title: Nei deficient Escherichia coli are sensitive to chromate and accumulate the oxidized guanine lesion spiroiminodihydantoin.
abstract: Growth inhibition and oxidized guanine lesion formation were studied in a number of base excision repair (BER) deficient Escherichia coli (E. coli) following chromate exposure. The only BER deficient bacterial strain that demonstrated significant growth inhibition by chromate, in comparison to its matched wild-type cell line, was the Nei deficient (TK3D11). HPLC coupled with electrospray ionization mass spectrometry showed that the Nei deficient E. coli accumulated the further oxidized guanine lesion, spiroiminodihydantoin (Sp), in genomic DNA at levels that were approximately 20-fold greater than its wild-type counterpart. However, no accumulation of the putative intermediate of Sp, 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxodG), was observed in the Nei deficient strain. A MutM-/MutY- double deletion mutant that was deficient in BER enzymes for the recognition and repair of 8-oxodG demonstrated no sensitivity toward chromate nor was there an associated increase in Sp accumulation over that of its wild type. However, the MutM-/MutY- double deletion mutant did show approximately 20-fold accumulation of 8-oxodG upon chromate exposure over that of the wild type and the Nei deficient E. coli. These data demonstrate that the Nei BER enzyme is critical for the recognition and repair of the Sp lesion in bacterial cell lines and demonstrates the protective effect of a specific BER enzyme on DNA lesions formed by chromate. To our knowledge, these are the first studies to show the formation and biological significance of the Sp lesion in a cellular system. This study has significant mechanistic and toxicological implications for how chromate may serve as an initiator of carcinogenesis and suggests a role for specific repair enzymes that may ameliorate the carcinogenic potential of chromate.
authors.affiliation: Department of Chemistry, The University of Montana, Missoula, Montana 59812, USA.
authors.forename: M Katie
authors.initials: MK
authors.lastname: Hailer
authors.forename: Peter G
authors.initials: PG
authors.lastname: Slade
authors.forename: Brooke D
authors.initials: BD
authors.lastname: Martin
authors.forename: Kent D
authors.initials: KD
authors.lastname: Sugden
mesh_terms: D049109:Cell Proliferation / Q000187:drug effects; D002840:Chromates / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D043224:Deoxyribonuclease (Pyrimidine Dimer) / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D016680:Genome, Bacterial / Q000235:genetics; D006147:Guanine / Q000737:chemistry / Q000378:metabolism*; D006151:Guanosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D015394:Molecular Structure; D010084:Oxidation-Reduction; D021241:Spectrometry, Mass, Electrospray Ionization; D013141:Spiro Compounds / Q000737:chemistry / Q000378:metabolism*
chemical_list: D002840:Chromates; D029968:Escherichia coli Proteins; D013141:Spiro Compounds; C408822:spiroiminodihydantoin; D006151:Guanosine; D006147:Guanine; D043224:Deoxyribonuclease (Pyrimidine Dimer); C468346:Nei protein, E coli