Interindividual variability in response to sodium dichromate-induced oxidative DNA damage: role of the Ser326Cys polymorphism in the DNA-repair protein of 8-oxo-7,8-dihydro-2'-deoxyguanosine DNA glycosylase 1.
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Although the genotoxic mechanism(s) of hexavalent chromium (CrVI) carcinogenicity remain to be fully elucidated, intracellular reduction of CrVI and concomitant generation of reactive intermediates including reactive oxygen species and subsequent oxidative damage to DNA is believed to contribute to the process of carcinogenesis. In the current study, substantial interindividual variation (7.19-25.84% and 8.79-34.72% tail DNA as assessed by conventional and FPG-modified comet assay, respectively) in levels of DNA strand breaks after in vitro treatment of WBC with sodium dichromate (100 micromol/L, 1 hour) was shown within a group of healthy adult volunteers (n = 72) as assessed by both comet and formamidopyrimidine glycosylase-modified comet assays. No statistically significant correlation between glutathione S-transferases M1 or T1, NADPH quinone oxidoreductase 1 (codon 187) and X-ray repair cross complementation factor 1 (codon 194) genotypes and individual levels of DNA damage were observed. However, individuals homozygous for the Cys(326) 8-oxo 7,8-dihydro-2'-deoxyguanosine glycosylase 1 (OGG1) polymorphism had a statistically significant elevation of formamidopyrimidine glycosylase-dependent oxidative DNA damage after treatment with sodium dichromate when compared with either Ser(326)/Ser(326) or Ser(326)/Cys(326) individuals (P = 0.008 and P = 0.003, respectively). In contrast, no effect of OGG1 genotype on background levels of oxidative DNA damage was observed. When individuals were divided on the basis of OGG1 genotype, Cys(326)/Cys(326) individuals had a statistically significant (P < 0.05, one-way ANOVA followed by Tukey test) higher ratio of oxidative DNA damage to plasma antioxidant capacity than either Ser(326)/Ser(326) or Ser(326)/Cys(326) individuals. The results of this study suggest that the Cys(326)/Cys(326) OGG1 genotype may represent a phenotype that is deficient in the repair of 8-oxo-7,8-dihydro-2'-deoxyguanosine, but only under conditions of cellular oxidative stress. We hypothesize that this may be due to oxidation of the Cys(326) residue. In conclusion, the homozygous Cys(326) genotype may represent a biomarker of individual susceptibility of lung cancer risk in individuals that are occupationally exposed to CrVI. (+info)
ChrR, a soluble quinone reductase of Pseudomonas putida that defends against H2O2.
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Most bacteria contain soluble quinone-reducing flavoenzymes. However, no biological benefit for this activity has previously been demonstrated. ChrR of Pseudomonas putida is one such enzyme that has also been characterized as a chromate reductase; yet we propose that it is the quinone-reducing activity of ChrR that has the greatest biological significance. ChrR reduces quinones by simultaneous two-electron transfer, avoiding formation of highly reactive semiquinone intermediates and producing quinols that promote tolerance of H(2)O(2). Expression of chrR was induced by H(2)O(2), and levels of chrR expression in overexpressing, wild type, and knock-out mutant strains correlated with the H(2)O(2) tolerance and scavenging ability of each strain. The chrR expression level also correlated with intracellular H(2)O(2) levels as measured by protein carbonylation assays and fluorescence-activated cell scanning analysis with the H(2)O(2)-responsive dye H(2)DCFDA. Thus, enhancing the activity of ChrR in a chromate-remediating bacterial strain may not only increase the rate of chromate transformation, it may also augment the capacity of these cells to withstand the unavoidable production of H(2)O(2) that accompanies chromate reduction. (+info)
Carcinogenic lead chromate induces DNA double-strand breaks in human lung cells.
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Hexavalent chromium (Cr(VI)) is a widespread environmental contaminant and a known human carcinogen, generally causing bronchial cancer. Recent studies have shown that the particulate forms of Cr(VI) are the potent carcinogens. Particulate Cr(VI) is known to induce a spectrum of DNA damage such as DNA single strand breaks, Cr-DNA adducts, DNA-protein crosslinks and chromosomal aberrations. However, particulate Cr(VI)-induced DNA double strand breaks (DSBs) have not been reported. Thus, the aim of this study was to determine if particulate Cr(VI)-induces DSBs in human bronchial cells. Using the single cell gel electrophoresis assay (comet assay), showed that lead chromate-induced concentration dependent increases in DSBs with 0.1, 0.5, 1 and 5 microg/cm2 lead chromate inducing a 20, 50, 67 and 109% relative increase in the tail integrated intensity ratio, respectively. Sodium chromate at concentrations of 1, 2.5 and 5 microM induced 38, 78 and 107% relative increase in the tail integrated intensity ratio, respectively. We also show that genotoxic concentrations of lead chromate activate the ataxia telangiectasia mutated (ATM) protein, which is thought to play a central role in the early stages of DSB detection and controls cellular responses to this damage. The H2A.X protein becomes rapidly phosphorylated on residue serine 139 in cells when DSBs are introduced into the DNA by ionizing radiation. By using immunofluorescence, we found that lead chromate-induced concentration-dependent increases in phosphorylated H2A.X (r-H2A.X) foci formation with 0.1, 0.5, 1, 5 and 10 microg/cm2 lead chromate inducing a relative increase in the number of cells with r-H2A.X foci formation of 43, 51, 115 and 129%, respectively. (+info)
Nei deficient Escherichia coli are sensitive to chromate and accumulate the oxidized guanine lesion spiroiminodihydantoin.
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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. (+info)
Resistance to apoptosis, increased growth potential, and altered gene expression in cells that survived genotoxic hexavalent chromium [Cr(VI)] exposure.
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Certain hexavalent chromium [Cr(VI)] compounds are known genotoxic respiratory carcinogens, which induce apoptosis as a predominant mode of cell death. Selection of cells that are resistant to apoptosis may be a factor in tumour progression. We developed sub-populations of telomerase-transfected human fibroblasts (BJ-hTERT) that survived a 99% clonogenically lethal exposure to Cr(VI) (B-5Cr). B-5Cr cells were markedly resistant to apoptosis induced by several agents and exhibited increased clonogenic survival, especially at apoptogenic doses. B-5Cr cells did not exhibit altered cellular uptake of Cr(VI) and retained a normal p53 response to Cr(VI) exposure. We conducted large-scale gene expression analysis at different time-points after a secondary genotoxic Cr(VI) insult in B-5Cr and BJ-hTERT cells using Affymetrix Genechip human genome arrays. Cr(VI) exposure led to differential regulation of many genes, which affect a diverse set of cellular activities such as transcription, signal transduction, stress response, cell adhesion, DNA repair, apoptosis and cell cycle modulation. We compared Cr(VI)-induced altered gene expression in the B-5Cr cells to that in the parental cells and identified 223, 147 and 204 genes with at least a two-fold difference in expression at 4, 8 and 18 h after exposure, respectively. Cluster analysis by gene function revealed altered expression of genes involved in apoptosis, cell cycle regulation and DNA repair. Our data suggest an alteration in gene expression that may favor cell survival and/or incomplete DNA repair after genotoxic exposure. Selection of cells with altered expression of these genes may constitute the early stages of tumour progression. (+info)
Expression of yeast transcriptional activator MSN1 promotes accumulation of chromium and sulfur by enhancing sulfate transporter level in plants.
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MSN1 is a putative yeast transcriptional activator involved in chromium (Cr) accumulation. Here we show that overexpression of MSN1 enhances Cr and sulfur accumulation and Cr tolerance in transgenic tobacco. In addition, we found that expression of NtST1 (Nicotiana tabacum sulfate transporter 1) was elevated in MSN1- expressing transgenic tobacco, suggesting that chromate and sulfate are taken up via the sulfate transporter in plants. Supporting this, expression of NtST1 increased levels of Cr and S in Saccharomyces cerevisiae. Our findings suggest that yeast transcriptional activators can be used for developing effective metal remediators, and for improving the nutritional status of plants. (+info)
A cohort study of bronchial carcinomas in workers producing chromate pigments.
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A cohort study of the incidence of bronchial cancer in male workers in a small company producing chromate pigments is presented. Altogether 133 workers had been employed by the company from the time production was started in 1948 until the end of 1972. Workers with more than three years employment were included in the study, and three cases of bronchial carcinoma were found among the 24 workers who fulfilled this requirement. Based on the data of the Cancer Registry of Norway the risk of bronchial cancer for a corresponding group of the general population was found to be 0.079, which gives a risk ratio for exposed workers of approximately 38. The average age of the cancer patients was as low as 50 years at the time of diagnosis. All workers in the company had been exposed mainly to zinc chromate dust, and the exposure levels of the workers developing bronchial cancers had probably been from 0-5 to 1-5 mg Cr/m-3 for six to nine years. Two of the three patients were smokers. It is assumed that exposure to chromate pigments, and probably to zinc chromate, may be related to the increased incidence of bronchial cancer in this group of workers. The possibility of a contributing effect of tobacco smoking in at least two of the three cases cannot be ruled out. (+info)
Molecular dynamics of the Shewanella oneidensis response to chromate stress.
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Temporal genomic profiling and whole-cell proteomic analyses were performed to characterize the dynamic molecular response of the metal-reducing bacterium Shewanella oneidensis MR-1 to an acute chromate shock. The complex dynamics of cellular processes demand the integration of methodologies that describe biological systems at the levels of regulation, gene and protein expression, and metabolite production. Genomic microarray analysis of the transcriptome dynamics of midexponential phase cells subjected to 1 mm potassium chromate (K(2)CrO(4)) at exposure time intervals of 5, 30, 60, and 90 min revealed 910 genes that were differentially expressed at one or more time points. Strongly induced genes included those encoding components of a TonB1 iron transport system (tonB1-exbB1-exbD1), hemin ATP-binding cassette transporters (hmuTUV), TonB-dependent receptors as well as sulfate transporters (cysP, cysW-2, and cysA-2), and enzymes involved in assimilative sulfur metabolism (cysC, cysN, cysD, cysH, cysI, and cysJ). Transcript levels for genes with annotated functions in DNA repair (lexA, recX, recA, recN, dinP, and umuD), cellular detoxification (so1756, so3585, and so3586), and two-component signal transduction systems (so2426) were also significantly up-regulated (p < 0.05) in Cr(VI)-exposed cells relative to untreated cells. By contrast, genes with functions linked to energy metabolism, particularly electron transport (e.g. so0902-03-04, mtrA, omcA, and omcB), showed dramatic temporal alterations in expression with the majority exhibiting repression. Differential proteomics based on multidimensional HPLC-MS/MS was used to complement the transcriptome data, resulting in comparable induction and repression patterns for a subset of corresponding proteins. In total, expression of 2,370 proteins were confidently verified with 624 (26%) of these annotated as hypothetical or conserved hypothetical proteins. The initial response of S. oneidensis to chromate shock appears to require a combination of different regulatory networks that involve genes with annotated functions in oxidative stress protection, detoxification, protein stress protection, iron and sulfur acquisition, and SOS-controlled DNA repair mechanisms. (+info)