Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P16104 (H2AX)
 3,930 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Phosphorylation of histone H2AX (termed gamma-H2AX) was recently identified as an early event after induction of DNA double strand breaks (DSBs). We have previously shown that co-exposure to benzo[a]pyrene (BaP), a wide-spread environmental carcinogen, and ultraviolet A (UVA), a major component of solar UV radiation, induced DSBs in mammalian cells. In the present study, we examined whether co-exposure to BaP and UVA generates gamma-H2AX in CHO-K1 cells. Single treatment with BaP (10(-9)-10(-7)M) or UVA ( approximately 2.4 J/cm(2)) did not result in gamma-H2AX, however, co-exposure drastically induced foci of gamma-H2AX in a dose-dependent manner. gamma-H2AX could be detected even at very low concentration of BaP (10(-9)M) plus UVA (0.6J/cm(2)), which did not change cell survival rates. NaN(3) effectively inhibited the formation of gamma-H2AX induced by co-exposure, indicating the contribution of singlet oxygen. This is the first evidence that co-exposure to BaP and UVA induced DSBs, involving gamma-H2AX.
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PMID:Co-exposure to benzo[a]pyrene and UVA induces phosphorylation of histone H2AX. 1625 11

It has been reported that the phosphorylated form of histone variant H2AX (gammaH2AX) plays an important role in the recruitment of DNA repair and checkpoint proteins to sites of DNA damage, particularly at double strand breaks (DSBs). Using gammaH2AX foci formation as an indicator for DNA damage, several chemicals/stress factors were chosen to assess their ability to induce gammaH2AX foci in a 24h time frame in a human amnion FL cell line. Two direct-acting genotoxins, methyl methanesulfonate (MMS) and N-ethyl-N-nitrosourea (ENU), can induce gammaH2AX foci formation in a time- and dose-dependent manner. Similarly, an indirect-acting genotoxin, benzo[a]pyrene (BP), also induced the formation of gammaH2AX foci in a time- and dose-dependent manner. Another indirect genotoxin, 2-acetyl-aminofluorene (AAF), did not induce gammaH2AX foci formation in FL cells; however, AAF can induce gammaH2AX foci formation in Chinese hamster CHL cells. Neutral comet assays also revealed the induction of DNA strand breaks by these agents. In contrast, epigenetic carcinogens azathioprine and cyclosporine A, as well as non-carcinogen dimethyl sulfoxide, did not induce gammaH2AX foci formation in FL cells. In addition, heat shock and hypertonic saline did not induce gammaH2AX foci. Cell survival analyses indicated that the induction of gammaH2AX is not correlated with the cytotoxic effects of these agents/factors. Taken together, these results suggest that gammaH2AX foci formation could be used for evaluating DNA damage; however, the different cell types used may play an important role in determining gammaH2AX foci formation induced by a specific agent.
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PMID:DNA damage evaluated by gammaH2AX foci formation by a selective group of chemical/physical stressors. 1642 55

Polycyclic aromatic hydrocarbons (PAHs), widespread environmental pollutants, were recently reported to show photomutagenesis. As contaminants in the environment are usually exposed to sunlight, a way to evaluate the phototoxic characteristics of pollutants is required. We have previously found that phosphorylation of histone H2AX (gamma-H2AX), which accompanied the induction of DNA double strand breaks (DSBs), was significantly induced by low concentrations of benzo[a]pyrene (10(-9)-10(-7) M) and UVA (0.6 J/cm2) in CHO-K1 cells. Higher concentrations have been required for the detection of DSBs. The aim of the present study is to investigate the applicability of gamma-H2AX in a new phototoxicity assay of PAHs. The human keratinocytes, HaCaT, were treated with four model PAHs (naphthalene, phenanthrene, pyrene, and benzo[a]pyrene, 10(-11)-10(-7) M) and/or UVA (5 J/cm2), and the induction of gamma-H2AX was assessed. Furthermore, DSBs were directly detected using a biased sinusoidal field gel electrophoresis, and the cell viability was examined as a general assay of phototoxicity. The induction of gamma-H2AX was detected in the presence of all the PAHs except naphthalene at concentrations of 10(-9)-10(-7) M, whereas neither DSBs nor cell death could be detected at those concentrations, and higher concentrations were required for the detection. Naphthalene showed no phototoxicity in any of the three different assays. These findings suggest that histone H2AX is a potential moleculartargetfor detecting the phototoxicity of PAHs more sensitively than the detection of cell viability and DSBs.
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PMID:New method for testing phototoxicity of polycyclic aromatic hydrocarbons. 1678

Cell cycle checkpoints are evolutionarily conserved signaling pathways that uphold genomic integrity. Complete inactivation of the mouse checkpoint gene Hus1 results in chromosomal instability, genotoxin hypersensitivity, and embryonic lethality. To determine the functional consequences of partial Hus1 impairment, we generated an allelic series in which Hus1 expression was incrementally reduced by combining a hypomorphic Hus1 allele, Hus1(neo), with either wild-type or null (Hus1(Delta1)) alleles. Primary Hus1(neo/Delta1) embryonic fibroblasts exhibited spontaneous chromosomal abnormalities and underwent premature senescence, while higher Hus1 expression in Hus1(neo/neo) cells allowed for normal proliferation. Antioxidant treatment almost fully suppressed premature senescence in Hus1(neo/Delta1) cultures, suggesting a critical role for Hus1 in oxidative stress responses. Treatment of Hus1(neo/neo) and Hus1(neo/Delta1) cells with the DNA adducting agent benzo(a)pyrene dihydrodriol epoxide resulted in a loss of cell viability that was associated with S-phase DNA damage checkpoint failure. Likewise, the DNA polymerase inhibitor aphidicolin triggered increased cell death, chromosomal aberrations, and H2AX phosphorylation, a marker for double-stranded DNA breaks, in Hus1(neo/neo) and Hus1(neo/Delta1) cultures compared to controls. Despite these pronounced genome maintenance defects in cultured Hus1(neo/Delta1) and Hus1(neo/neo) cells, mice of the same genotypes were born at expected frequencies and appeared grossly normal. A significant increase in micronucleus formation was observed in peripheral blood cells from Hus1(neo/Delta1) mice, but reduced Hus1 expression did not cause an elevated predisposition to spontaneous tumor development or accelerate tumorigenesis in p53-deficient mice. These results identify differential effects of altered Hus1 gene dosage on genome maintenance during in vitro culture, genotoxic stress responses, embryonic development, and adult homeostasis.
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PMID:Genome maintenance defects in cultured cells and mice following partial inactivation of the essential cell cycle checkpoint gene Hus1. 1722 Feb 76

Compounds within the 2-(4-aminophenyl)benzothiazole class represent extremely potent and selective experimental antitumour agents. The lysylamide prodrug of 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole is undergoing phase I clinical evaluation. Extensive studies to elucidate mechanisms underlying the stark selectivity demonstrated potent cytosolic AhR ligand binding and cytochrome P450 1A1-catalysed bioactivation. Two human derived breast cell lines, initially exquisitely sensitive to this class of agent (GI50 < 5 nM) have been derived displaying acquired resistance to 2-(4-amino-3-methylphenyl)benzothiazole (DF 203; GI50 > 50 microM). Cross resistance to 2-(4-amino-3-iodophenyl)benzothiazole and 2-(4-amino-3-cyanophenyl)benzothiazole is observed (GI50 > 30 microM) as is > 100-fold reduced sensitivity of the two variant lines to 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203). In contrast, cell lines possessing acquired resistance to DF 203 (203R) retain sensitivity to benzo[a]pyrene and doxorubicin. Examination of DF 203-treated cells by confocal microscopy and HPLC analyses of nutrient media concur revealing diminished depletion of DF 203 from medium and impaired intracellular DF 203 retention. In contrast to cytosolic arylhydrocarbon (AhR) receptors of wild type cells, AhR appears constitutively localised within nuclei of 203R cells; consequently, DF 203 fails to drive transcription of cyp1a1. DF 203- and 5F 203-derived DNA adducts fall significantly in 203R cells. Reduced number and intensity of gamma H2AX foci report protection against DF 203-evoked DNA double strand breaks. In conclusion, aberrant AhR signalling underlies at least in part acquired resistance to DF 203. Intriguingly, comparisons of gene transcription profiles between sensitive and resistant paired lines reveal > 5-fold up-regulation of cyp1b1 expression, a protein implicated in resistance to therapeutic agents.
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PMID:Mechanisms of acquired resistance to 2-(4-Amino-3-methylphenyl)benzothiazole in breast cancer cell lines. 1767 93

The epidemiological association between cancer and exposure to ambient air pollution particles (particles with a 50% cut-off aerodynamic diameter of 10 microm (PM(10))) has been related to the ability of PM(10) and its constituent nanoparticles (NPs) to cause reactive oxidative species (ROS)-driven DNA damage. However, there are no data on the molecular response to these genotoxic effects. In order to assess whether PM(10), NP and ROS-driven DNA damage induce carcinogenesis pathways, A549 cells were treated with tert-butyl-hyperperoxide (Tbh), urban dust (UD), carbon black (CB), nanoparticulate CB (NPCB), benzo(a)pyrene (BaP) and NPCB coated with BaP for <or=24 h. Single- and double-strand breakage of DNA was determined by comet assay; cell cycle status was analysed using flow cytometry. Nuclear extracts or acid-extracted histones were used for Western blot analysis of p-ser15-p53 (p53 phosphorylated at ser15), p53 binding protein (53BP) 1, phospho-histone H2A.X (p-H2A.X) and phospho-BRCA1 (p-BRCA1). UD caused both single- and double-strand DNA breaks, while other tested NPs caused only single-strand DNA breaks. NPs significantly altered cell cycle kinetics. Tbh enhanced p-H2A.X after 1 and 6 h (2.1- and 2.2-fold, respectively). NP increased 53BP1 expression at 1 h (2.4-8.7-fold) and p-BRCA1 at 1-6 h. N-acetylcysteine blocked NP-driven p-ser15-p53 response. In conclusion, nanoparticles and reactive oxidative species induce DNA damage, activating p53 and proteins related to DNA repair, mimicking irradiation-related carcinogenesis pathways.
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PMID:Nanoparticle-driven DNA damage mimics irradiation-related carcinogenesis pathways. 1805 54

Mdm2 is an oncoprotein interacting with p53 and maintaining low p53 levels in unstressed cells. Here we investigated the effect of genotoxic compounds on Mdm2 phosphorylation levels. Employing the Mdm2 2A10 antibody and phosphatase treatment we found that Mdm2 accumulated in HepG2 cells when exposed to low concentrations of genotoxic compounds such as mitomycin C, etoposide, 5-fluorouracil, and benzo[a]pyrene (BP). The low-dose responses were not accompanied by p53 accumulation and the effect of low concentrations of BP on Mdm2 was not affected by small interfering RNA for p53. In human lymphoblasts 10nM BP induced an Mdm2 response. Low concentrations of BP also induced binding of Mdm2 to chromatin in HepG2 cells, but no p53 binding or H2AX phosphorylation. The more mutagenic dibenzo[a,l]pyrene as well as higher BP concentrations instead induced gammaH2AX and p53 Ser15 association with chromatin. Acrolein potentiated the effect of BP on p53 stabilization and chromatin binding. Taken together, these data suggest that (1) Mdm2 is a sensitive biomarker for certain types of genotoxicity, and (2) that polycyclic aromatic hydrocarbons-induced Mdm2 binding to chromatin reflects repairable damage, whereas chromatin binding of p53 Ser15 and gammaH2AX indicates more persistent DNA damage. The analysis of Mdm2 and related endpoints might be useful for evaluating mutagenic potentials of DNA damages. It is suggested that patterns documented here can be used for separating BP doses that induce readily repaired DNA adducts from doses that overwhelm this capacity.
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PMID:Mdm2 as a sensitive and mechanistically informative marker for genotoxicity induced by benzo[a]pyrene and dibenzo[a,l]pyrene. 1809 71

Polycyclic aromatic hydrocarbons (PAHs), wide-spread mutagenic and carcinogenic environmental pollutants, are consistently exposed to sunlight in the environment. The exposure causes structural change, resulting in the generation of a variety of photomodified products having different bioactivities compared with the parent compounds. In this study, we found that benzo[a]pyrene (BaP) exposed to solar-simulated light (SSL)-induced phosphorylation of histone H2AX (gamma-H2AX), which was recently identified as an early event after the induction of DNA double strand breaks (DSBs). Although BaP itself did not produce gamma-H2AX, SSL-exposed BaP significantly generated gamma-H2AX depending on the period of exposure. Furthermore, we revealed that reactive oxygen species produced by the SSL-exposed BaP mainly contributed to the generation of gamma-H2AX. The appearance of gamma-H2AX means the induction of the most serious form of DNA damage, DSBs, suggesting the potential risk of carcinogenesis.
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PMID:Solar-simulated light-exposed benzo[a]pyrene induces phosphorylation of histone H2AX. 1816 Mar 32

Early events in the cellular response to DNA damage, such as double strand breaks, rely on lesion recognition and activation of proteins involved in maintenance of genomic stability. One important component of this process is the phosphorylation of the histone variant H2AX. To investigate factors explaining the variation in carcinogenic potency between different categories of polycyclic aromatic hydrocarbons (PAHs), we have studied the phosphorylation of H2AX (H2AXgamma). A549 cells were exposed to benzo[a]pyrene diol epoxide [(+)-anti-BPDE] (a bay-region PAH) and dibenzo[a,l]pyrene diol epoxide [(-)-anti-DBPDE] (a fjord-region PAH) and H2AXgamma was studied using immunocytochemistry and Western blot. Hydrogen peroxide (H(2)O(2)) was used to induce oxidative DNA damage and strand breaks. As showed with single cell gel electrophoresis, neither of the diol epoxides resulted in DNA strand breaks relative to H(2)O(2). Visualisation of H2AXgamma formation demonstrated that the proportion of cells exhibiting H2AXgamma staining at 1h differed between BPDE, 40% followed by a decline, and DBPDE, <10% followed by an increase. With H(2)O(2) treatment, almost all cells demonstrated H2AXgamma at 1h. Western blot analysis of the H2AXgamma formation also showed concentration and time-dependent response patterns. The kinetics of H2AXgamma formation correlated with the previously observed kinetics of elimination of BPDE and DBPDE adducts. Thus, the extent of H2AXgamma formation and persistence was related to both the number of adducts and their structural features.
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PMID:H2AX phosphorylation in A549 cells induced by the bulky and stable DNA adducts of benzo[a]pyrene and dibenzo[a,l]pyrene diol epoxides. 1884 25

3,6-Dinitrobenzo[e]pyrene (3,6-DNBeP), newly identified in airborne particles and surface soil, is a potent mutagen in Salmonella typhimurium. The present study investigated the genotoxic potency of 3,6-DNBeP in vitro and in vivo using mammalian cell strains (Chinese hamster CHL/IU and human HepG2) and ICR mice, respectively. In the hprt gene mutation assay using HepG2 cells, the spontaneous mutant frequency was 61.1 per 10(5) clonable cells, which increased to 229 per 10(5) clonable cells after treatment with 1.0 microg/ml (3 microM) 3,6-DNBeP. Notably, in HepG2 cells with increased N-acetyltransferase 2 activity, the mutant frequency increased to 648 per 10(5) clonable cells by treatment of 1.0 microg/ml (3 microM) 3,6-DNBeP. The sister chromatid exchange frequency increased approximately three times the control level in HepG2 cells treated with 3,6-DNBeP at a concentration of 1.0 microg/ml (3 microM). In HepG2 and CHL/IU cells, the frequency of the cells with micronuclei was 0.9 and 1.2%, and the frequencies increased to 2.3 and 7.6% after 1.0 microg/ml (3 microM) 3,6-DNBeP-treatment, respectively. The H2AX phosphorylation level increased 8-fold compared with the background level with 1.0 microg/ml (3 microM) 3,6-DNBeP-treatment in HepG2 cells. Moreover, the comet assay showed that 3,6-DNBeP produced DNA damage in the cells of liver, kidney, lung and bone marrow in ICR mice 3 h after intraperitoneal injection at 40 mg/kg (0.12 mmol/kg) body weight. These data indicate that 3,6-DNBeP is genotoxic to mammalian cells in vitro and in vivo.
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PMID:Genotoxicity of 3,6-dinitrobenzo[e]pyrene, a novel mutagen in ambient air and surface soil, in mammalian cells in vitro and in vivo. 1927 66


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