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)

Human microvascular endothelial cells (HMEC) were exposed to ionizing radiation at doses ranging from 0 to 16 Gy in either the presence or absence of the active thiol forms of amifostine (WR1065), phosphonol (WR255591), N-acetyl-l-cysteine (NAC), captopril or mesna. Each of these clinically relevant thiols, administered to HMEC at a dose of 4 mM for 30 min prior to irradiation, is known to exhibit antioxidant properties. The purpose of this investigation was to determine the relationship(s), if any, between the frequency of radiation-induced histone H2AX phosphorylation at serine 139 (gamma-H2AX) in cells and subsequent survival, as assessed by colony-forming ability, in exposed cell populations as a function of the presence or absence of each of the five thiol compounds during irradiation. gamma-H2AX formation in irradiated cells, as a function of relative DNA content, was quantified by bivariant flow cytometry analysis with FITC-conjugated gamma-H2AX antibody and nuclear DAPI staining. gamma-H2AX formation in cells was measured as the relative fold increase as a function of the treatment conditions. The frequency of gamma-H2AX-positive cells increased with increasing dose of radiation followed by a dose- and time-dependent decay. The most robust response for gamma-H2AX formation occurred 1 h after irradiation with their relative frequencies decreasing as a function of time 4 and 24 h later. To assess the effects of the various thiols on gamma-H2AX formation, all measurements were made 1 h after irradiation. WR1065 was not only effective in protecting HMEC against gamma-H2AX formation across the entire dose range of radiation exposures used, but it was also significantly more cytoprotective than either its prodrug (WR2721) or disulfide (WR33278) analogue. WR1065 had no significant effect on gamma-H2AX formation when administered immediately or up to 30 min after radiation exposure. An inhibitory effect against gamma-H2AX formation induced by 8 Gy of radiation was expressed by each of the thiols tested. NAC, captopril and mesna were equally effective in reducing the frequency of gamma-H2AX formation, with both WR1065 and WR255591 exhibiting a slightly more robust protective effect. Each of the five thiols was effective in reducing the frequency of gamma-H2AX-positive cells across all phases of the cell cycle. In contrast to the relative ability of each of these thiols to inhibit gamma-H2AX formation after irradiation, NAC, captopril and mesna afforded no protection to HMEC as determined using a colony-forming survival assay. Only WR1065 and WR255591 were effective in reducing the frequencies of radiation-induced gamma-H2AX-positive cells as well as protecting against cell death. These results suggest that the use of gamma-H2AX as a biomarker for screening the efficacy of novel antioxidant radioprotective compounds is highly problematic since their formation and disappearance may be linked to processes beyond simply the formation and repair of radiation-induced DSBs.
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PMID:Relationship between phosphorylated histone H2AX formation and cell survival in human microvascular endothelial cells (HMEC) as a function of ionizing radiation exposure in the presence or absence of thiol-containing drugs. 1772 2

We previously reported that the Polo-like Kinase 2 gene (Plk2/Snk) is a direct target for transcriptional regulation by p53 and that silencing Plk2 sensitizes cancer cells to Taxol-induced apoptosis. Our goals have been to better understand why Plk2 is regulated by p53 and how Plk2 signals protection from cell death through checkpoint activation. We found that following knock-down of Plk2 in wild-type p53 expressing H460 human non-small cell lung cancer cells there was a significant increase in cell death observed in aphidicolin-treated cells and a further increase after release from aphidicolin-block. The highest levels of cell death were observed when Plk2-deficient cells were released from both aphidicolin and etoposide treatment. These results suggested that a defective S-phase checkpoint may contribute to enhanced sensitivity of Plk2-deficient cells to replication stress. Consistent with this hypothesis, we observed higher levels of Serine 139 H2AX phosphorylation in Plk2-deficient as compared to control cells before and after aphidicolin treatment indicating that there is more DNA damage when Plk2 is depleted. We also observed higher levels of Chk1 protein in Plk2-deficient cells that were associated with reduced levels of Serine 317-phosphorylated Chk1. In aphidicolin-treated cells, there were lower levels of Serine 317-phosphorylated Chk1 when Plk2 was knocked-down. Plk2 was demonstrated to interact with Chk2, Chk1, Serine 317-phosphorylated Chk1 and p53. Thus, increased cell death observed after aphidicolin treatment and release in Plk2-deficient cells may result from both higher levels of replication stress-induced DNA damage and a dysfunctional S-phase checkpoint.
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PMID:Replication stress, defective S-phase checkpoint and increased death in Plk2-deficient human cancer cells. 1791 33

H2AX may be one family of histone and distributes among the genome. It is reported that DNA double-strand breaks (DSB) could induce phosphorylation of H2AX at conservative C-terminal region of serine 139 and the formation of gamma-H2AX. The gamma-H2AX foci can be observed by the fluorescence microscope. An immunofluorescence assay with specific antibodies recognizing gamma-H2AX could become the gold standard for detecting the DSB. It was indicated that some of physical, chemical, and biological factors could induce the formation of the gamma-H2AX foci. In this paper, the advance of the studies of mechanism of gamma-H2AX formation and the association between gamma-H2AX and DSB were reviewed.
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PMID:[Association between gamma-H2AX and DNA double-strand breaks]. 1795 27

Carbon nanotubes (CNTs) have shown promise as an important new class of multifunctional building blocks and innovative tools in a large variety of applications, ranging from nanocomposite materials through nanoelectronics to biomedical devices. Because of their unusual one-dimensional hollow nanostructure and unique physicochemical properties, CNTs are particularly useful as novel drug delivery tools and imaging agents. However, such biomedical applications will not be realized if there is no proper assessment of the potential hazards of CNTs to humans and other biological systems. Although a few reports on the cytotoxicity of CNTs have been published, very little is known about the toxicity at the molecular level, or genotoxicity, of CNTs in mammalian cells. We have for the first time assessed the DNA damage response to multiwalled carbon nanotubes (MWNTs) in mouse embryonic stem (ES) cells. We found that MWNTs can accumulate and induce apoptosis in mouse ES cells and activate the tumor suppressor protein p53 within 2 h of exposure. Furthermore, we also observed increased expression of two isoforms of base excision repair protein 8-oxoguanine-DNA glycosylase 1 (OGG1), double strand break repair protein Rad 51, phosphorylation of H2AX histone at serine 139, and SUMO modification of XRCC4 following the treatment with MWNTs. A mutagenesis study using an endogenous molecular marker, adenine phosphoribosyltransferase (Aprt), showed that MWNTs increased the mutation frequency by 2-fold compared with the spontaneous mutation frequency in mouse ES cells. These results suggest that careful scrutiny of the genotoxicity of nanomaterials is needed even for those materials, like multiwalled carbon nanotubes, that have been previously demonstrated to have limited or no toxicity at the cellular level.
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PMID:DNA damage induced by multiwalled carbon nanotubes in mouse embryonic stem cells. 1804 46

Nucleotide excision repair (NER) eliminates helix-distorting DNA base lesions. Seven XP-deficient genetic complementation groups (XPA to XPG) have already been identified in mammals, and their corresponding genes have been cloned. Hereditary defects in NER are associated with several diseases, including xeroderma pigmentosum (XP). UV-DDB (XPE) is formed by two associated subunits, DDB1 and DDB2. UV-DDB was identified biochemically as a protein factor that exhibits very strong and specific binding to ultraviolet (UV)-treated DNA. As a preliminary step to characterize the components of the NER in the filamentous fungus Aspergillus nidulans, here we identified a putative DDB1 homologue, DdbA. Deletion and expression analysis indicated that A. nidulans ddbA gene is involved in the DNA damage response, more specifically in the UV light response and 4-nitroquinoline oxide (4-NQO) sensitivity. Furthermore, the DeltaddbA strain cannot self-cross and expression analysis showed that ddbA can be induced by oxidative stress and is developmentally regulated in both asexual and sexual processes. The DeltaddbA mutation can genetically interact with uvsB (ATR), atmA(ATM), nkuA (KU70), H2AX-S129A (a replacement of the conserved serine in the C-terminal of H2AX with alanine), and cshB (a mutation in CSB Cockayne's syndrome protein involved in the transcription-coupled repair subpathway of NER) mutations. Finally, to determine the DdbA cellular localization, we constructed a GFP::DdbA strain. In the presence and absence of DNA damage, DdbA was mostly detected in the nuclei, indicating that DdbA localizes to nuclei and its cellular localization is not affected by the cellular response to DNA damage induced by 4-NQO and UV light.
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PMID:Functional characterization of the putative Aspergillus nidulans DNA damage binding protein homologue DdbA. 1806 Apr 32

In recent years, several histone modifications have been implicated in the cellular response to DNA double-strand breaks (DSBs). One of the best characterized histone modifications important in DSB repair is the phosphorylation of histone H2A variant, H2A.X. In response to DSBs, H2A.X is phosphorylated and this phosphorylation is required for DSB signaling and the retention of repair proteins at the break site. Despite the existing picture that the function of H2A.X is to promote DNA repair, very recent data suggest that the phosphorylation of histone H2A.X has additional functions. This is analogous to histone H3 phosphorylation on serine 10, which participates in seemingly incompatible functions--transcriptional activation and mitosis. In this review, we discuss the role of histone H2A.X in maintaining genomic stability and review emerging evidence that histone H2A.X is multifunctional.
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PMID:The gamma-H2A.X: is it just a surrogate marker of double-strand breaks or much more? 1809 27

Kodym, E., Kodym, R., Choy, H. and Saha, D. Sustained Metaphase Arrest in Response to Ionizing Radiation in a Non-small Cell Lung Cancer Cell Line. Radiat. Res. 169, 46-58 (2008). In solid tumors, non-apoptotic forms of tumor cell inactivation such as mitotic catastrophe appear to be predominant in the response to DNA-damaging agents. Despite its importance, the underlying molecular mechanisms of mitotic catastrophe have been only partially elucidated. We found that a large fraction of HCC2279 non-small cell lung cancer cells underwent mitotic catastrophe after irradiation. Cells were arrested in metaphase with chromosomal damage indicated by DNA fragments displaced from the metaphase plate and considerable numbers of residual gamma-H2AX foci. Although TP53 was nonfunctional, we detected a prompt radiation response on the level of checkpoint kinases. In contrast, CDC25A was the only checkpoint phosphatase that was responsive to radiation. CDC25B was not detectable, and CDC25C was constitutively phosphorylated at serine 216, leading to its cytoplasmic sequestration and functional inactivation. Therefore, radiation-induced mitotic catastrophe in HCC2279 cells appears to be induced by a combination of relative insufficiencies in the p53-mediated and checkpoint kinase-mediated pathways leading to premature entry into mitosis. Displaced chromosome fragments triggering an intra-M checkpoint in cells entering mitosis presumably result in a sustained metaphase arrest. The phenomenon found in these cells, which were derived directly from a human patient, might be responsible for therapy-induced genetic instability of tumors.
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PMID:Sustained metaphase arrest in response to ionizing radiation in a non-small cell lung cancer cell line. 1815 51

Sp1, a transcription factor that regulates expression of a wide array of essential genes, contains two SQ/TQ cluster domains, which are characteristic of ATM kinase substrates. ATM substrates are transducers and effectors of the DNA damage response, which involves sensing damage, checkpoint activation, DNA repair, and/or apoptosis. A role for Sp1 in the DNA damage response is supported by our findings: Activation of ATM induces Sp1 phosphorylation with kinetics similar to H2AX; inhibition of ATM activity blocks Sp1 phosphorylation; depletion of Sp1 sensitizes cells to DNA damage and increases the frequency of double strand breaks. We have identified serine 101 as a critical site phosphorylated by ATM; Sp1 with serine 101 mutated to alanine (S101A) is not significantly phosphorylated in response to damage and cannot restore increased sensitivity to DNA damage of cells depleted of Sp1. Together, these data show that Sp1 is a novel ATM substrate that plays a role in the cellular response to DNA damage.
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PMID:Phosphorylation of Sp1 in response to DNA damage by ataxia telangiectasia-mutated kinase. 1817 90

Although DNA-damaging agents are among the most effective anticancer drugs in clinical use, their overall effectiveness is limited by the development of cross-resistance to these drugs. Given that histone deacetylase (HDAC) inhibitors increase the acetylation of core histones, resulting in an open chromatin configuration that is more accessible to DNA-targeting agents, we examined whether HDAC inhibitors might enhance the cytotoxicity of DNA-damaging drugs in six human ovarian tumor cell lines that exhibit different cisplatin sensitivities. Low concentrations of HDAC inhibitors, which alone exhibited little cytotoxicity, markedly enhanced the induction of apoptotic cell death not only by cisplatin but also by a wide variety of DNA-targeting anticancer drugs in these tumor cell lines, irrespective of their sensitivities to the respective drugs. In contrast, HDAC inhibitors did not increase the cytotoxicity of metabolic antagonists or microtubule-targeting agents. HDAC inhibitors potentiated both the phosphorylation of histone H2AX on serine-139 (a marker of DNA double-strand breaks) as well as the accumulation of reactive oxygen species induced by DNA-damaging agents in tumor cells. The enhanced generation of reactive oxygen species appeared to be responsible for the enhanced apoptotic cell death induced by the combination of these drugs. These results indicate that the combination of an HDAC inhibitor with a wide variety of DNA-damaging agents is a promising chemotherapeutic strategy for the eradication of tumor cells, regardless of whether the cells are sensitive or resistant to the DNA-damaging anticancer drugs.
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PMID:Histone deacetylase inhibitors enhance the chemosensitivity of tumor cells with cross-resistance to a wide range of DNA-damaging drugs. 1820 Dec 78

Phosphorylated histone H3 at serine 10 and serine 28 (H3Ser10 and H3Ser28) have been recognized as cell cycle markers to evaluate the late-G(2)/M status of cell and tissue samples. I report about the reduction phenomena of H3Ser10 and Ser28 phosphorylation (H3Ser10P and 28P) at the late-G(2) through M cell cycle phases in association with DNA damage caused by hydrogen peroxide (H(2)O(2)). The levels of H3Ser10P and Ser28P decreased between 15 and 60 min after H(2)O(2) addition in an inverse correlation manner with H2AX Ser139 phosphorylation (gammaH2AX). Experiments using wortmannin suggested that the reduction events of H3Ser10P/28P are under the control of phosphatidylinositol 3-kinase-like kinases. Fluorescence microscopic observation showed that H3Ser10 and Ser28 on telomeric portions of condensed M-phase chromosomes retained their strongly phosphorylated status even after 60 min of H(2)O(2) treatment. In addition, these chromosome parts were poorly gammaH2AX positive showing mutually exclusive distribution patterns between H3Ser10P/28P and gammaH2AX. Considering these data, I hypothesize that the reduction of the H3Ser10P/28P is a part of the DNA damage response processes. It is advisable to pay careful attention to these phenomena at the time of designing cell cycle assay protocols with H3Ser10P or Ser28P mitosis markers when DNA damaging process is expected to occur.
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PMID:Reduction of phosphorylated histone H3 serine 10 and serine 28 cell cycle marker intensities after DNA damage. 1839 32


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