Jan;27(1):29–40. doi: 10.1016/0147-619x(92)90004-t. [DOI] [PubMed] [Google Scholar]
  20. Kohara Y., Akiyama K., Isono K. The physical map of the whole E. coli chromosome: application of a new strategy for rapid analysis and sorting of a large genomic library. Cell. 1987 Jul 31;50(3):495–508. doi: 10.1016/0092-8674(87)90503-4. [DOI] [PubMed] [Google Scholar]
  21. Lim C. K., Cooksey D. A. Characterization of chromosomal homologs of the plasmid-borne copper resistance operon of Pseudomonas syringae. J Bacteriol. 1993 Jul;175(14):4492–4498. doi: 10.1128/jb.175.14.4492-4498.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Nordmann P., Naas T. Sequence analysis of PER-1 extended-spectrum beta-lactamase from Pseudomonas aeruginosa and comparison with class A beta-lactamases. Antimicrob Agents Chemother. 1994 Jan;38(1):104–114. doi: 10.1128/aac.38.1.104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ralston D. M., O'Halloran T. V. Ultrasensitivity and heavy-metal selectivity of the allosterically modulated MerR transcription complex. Proc Natl Acad Sci U S A. 1990 May;87(10):3846–3850. doi: 10.1073/pnas.87.10.3846. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Roninson I. B. From amplification to function: the case of the MDR1 gene. Mutat Res. 1992 May;276(3):151–161. doi: 10.1016/0165-1110(92)90005-t. [DOI] [PubMed] [Google Scholar]
  25. Rosen B. P., Weigel U., Karkaria C., Gangola P. Molecular characterization of an anion pump. The arsA gene product is an arsenite(antimonate)-stimulated ATPase. J Biol Chem. 1988 Mar 5;263(7):3067–3070. [PubMed] [Google Scholar]
  26. Rosenstein R., Peschel A., Wieland B., Götz F. Expression and regulation of the antimonite, arsenite, and arsenate resistance operon of Staphylococcus xylosus plasmid pSX267. J Bacteriol. 1992 Jun;174(11):3676–3683. doi: 10.1128/jb.174.11.3676-3683.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. San Francisco M. J., Hope C. L., Owolabi J. B., Tisa L. S., Rosen B. P. Identification of the metalloregulatory element of the plasmid-encoded arsenical resistance operon. Nucleic Acids Res. 1990 Feb 11;18(3):619–624. doi: 10.1093/nar/18.3.619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sanders G. E., Reinert O., Tepe R., Maloney P. Chiropractic adjustive manipulation on subjects with acute low back pain: visual analog pain scores and plasma beta-endorphin levels. J Manipulative Physiol Ther. 1990 Sep;13(7):391–395. [PubMed] [Google Scholar]
  29. Seoane A., García Lobo J. M. Nucleotide sequence of a new class A beta-lactamase gene from the chromosome of Yersinia enterocolitica: implications for the evolution of class A beta-lactamases. Mol Gen Genet. 1991 Aug;228(1-2):215–220. doi: 10.1007/BF00282468. [DOI] [PubMed] [Google Scholar]
  30. Silver S., Budd K., Leahy K. M., Shaw W. V., Hammond D., Novick R. P., Willsky G. R., Malamy M. H., Rosenberg H. Inducible plasmid-determined resistance to arsenate, arsenite, and antimony (III) in escherichia coli and Staphylococcus aureus. J Bacteriol. 1981 Jun;146(3):983–996. doi: 10.1128/jb.146.3.983-996.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Silver S., Ji G., Bröer S., Dey S., Dou D., Rosen B. P. Orphan enzyme or patriarch of a new tribe: the arsenic resistance ATPase of bacterial plasmids. Mol Microbiol. 1993 May;8(4):637–642. doi: 10.1111/j.1365-2958.1993.tb01607.x. [DOI] [PubMed] [Google Scholar]
  32. Silver S., Nucifora G., Chu L., Misra T. K. Bacterial resistance ATPases: primary pumps for exporting toxic cations and anions. Trends Biochem Sci. 1989 Feb;14(2):76–80. doi: 10.1016/0968-0004(89)90048-0. [DOI] [PubMed] [Google Scholar]
  33. Silver S., Nucifora G., Phung L. T. Human Menkes X-chromosome disease and the staphylococcal cadmium-resistance ATPase: a remarkable similarity in protein sequences. Mol Microbiol. 1993 Oct;10(1):7–12. doi: 10.1111/j.1365-2958.1993.tb00898.x. [DOI] [PubMed] [Google Scholar]
  34. Silver S., Walderhaug M. Gene regulation of plasmid- and chromosome-determined inorganic ion transport in bacteria. Microbiol Rev. 1992 Mar;56(1):195–228. doi: 10.1128/mr.56.1.195-228.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Smith A. H., Hopenhayn-Rich C., Bates M. N., Goeden H. M., Hertz-Picciotto I., Duggan H. M., Wood R., Kosnett M. J., Smith M. T. Cancer risks from arsenic in drinking water. Environ Health Perspect. 1992 Jul;97:259–267. doi: 10.1289/ehp.9297259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sofia H. J., Burland V., Daniels D. L., Plunkett G., 3rd, Blattner F. R. Analysis of the Escherichia coli genome. V. DNA sequence of the region from 76.0 to 81.5 minutes. Nucleic Acids Res. 1994 Jul 11;22(13):2576–2586. doi: 10.1093/nar/22.13.2576. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sol K., Lapointe M., MacLeod M., Nadeau C., DuBow M. S. A cloned fragment of HeLa DNA containing consensus sequences of satellite II and III DNA hybridizes with the Drosophila P-element and with the 1.8 kb family of human KpnI fragments. Biochim Biophys Acta. 1986 Nov 13;868(2-3):128–135. doi: 10.1016/0167-4781(86)90015-1. [DOI] [PubMed] [Google Scholar]
  38. Tisa L. S., Rosen B. P. Molecular characterization of an anion pump. The ArsB protein is the membrane anchor for the ArsA protein. J Biol Chem. 1990 Jan 5;265(1):190–194. [PubMed] [Google Scholar]
  39. Tolias P. P., DuBow M. S. The cloning and characterization of the bacteriophage D108 regulatory DNA-binding protein ner. EMBO J. 1985 Nov;4(11):3031–3037. doi: 10.1002/j.1460-2075.1985.tb04040.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Vieira J., Messing J. Production of single-stranded plasmid DNA. Methods Enzymol. 1987;153:3–11. doi: 10.1016/0076-6879(87)53044-0. [DOI] [PubMed] [Google Scholar]
  41. Wu J., Rosen B. P. Metalloregulated expression of the ars operon. J Biol Chem. 1993 Jan 5;268(1):52–58. [PubMed] [Google Scholar]
  42. Wu J., Rosen B. P. The ArsR protein is a trans-acting regulatory protein. Mol Microbiol. 1991 Jun;5(6):1331–1336. doi: 10.1111/j.1365-2958.1991.tb00779.x. [DOI] [PubMed] [Google Scholar]
  43. Wu J., Rosen B. P. The arsD gene encodes a second trans-acting regulatory protein of the plasmid-encoded arsenical resistance operon. Mol Microbiol. 1993 May;8(3):615–623. doi: 10.1111/j.1365-2958.1993.tb01605.x. [DOI] [PubMed] [Google Scholar]
  44. Wu J., Tisa L. S., Rosen B. P. Membrane topology of the ArsB protein, the membrane subunit of an anion-translocating ATPase. J Biol Chem. 1992 Jun 25;267(18):12570–12576. [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES