title: Sub-inhibitory fosmidomycin exposures elicits oxidative stress in Salmonella enterica serovar Typhimurium LT2.
abstract: Fosmidomycin is a time-dependent nanomolar inhibitor of methylerythritol phosphate (MEP) synthase, which is the enzyme that catalyzes the first committed step in the MEP pathway to isoprenoids. Importantly, fosmidomycin is one of only a few MEP pathway-specific inhibitors that exhibits antimicrobial activity. Most inhibitors identified to date only exhibit activity against isolated pathway enzymes. The MEP pathway is the sole route to isoprenoids in many bacteria, yet has no human homologs. The development of inhibitors of this pathway holds promise as novel antimicrobial agents. Similarly, analyses of the bacterial response toward MEP pathway inhibitors provides valuable information toward the understanding of how emergent resistance may ultimately develop to this class of antibiotics. We have examined the transcriptional response of Salmonella enterica serovar typhimurium LT2 to sub-inhibitory concentrations of fosmidomycin via cDNA microarray and RT-PCR. Within the regulated genes identified by microarray were a number of genes encoding enzymes associated with the mediation of reactive oxygen species (ROS). Regulation of a panel of genes implicated in the response of cells to oxidative stress (including genes for catalases, superoxide dismutases, and alkylhydrogen peroxide reductases) was investigated and mild upregulation in some members was observed as a function of fosmidomycin exposure over time. The extent of regulation of these genes was similar to that observed for comparable exposures to kanamycin, but differed significantly from tetracycline. Furthermore, S. typhimurium exposed to sub-inhibitory concentrations of fosmidomycin displayed an increased sensitivity to exogenous H2O2 relative to either untreated controls or kanamycin-treated cells. Our results suggest that endogenous oxidative stress is one consequence of exposures to fosmidomycin, likely through the temporal depletion of intracellular isoprenoids themselves, rather than other mechanisms that have been proposed to facilitate ROS accumulation in bacteria (e.g. cell death processes or the ability of the antibiotic to redox cycle).
authors.affiliation: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.
authors.forename: David T
authors.initials: DT
authors.lastname: Fox
authors.affiliation: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.
authors.forename: Emily N
authors.initials: EN
authors.lastname: Schmidt
authors.affiliation: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.
authors.forename: Hongzhao
authors.initials: H
authors.lastname: Tian
authors.affiliation: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.
authors.forename: Suraj
authors.initials: S
authors.lastname: Dhungana
authors.affiliation: Department of Biology, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Michael C
authors.initials: MC
authors.lastname: Valentine
authors.affiliation: Department of Chemistry, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Nicole V
authors.initials: NV
authors.lastname: Warrington
authors.affiliation: Department of Chemistry, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Paul D
authors.initials: PD
authors.lastname: Phillips
authors.affiliation: Department of Chemistry, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Kellan B
authors.initials: KB
authors.lastname: Finney
authors.affiliation: Department of Biology, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Emily K
authors.initials: EK
authors.lastname: Cope
authors.affiliation: Department of Biology, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Jeff G
authors.initials: JG
authors.lastname: Leid
authors.affiliation: Echelon Biosciences, Inc, Salt Lake City, Utah, United States of America.
authors.forename: Charles A
authors.initials: CA
authors.lastname: Testa
authors.affiliation: Department of Chemistry, Northern Arizona University, Flagstaff, Arizona, United States of America.
authors.forename: Andrew T
authors.initials: AT
authors.lastname: Koppisch
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D053898:Biosynthetic Pathways / Q000187:drug effects; D005578:Fosfomycin / Q000031:analogs & derivatives* / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D045782:Hemiterpenes / Q000096:biosynthesis / Q000737:chemistry; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007612:Kanamycin / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D020411:Oligonucleotide Array Sequence Analysis; D009943:Organophosphorus Compounds / Q000737:chemistry; D018384:Oxidative Stress / Q000187:drug effects*; D012486:Salmonella typhimurium / Q000187:drug effects* / Q000254:growth & development; D013997:Time Factors
chemical_list: D000900:Anti-Bacterial Agents; D045782:Hemiterpenes; D009943:Organophosphorus Compounds; D005578:Fosfomycin; C004809:isopentenyl pyrophosphate; C043060:3,3-dimethylallyl pyrophosphate; C024640:fosmidomycin; D007612:Kanamycin; D006861:Hydrogen Peroxide
references.citation: Science. 2013 Mar 8;339(6124):1210-3
references.pmid: 23471409
references.citation: J Antimicrob Chemother. 2012 Jan;67(1):138-48
references.pmid: 21972270
references.citation: Antimicrob Agents Chemother. 2012 Sep;56(9):4922-6
references.pmid: 22777047
references.citation: Front Microbiol. 2012 Aug 09;3:226
references.pmid: 22905031
references.citation: J Bacteriol. 2001 Dec;183(24):7094-101
references.pmid: 11717267
references.citation: Curr Opin Microbiol. 2009 Oct;12(5):482-9
references.pmid: 19647477
references.citation: Antimicrob Agents Chemother. 2006 Aug;50(8):2713-8
references.pmid: 16870763
references.citation: J Med Chem. 2006 Dec 14;49(25):7331-41
references.pmid: 17149863
references.citation: BMC Evol Biol. 2013 Sep 03;13:180
references.pmid: 24004839
references.citation: J Bacteriol. 2001 Dec;183(24):7173-81
references.pmid: 11717276
references.citation: ACS Chem Biol. 2007 Nov 20;2(11):708-10
references.pmid: 18030985
references.citation: Annu Rev Biochem. 2008;77:755-76
references.pmid: 18173371
references.citation: Science. 1999 Sep 3;285(5433):1573-6
references.pmid: 10477522
references.citation: Trends Biotechnol. 2001 Mar;19(3):109-14
references.pmid: 11179804
references.citation: Microbiology (Reading). 2008 Nov;154(Pt 11):3410-3419
references.pmid: 18957594
references.citation: Int J Antimicrob Agents. 2005 Aug;26(2):146-51
references.pmid: 16024233
references.citation: J Bacteriol. 2008 Apr;190(8):2671-9
references.pmid: 18245294
references.citation: Cell. 2007 Sep 7;130(5):797-810
references.pmid: 17803904
references.citation: Biochemistry. 2002 Jan 8;41(1):236-43
references.pmid: 11772021
references.citation: Biochemistry. 2005 Jun 14;44(23):8360-8
references.pmid: 15938625
references.citation: Proc Natl Acad Sci U S A. 2006 Dec 19;103(51):19484-9
references.pmid: 17148599
references.citation: Biol Proced Online. 2003;5:189-196
references.pmid: 14615815
references.citation: Nat Biotechnol. 2013 Feb;31(2):160-5
references.pmid: 23292609
references.citation: PLoS Pathog. 2011 Dec;7(12):e1002323
references.pmid: 22216001
references.citation: Proc Natl Acad Sci U S A. 2002 Dec 24;99(26):17025-30
references.pmid: 12482953
references.citation: Biosci Biotechnol Biochem. 2003 Sep;67(9):2030-3
references.pmid: 14519998
references.citation: J Bacteriol. 2000 Jan;182(1):23-9
references.pmid: 10613858
references.citation: Curr Opin Microbiol. 2006 Oct;9(5):445-53
references.pmid: 16942902
references.citation: Science. 2013 Mar 8;339(6124):1213-6
references.pmid: 23471410
references.citation: Antimicrob Agents Chemother. 2004 Aug;48(8):2838-44
references.pmid: 15273089
references.citation: Cell. 1985 Jul;41(3):753-62
references.pmid: 2988786
references.citation: J Bacteriol. 2006 Nov;188(22):7988-91
references.pmid: 16980465
references.citation: Int J Antimicrob Agents. 2011 Sep;38(3):261-4
references.pmid: 21724375
references.citation: Proc Natl Acad Sci U S A. 1998 Aug 18;95(17):9879-84
references.pmid: 9707569
references.citation: Luminescence. 2003 Nov-Dec;18(6):334-40
references.pmid: 14694423
references.citation: Gene. 1996 Oct 10;175(1-2):83-7
references.pmid: 8917080
references.citation: Proc Natl Acad Sci U S A. 1986 Nov;83(21):8059-63
references.pmid: 3534881
references.citation: Antimicrob Agents Chemother. 2002 Sep;46(9):2889-94
references.pmid: 12183243
references.citation: J Org Chem. 2005 Mar 18;70(6):1978-85
references.pmid: 15760175
references.citation: Antimicrob Agents Chemother. 2009 Apr;53(4):1395-402
references.pmid: 19223646
references.citation: Bioorg Chem. 2004 Dec;32(6):483-93
references.pmid: 15530989
references.citation: Sci Transl Med. 2013 Jun 19;5(190):190ra81
references.pmid: 23785037
references.citation: Biochem Biophys Res Commun. 2002 Oct 4;297(4):1003-7
references.pmid: 12359254
references.citation: J Infect Dis. 2004 Nov 1;190(9):1534-40
references.pmid: 15478056
references.citation: Mol Microbiol. 2011 Mar;79(5):1136-50
references.pmid: 21226770
references.citation: Mol Syst Biol. 2007;3:91
references.pmid: 17353933
references.citation: Mol Microbiol. 1992 Jun;6(12):1583-91
references.pmid: 1495387
references.citation: Antimicrob Agents Chemother. 2003 Sep;47(9):2903-13
references.pmid: 12936993
references.citation: J Bacteriol. 2004 Jan;186(2):473-80
references.pmid: 14702317
references.citation: Res Microbiol. 2010 Oct;161(8):643-50
references.pmid: 20600858
references.citation: Science. 2010 Oct 1;330(6000):70-4
references.pmid: 20929806
references.citation: Curr Pharm Biotechnol. 2003 Aug;4(4):248-59
references.pmid: 14529427
references.citation: J Bacteriol. 2000 Oct;182(20):5749-56
references.pmid: 11004173
references.citation: J Microbiol Methods. 2006 Jul;66(1):125-35
references.pmid: 16378650
references.citation: J Antibiot (Tokyo). 2011 Jan;64(1):73-8
references.pmid: 21102598
references.citation: Environ Microbiol. 2009 Jan;11(1):126-36
references.pmid: 18793317
references.citation: J Mol Microbiol Biotechnol. 2003;5(2):105-22
references.pmid: 12736533
references.citation: Appl Environ Microbiol. 2011 Aug;77(15):5149-56
references.pmid: 21642396
references.citation: Biochemistry. 2011 May 3;50(17):3570-7
references.pmid: 21438569
references.citation: Mol Cell. 2010 Feb 12;37(3):311-20
references.pmid: 20159551
references.citation: Mol Microbiol. 2003 Oct;50(1):219-30
references.pmid: 14507376
references.citation: J Biol Chem. 2004 Nov 19;279(47):48742-50
references.pmid: 15361522