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J Bacteriol. Jul 1996; 178(14): 3996–4003.
PMCID: PMC178152

Ferric uptake regulator (Fur) mutants of Pseudomonas aeruginosa demonstrate defective siderophore-mediated iron uptake, altered aerobic growth, and decreased superoxide dismutase and catalase activities.

Abstract

Pseudomonas aeruginosa is considered a strict aerobe that possesses several enzymes important in the disposal of toxic oxygen reduction products including iron- and manganese-cofactored superoxide dismutase and catalase. At present, the nature of the regulation of these enzymes in P. aeruginosa Is not understood. To address these issues, we used two mutants called A4 and C6 which express altered Fur (named for ferric uptake regulation) proteins and constitutively produce the siderophores pyochelin and pyoverdin. Both mutants required a significant lag phase prior to log-phase aerobic growth, but this lag was not as apparent when the organisms were grown under microaerobic conditions. The addition of iron salts to mutant A4 and, to a greater extent, C6 cultures allowed for an increased growth rate under both conditions relative to that of bacteria without added iron. Increased manganese superoxide dismutase (Mn-SOD) and decreased catalase activities were also apparent in the mutants, although the second catalase, KatB, was detected in cell extracts of each fur mutant. Iron deprivation by the addition of the iron chelator 2,2'-dipyridyl to wild-type bacteria produced an increase in Mn-SOD activity and a decrease in total catalase activity, similar to the fur mutant phenotype. Purified wild-type Fur bound more avidly than mutant Fur to a PCR product containing two palindromic 19-bp "iron box" regions controlling expression of an operon containing the sodA gene that encodes Mn-SOD. All mutants were defective in both ferripyochelin- and ferripyoverdin-mediated iron uptake. Two mutants of strain PAO1, defective in pyoverdin but not pyochelin biosynthesis, produced increased Mn-SOD activity. Sensitivity to both the redox-cycling agent paraquat and hydrogen peroxide was greater in each mutant than in the wild-type strain. In summary, the results indicate that mutations in the P. aeruginosa fur locus affect aerobic growth and SOD and catalase activities in P. aeruginosa. We postulate that reduced siderophore-mediated iron uptake, especially that by pyoverdin, may be one possible mechanism contributing to such effect.

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Selected References

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  • Ankenbauer R, Sriyosachati S, Cox CD. Effects of siderophores on the growth of Pseudomonas aeruginosa in human serum and transferrin. Infect Immun. 1985 Jul;49(1):132–140. [PMC free article] [PubMed]
  • Ankenbauer RG, Quan HN. FptA, the Fe(III)-pyochelin receptor of Pseudomonas aeruginosa: a phenolate siderophore receptor homologous to hydroxamate siderophore receptors. J Bacteriol. 1994 Jan;176(2):307–319. [PMC free article] [PubMed]
  • Ankenbauer RG, Staley AL, Rinehart KL, Cox CD. Mutasynthesis of siderophore analogues by Pseudomonas aeruginosa. Proc Natl Acad Sci U S A. 1991 Mar 1;88(5):1878–1882. [PMC free article] [PubMed]
  • Bagg A, Neilands JB. Ferric uptake regulation protein acts as a repressor, employing iron (II) as a cofactor to bind the operator of an iron transport operon in Escherichia coli. Biochemistry. 1987 Aug 25;26(17):5471–5477. [PubMed]
  • BEERS RF, Jr, SIZER IW. A spectrophotometric method for measuring the breakdown of hydrogen peroxide by catalase. J Biol Chem. 1952 Mar;195(1):133–140. [PubMed]
  • Berish SA, Subbarao S, Chen CY, Trees DL, Morse SA. Identification and cloning of a fur homolog from Neisseria gonorrhoeae. Infect Immun. 1993 Nov;61(11):4599–4606. [PMC free article] [PubMed]
  • Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. [PubMed]
  • Brown SM, Howell ML, Vasil ML, Anderson AJ, Hassett DJ. Cloning and characterization of the katB gene of Pseudomonas aeruginosa encoding a hydrogen peroxide-inducible catalase: purification of KatB, cellular localization, and demonstration that it is essential for optimal resistance to hydrogen peroxide. J Bacteriol. 1995 Nov;177(22):6536–6544. [PMC free article] [PubMed]
  • Calderwood SB, Mekalanos JJ. Iron regulation of Shiga-like toxin expression in Escherichia coli is mediated by the fur locus. J Bacteriol. 1987 Oct;169(10):4759–4764. [PMC free article] [PubMed]
  • Carlioz A, Touati D. Isolation of superoxide dismutase mutants in Escherichia coli: is superoxide dismutase necessary for aerobic life? EMBO J. 1986 Mar;5(3):623–630. [PMC free article] [PubMed]
  • Clare DA, Duong MN, Darr D, Archibald F, Fridovich I. Effects of molecular oxygen on detection of superoxide radical with nitroblue tetrazolium and on activity stains for catalase. Anal Biochem. 1984 Aug 1;140(2):532–537. [PubMed]
  • Compan I, Touati D. Interaction of six global transcription regulators in expression of manganese superoxide dismutase in Escherichia coli K-12. J Bacteriol. 1993 Mar;175(6):1687–1696. [PMC free article] [PubMed]
  • Cornelis P, Hohnadel D, Meyer JM. Evidence for different pyoverdine-mediated iron uptake systems among Pseudomonas aeruginosa strains. Infect Immun. 1989 Nov;57(11):3491–3497. [PMC free article] [PubMed]
  • Cox CD, Graham R. Isolation of an iron-binding compound from Pseudomonas aeruginosa. J Bacteriol. 1979 Jan;137(1):357–364. [PMC free article] [PubMed]
  • Cox CD. Iron uptake with ferripyochelin and ferric citrate by Pseudomonas aeruginosa. J Bacteriol. 1980 May;142(2):581–587. [PMC free article] [PubMed]
  • Cox CD, Adams P. Siderophore activity of pyoverdin for Pseudomonas aeruginosa. Infect Immun. 1985 Apr;48(1):130–138. [PMC free article] [PubMed]
  • Farr SB, Kogoma T. Oxidative stress responses in Escherichia coli and Salmonella typhimurium. Microbiol Rev. 1991 Dec;55(4):561–585. [PMC free article] [PubMed]
  • Foster JW, Hall HK. Effect of Salmonella typhimurium ferric uptake regulator (fur) mutations on iron- and pH-regulated protein synthesis. J Bacteriol. 1992 Jul;174(13):4317–4323. [PMC free article] [PubMed]
  • Gardner PR, Fridovich I. Superoxide sensitivity of the Escherichia coli 6-phosphogluconate dehydratase. J Biol Chem. 1991 Jan 25;266(3):1478–1483. [PubMed]
  • Gardner PR, Fridovich I. Superoxide sensitivity of the Escherichia coli aconitase. J Biol Chem. 1991 Oct 15;266(29):19328–19333. [PubMed]
  • Hantke K. Selection procedure for deregulated iron transport mutants (fur) in Escherichia coli K 12: fur not only affects iron metabolism. Mol Gen Genet. 1987 Nov;210(1):135–139. [PubMed]
  • Hassett DJ, Charniga L, Bean K, Ohman DE, Cohen MS. Response of Pseudomonas aeruginosa to pyocyanin: mechanisms of resistance, antioxidant defenses, and demonstration of a manganese-cofactored superoxide dismutase. Infect Immun. 1992 Feb;60(2):328–336. [PMC free article] [PubMed]
  • Hassett DJ, Cohen MS. Bacterial adaptation to oxidative stress: implications for pathogenesis and interaction with phagocytic cells. FASEB J. 1989 Dec;3(14):2574–2582. [PubMed]
  • Hassett DJ, Schweizer HP, Ohman DE. Pseudomonas aeruginosa sodA and sodB mutants defective in manganese- and iron-cofactored superoxide dismutase activity demonstrate the importance of the iron-cofactored form in aerobic metabolism. J Bacteriol. 1995 Nov;177(22):6330–6337. [PMC free article] [PubMed]
  • Hassett DJ, Woodruff WA, Wozniak DJ, Vasil ML, Cohen MS, Ohman DE. Cloning and characterization of the Pseudomonas aeruginosa sodA and sodB genes encoding manganese- and iron-cofactored superoxide dismutase: demonstration of increased manganese superoxide dismutase activity in alginate-producing bacteria. J Bacteriol. 1993 Dec;175(23):7658–7665. [PMC free article] [PubMed]
  • Hickey EK, Cianciotto NP. Cloning and sequencing of the Legionella pneumophila fur gene. Gene. 1994 May 27;143(1):117–121. [PubMed]
  • Kammler M, Schön C, Hantke K. Characterization of the ferrous iron uptake system of Escherichia coli. J Bacteriol. 1993 Oct;175(19):6212–6219. [PMC free article] [PubMed]
  • Kono Y, Fridovich I. Superoxide radical inhibits catalase. J Biol Chem. 1982 May 25;257(10):5751–5754. [PubMed]
  • Kuo CF, Mashino T, Fridovich I. alpha, beta-Dihydroxyisovalerate dehydratase. A superoxide-sensitive enzyme. J Biol Chem. 1987 Apr 5;262(10):4724–4727. [PubMed]
  • Lam MS, Litwin CM, Carroll PA, Calderwood SB. Vibrio cholerae fur mutations associated with loss of repressor activity: implications for the structural-functional relationships of fur. J Bacteriol. 1994 Aug;176(16):5108–5115. [PMC free article] [PubMed]
  • Leoni L, Ciervo A, Orsi N, Visca P. Iron-regulated transcription of the pvdA gene in Pseudomonas aeruginosa: effect of Fur and PvdS on promoter activity. J Bacteriol. 1996 Apr;178(8):2299–2313. [PMC free article] [PubMed]
  • Litwin CM, Boyko SA, Calderwood SB. Cloning, sequencing, and transcriptional regulation of the Vibrio cholerae fur gene. J Bacteriol. 1992 Mar;174(6):1897–1903. [PMC free article] [PubMed]
  • Loewen PC. Isolation of catalase-deficient Escherichia coli mutants and genetic mapping of katE, a locus that affects catalase activity. J Bacteriol. 1984 Feb;157(2):622–626. [PMC free article] [PubMed]
  • Marklund S, Marklund G. Involvement of the superoxide anion radical in the autoxidation of pyrogallol and a convenient assay for superoxide dismutase. Eur J Biochem. 1974 Sep 16;47(3):469–474. [PubMed]
  • Meyer JM, Hohnadel D, Khan A, Cornelis P. Pyoverdin-facilitated iron uptake in Pseudomonas aeruginosa: immunological characterization of the ferripyoverdin receptor. Mol Microbiol. 1990 Aug;4(8):1401–1405. [PubMed]
  • Moody CS, Hassan HM. Anaerobic biosynthesis of the manganese-containing superoxide dismutase in Escherichia coli. J Biol Chem. 1984 Oct 25;259(20):12821–12825. [PubMed]
  • Niederhoffer EC, Naranjo CM, Bradley KL, Fee JA. Control of Escherichia coli superoxide dismutase (sodA and sodB) genes by the ferric uptake regulation (fur) locus. J Bacteriol. 1990 Apr;172(4):1930–1938. [PMC free article] [PubMed]
  • Ochsner UA, Vasil AI, Vasil ML. Role of the ferric uptake regulator of Pseudomonas aeruginosa in the regulation of siderophores and exotoxin A expression: purification and activity on iron-regulated promoters. J Bacteriol. 1995 Dec;177(24):7194–7201. [PMC free article] [PubMed]
  • Ochsner UA, Vasil ML. Gene repression by the ferric uptake regulator in Pseudomonas aeruginosa: cycle selection of iron-regulated genes. Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4409–4414. [PMC free article] [PubMed]
  • Prince RW, Cox CD, Vasil ML. Coordinate regulation of siderophore and exotoxin A production: molecular cloning and sequencing of the Pseudomonas aeruginosa fur gene. J Bacteriol. 1993 May;175(9):2589–2598. [PMC free article] [PubMed]
  • Prince RW, Storey DG, Vasil AI, Vasil ML. Regulation of toxA and regA by the Escherichia coli fur gene and identification of a Fur homologue in Pseudomonas aeruginosa PA103 and PA01. Mol Microbiol. 1991 Nov;5(11):2823–2831. [PubMed]
  • Schmitt MP, Holmes RK. Iron-dependent regulation of diphtheria toxin and siderophore expression by the cloned Corynebacterium diphtheriae repressor gene dtxR in C. diphtheriae C7 strains. Infect Immun. 1991 Jun;59(6):1899–1904. [PMC free article] [PubMed]
  • Schwyn B, Neilands JB. Universal chemical assay for the detection and determination of siderophores. Anal Biochem. 1987 Jan;160(1):47–56. [PubMed]
  • Silver S, Johnseine P, Whitney E, Clark D. Manganese-resistant mutants of Escherichia coli: physiological and genetic studies. J Bacteriol. 1972 Apr;110(1):186–195. [PMC free article] [PubMed]
  • Sokol PA. Production and utilization of pyochelin by clinical isolates of Pseudomonas cepacia. J Clin Microbiol. 1986 Mar;23(3):560–562. [PMC free article] [PubMed]
  • Staggs TM, Perry RD. Identification and cloning of a fur regulatory gene in Yersinia pestis. J Bacteriol. 1991 Jan;173(2):417–425. [PMC free article] [PubMed]
  • Tardat B, Touati D. Two global regulators repress the anaerobic expression of MnSOD in Escherichia coli::Fur (ferric uptake regulation) and Arc (aerobic respiration control). Mol Microbiol. 1991 Feb;5(2):455–465. [PubMed]
  • Tardat B, Touati D. Iron and oxygen regulation of Escherichia coli MnSOD expression: competition between the global regulators Fur and ArcA for binding to DNA. Mol Microbiol. 1993 Jul;9(1):53–63. [PubMed]
  • Thomas CE, Sparling PF. Identification and cloning of a fur homologue from Neisseria meningitidis. Mol Microbiol. 1994 Feb;11(4):725–737. [PubMed]
  • Tolmasky ME, Wertheimer AM, Actis LA, Crosa JH. Characterization of the Vibrio anguillarum fur gene: role in regulation of expression of the FatA outer membrane protein and catechols. J Bacteriol. 1994 Jan;176(1):213–220. [PMC free article] [PubMed]
  • Touati D, Jacques M, Tardat B, Bouchard L, Despied S. Lethal oxidative damage and mutagenesis are generated by iron in delta fur mutants of Escherichia coli: protective role of superoxide dismutase. J Bacteriol. 1995 May;177(9):2305–2314. [PMC free article] [PubMed]
  • Vasil ML, Chamberlain C, Grant CC. Molecular studies of Pseudomonas exotoxin A gene. Infect Immun. 1986 May;52(2):538–548. [PMC free article] [PubMed]
  • Wayne LG, Diaz GA. A double staining method for differentiating between two classes of mycobacterial catalase in polyacrylamide electrophoresis gels. Anal Biochem. 1986 Aug 15;157(1):89–92. [PubMed]
  • Woodbury W, Spencer AK, Stahman MA. An improved procedure using ferricyanide for detecting catalase isozymes. Anal Biochem. 1971 Nov;44(1):301–305. [PubMed]
  • Wooldridge KG, Williams PH, Ketley JM. Iron-responsive genetic regulation in Campylobacter jejuni: cloning and characterization of a fur homolog. J Bacteriol. 1994 Sep;176(18):5852–5856. [PMC free article] [PubMed]
  • Ye RW, Haas D, Ka JO, Krishnapillai V, Zimmermann A, Baird C, Tiedje JM. Anaerobic activation of the entire denitrification pathway in Pseudomonas aeruginosa requires Anr, an analog of Fnr. J Bacteriol. 1995 Jun;177(12):3606–3609. [PMC free article] [PubMed]

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