The NRAMP proteins of <i>Salmonella typhimurium</i> and <i>Escherichia coli</i> are selective manganese transporters involved in the response to reactive oxygen

Molecular Microbiology - Tập 36 Số 5 - Trang 1085-1100 - 2000
David G. Kehres1, Michelle L. Zaharik2, B. Brett Finlay2, Michael E. Maguire1
1Department of Pharmacology, School of Medicine, Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106‐4965, USA
2Biotechnology Laboratory, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada

Tóm tắt

NRAMPs (natural resistance‐associated macrophage proteins) have been characterized in mammals as divalent transition metal transporters involved in iron metabolism and host resistance to certain pathogens. The mechanism of pathogen resistance is proposed to involve sequestration of Fe2+ and Mn2+, cofactors of both prokaryotic and eukaryotic catalases and superoxide dismutases, not only to protect the macrophage against its own generation of reactive oxygen species, but to deny the cations to the pathogen for synthesis of its protective enzymes. NRAMP homologues are also present in bacteria. We report the cloning and characterization of the single NRAMP genes in Escherichia coli and Salmonella enterica ssp. typhimurium, and the cloning of two distinct NRAMP genes from Pseudomonas aeruginosa and an internal fragment of an NRAMP gene in Burkholderia cepacia. The genes are designated mntH because the two enterobacterial NRAMPs encode H+‐stimulated, highly selective manganese(II) transport systems, accounting for all Mn2+ uptake in each species under the conditions tested. For S. typhimurium MntH, the Km for 54Mn2+ (≈ 0.1 µM) was pH independent, but maximal uptake increased as pH decreased. Monovalent cations, osmotic strength, Mg2+ and Ca2+ did not inhibit 54Mn2+ uptake. Ni2+, Cu2+ and Zn2+ inhibited uptake with Kis greater than 100 µM, Co2+ with a Ki of 20 µM and Fe2+ with a Ki that decreased from 100 µM at pH 7.6 to 10 µM at pH 5.5. Fe3+ and Pb2+ inhibited weakly, exhibiting Kis of 50 µM, while Cd2+ was a potent inhibitor with a Ki of about 1 µM. E. coli MntH had a similar inhibition profile, except that Kis were three‐ to 10‐fold higher. Both S. typhimurium and E. coli MntH also transport 55Fe2+ however, the Kms are equivalent to the Kis for Fe2+ inhibition of Mn2+ uptake, and are thus too high to be physiologically relevant. In both S. typhimurium and E. coli, mntH::lacZ constructs were strongly induced by hydrogen peroxide, weakly induced by EDTA and unresponsive to paraquat, consistent with the presence of Fur and OxyR binding sites in the promoters. Strains overexpressing mntH were more susceptible to growth inhibition by Mn2+ and Cd2+ than wild type, and strains lacking a functional mntH gene were more susceptible to killing by hydrogen peroxide. In S. typhimurium strain SL1344, mntH mutants showed no defect in invasion of or survival in cultured HeLa or RAW264.7 macrophage cells; however, expression of mntH::lacZ was induced severalfold by 3 h after invasion of the macrophages. S. typhimurium mntH mutants showed only a slight attenuation of virulence in BALB/c mice. Thus, the NRAMP Mn2+ transporter MntH and Mn2+ play a role in bacterial response to reactive oxygen species and possibly have a role in pathogenesis.

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Tài liệu tham khảo

10.1086/513830

10.1046/j.1365-2958.1998.00790.x

10.1084/jem.190.5.717

10.1021/bi982788s

Ando M., 1979, Superoxide production in pulmonary alveolar macrophages and killing of BCG by the superoxide‐generating system with or without catalase, Infect Immun, 24, 404, 10.1128/iai.24.2.404-410.1979

10.3109/10408418609108735

10.1016/0003-9861(82)90049-2

10.1016/S0014-5793(98)00236-1

10.1046/j.1365-2567.1999.00672.x

Ausubel F.M., 1999, Current Protocols in Molecular Biology.

10.1016/0959-440X(95)80004-2

10.1074/jbc.271.42.26057

Bauer P.D., 1993, Acquisition of manganous ions by mutans group streptococci, J Bacteriol, 175, 819, 10.1128/jb.175.3.819-825.1993

10.1046/j.1365-2958.1999.01360.x

10.1056/NEJM199803053381002

10.1023/A:1005723304911

10.1073/pnas.87.1.389

10.1016/S0020-7519(97)00175-6

10.1016/S0165-2478(98)00127-8

10.1084/jem.180.5.1741

10.1016/0168-9525(96)30042-5

10.1016/0006-2952(73)90196-2

10.1007/BF02784167

10.1021/bi00247a016

10.1128/IAI.43.3.1033-1040.1984

10.1073/pnas.94.25.13997

10.1074/jbc.274.37.26027

10.1128/jb.172.6.3496-3499.1990

10.1128/JB.113.3.1373-1380.1973

10.1111/j.1753-4887.1998.tb01701.x

10.1073/pnas.95.3.1148

10.1006/abbi.1995.1488

10.1046/j.1365-2958.1999.01343.x

10.1007/s000110050330

Gros P., 1983, Cellular mechanisms of genetically controlled host resistance to Mycobacterium bovis (BCG), J Immunol, 131, 1966, 10.4049/jimmunol.131.4.1966

10.1016/S0076-6879(89)73038-X

10.1016/0888-7543(95)80053-O

10.1084/jem.185.4.717

10.1084/jem.189.5.831

10.1038/41343

10.1084/jem.188.2.351

10.1128/jb.171.9.4617-4622.1989

Hao Z., 1999, Cloning, expression and characterization of cadmium and manganese uptake genes from Lactobacillus plantarum, Appl Environ Microbiol, 65, 4746, 10.1128/AEM.65.11.4746-4752.1999

10.1016/S0092-8674(00)81442-7

10.1128/jb.171.9.4742-4751.1989

10.1101/gr.7.7.693

10.1007/s002510050360

10.1139/bcb-75-1-17

10.1089/omi.1.1998.3.151

10.1007/BF02789459

10.1016/S0161-5890(96)00088-0

Kolenbrander P.E., 1998, The adhesion‐associated sca operon in Streptococcus gordonii encodes an inducible high‐affinity ABC transporter for Mn2+ uptake, J Bacteriol, 180, 290, 10.1128/JB.180.2.290-295.1998

10.1016/0378-1119(87)90051-5

Kozlov Y., 1997, Changes in the redox potential and catalase activity of Mn2+ ions during formation of Mn‐bicarbonate complexes, Membr Cell Biol, 11, 115

10.1002/jlb.66.1.113

10.1128/iai.65.2.380-386.1997

10.1073/pnas.88.24.11470

Lissner C.R., 1985, Mouse chromosome 1 Ity locus regulates microbicidal activity of isolated peritoneal macrophages against a diverse group of intracellular and extracellular bacteria, J Immunol, 135, 544, 10.4049/jimmunol.135.1.544

10.1074/jbc.274.8.4863

10.1046/j.1365-2958.2000.01774.x

10.1080/00365529850171044

Miller J.H., 1992, A Short Course in Bacterial Genetics, 1

10.1093/emboj/16.7.1670

10.1093/emboj/18.16.4361

10.1128/jb.171.7.4073-4075.1989

10.1046/j.1365-2958.1998.01016.x

10.1006/jmbi.1998.1609

Perry R.D., 1982, Cadmium and manganese transport in Staphylococcus aureus membrane vesicles, J Bacteriol, 150, 973, 10.1128/jb.150.2.973-976.1982

10.1074/jbc.272.46.28933

10.1128/iai.64.7.2765-2773.1996

10.1002/j.1460-2075.1995.tb07303.x

10.1080/15287398309530433

10.1016/0006-291X(69)90786-4

Silver S., 1970, Manganese active transport in Escherichia coli, J Bacteriol, 104, 1299, 10.1128/jb.104.3.1299-1306.1970

10.1074/jbc.273.44.28663

10.1128/jb.171.9.4761-4766.1989

10.1046/j.1462-5822.1999.00003.x

10.1111/j.1365-2958.1996.tb02497.x

10.1182/blood.V92.6.2157

10.1016/S0092-8674(94)90702-1

10.1128/IAI.63.5.1739-1744.1995

10.1016/0022-2836(92)90934-C

10.1128/IAI.65.1.203-210.1997

10.1016/S1367-5931(99)80031-3

10.1128/IAI.67.4.1974-1981.1999

10.1128/IAI.67.3.1386-1392.1999

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Molecular Microbiology

Tập 36 Số 5

1085-1100

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