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)

Adenoviral proteins interact with host-cell proteins to either exploit or inhibit cellular functions for the purpose of viral propagation. E4orf6, the 34-kDa gene product of the E4 gene, interacts with the double-strand break repair (DSBR) protein DNA-dependent protein kinase and cooperates with binding partner E1B-55K to degrade MRE11, preventing viral DNA concatemer formation. We previously demonstrated that E4orf6 radiosensitizes human tumor cells through the inhibition of DSBR, notably in the absence of E1B-55K. Here, we report that E4orf6 prolongs the signaling of DNA damage by inhibiting the activity of protein phosphatase 2A (PP2A), the phosphatase responsible for dephosphorylating gammaH2AX. The inhibition of PP2A occurs without significant disruption of the DNA re-ligation rate. Prolonged signaling of DNA damage in the presence of E4orf6 initiates caspase-dependent and independent cell death. This is accompanied by poly(ADP-ribose) polymerase (PARP) hyperactivation and the translocation of apoptosis-inducing factor (AIF) from the mitochondria to the nucleus. Knockdown of AIF by shRNA rescues the radiosensitization induced by E4orf6. Taken together, these data suggest that E4orf6 disrupts cellular DSBR signaling by inhibiting PP2A, leading to prolonged H2AX phosphorylation, hyperactivation of PARP, and AIF translocation to the nucleus. The function of E4orf6 as an inhibitor of PP2A and activator of PARP in the absence of other adenoviral gene products is of importance in delineating the adenovirus-host cell interplay.
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PMID:The adenoviral E4orf6 protein induces atypical apoptosis in response to DNA damage. 1717 68

Interactions between tumor cells and their substratum influence cancer progression by modulating cell proliferation and survival. We now investigated whether signaling responses to UV irradiation differ on adhesion-permissive or restrictive substrates. The latter conditions diminished spreading and proliferation of neo 6.3/C8161 melanoma in which metastasis is suppressed by introduction of neo-tagged chromosome 6, but permitted proliferation of human metastatic C8161 melanoma. Apoptosis-associated PARP cleavage and DNA fragmentation induced by UV irradiation were diminished on the restrictive substrate in C8161 melanoma. Genotoxic responses to UV irradiation like persistent increases in the phosphorylation of histone H2AX, induction of the tumor suppressor p53 protein and greater binding of this protein to its DNA consensus sequence, were all decreased on the restrictive substrate. The latter also promoted a 2 fold increase of DNA condensation in chromatin and enhanced activation of the survival - and invasion-associated MMP-9 gelatinase B, preferentially in metastatic C81261 melanoma. Our data suggest that adaptation to restrictive substrates in metastatic C8161 melanoma decreases UV-induced apoptosis, partly through attenuation of DNA damage signaling responses and changes in genomic organization.
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PMID:Attenuation of genotoxicity under adhesion-restrictive conditions through modulation of p53, gamma H2AX and nuclear DNA organization. 1720 47

We have recently shown that thymoquinone (TQ) is an antineoplastic drug that induces p53-dependent apoptosis in human colon cancer cells. This study evaluated the antiproliferative and pro-apoptotic effects of TQ in two human osteosarcoma cell lines with different p53 mutation status. TQ decreased cell survival dose-dependently and, more significantly, in p53-null MG63 cells (IC(50) = 17 muM) than in p53-mutant MNNG/HOS cells (IC(50) = 38 muM). Cell viability was reduced more selectively in MG63 tumor cells than in normal human osteoblasts. Flow cytometric analysis showed that TQ induced a much greater increase in the PreG(1) (apoptotic) cell population, but no cell cycle arrest in MG63. G(2)/M arrest in MNNG/HOS cells was associated with p21(WAF1) upregulation. Using three DNA damage assays, TQ was confirmed to result in a significantly greater extent of apoptosis in p53 null MG63 cells. Although the Bax/Bcl-2 ratios were not differentially modulated in both cell lines, the mitochondrial pathway appeared to be involved in TQ-induced apoptosis in MG63 by showing the cleavage of caspases-9 and -3. Oxidative stress and mitochondrial O(2)(*-) generation in isolated rat mitochondria were enhanced by TQ as measured by the dose-dependent reduction in aconitase enzyme activity and Amplex Red oxidation respectively. TQ-induced oxidative damage, reflected by an increase in gamma-H2AX foci and increased protein expression levels of gamma-H2AX and the DNA repair enzyme, NBS1, was more pronounced in MNNG/HOS than in MG63. We suggest that the resistance of MNNG/HOS cells to drug-induced apoptosis is caused by the up-regulation of p21(WAF1) by the mutant p53 (transcriptional activity was shown by p53 siRNA treatment) which induces cell cycle arrest and allows to repair DNA damage. Collectively, these findings show that TQ induces p53-independent apoptosis in human osteosarcoma cells. As the loss of p53 function is frequently observed in osteosarcoma patients, our data suggest the potential clinical usefulness of TQ for the treatment of these malignancies.
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PMID:Lack of p53 augments thymoquinone-induced apoptosis and caspase activation in human osteosarcoma cells. 1721 78

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

Tumor suppressor gene BRCA1 is frequently mutated in familial breast and ovarian cancer. BRCA1 plays pivotal roles in maintaining genomic stability by interacting with numerous proteins in cell cycle control and DNA repair. Irofulven (6-hydroxymethylacylfulvene, HMAF, MGI 114, NSC 683863) is one of a new class of anticancer agents that are analogs of mushroom-derived illudin toxins. Preclinical studies and clinical trials have demonstrated that irofulven is effective against several tumor cell types. The exact nature of irofulven-induced DNA damage is not completely understood. We demonstrated previously that irofulven activates ATM and its targets, NBS1, SMC1, CHK2, and p53. In this study, we hypothesize that irofulven induces DNA double-strand breaks and that BRCA1 may affect chemosensitivity by controlling cell cycle checkpoints, DNA repair, and genomic stability in response to irofulven treatment. We observed that irofulven induces the formation of chromosome breaks and radials and the activation and foci formation of gamma-H2AX, BRCA1, and RAD51. We also provided evidence that irofulven induces the generation of DNA double-strand breaks. By using BRCA1-deficient or -proficient cells, we demonstrated that in response to irofulven, BRCA1 contributes to the control of S and G(2)/M cell cycle arrest and is critical for repairing DNA double-strand breaks and for RAD51-dependent homologous recombination. Furthermore, we found that BRCA1 deficiency results in increased chromosome damage and chemosensitivity after irofulven treatment.
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PMID:BRCA1 contributes to cell cycle arrest and chemoresistance in response to the anticancer agent irofulven. 1722 70

Ku70 is one component of a protein complex, the Ku70/Ku80 heterodimer, which binds to DNA double-strand breaks and activates DNA-dependent protein kinase (DNA-PK), leading to DNA damage repair. Our previous work has confirmed that Ku70 is important for DNA damage repair in that Ku70 deficiency compromises the ability of cells to repair DNA double-strand breaks, increases the radiosensitivity of cells, and enhances radiation-induced apoptosis. Because of the radioresistance of some human cancers, particularly glioblastoma, we examined the use of a radio-gene therapy paradigm to sensitize cells to ionizing radiation. Based on the analysis of the structure-function of Ku70 and the crystal structure of Ku70/Ku80 heterodimer, we designed and identified a candidate dominant negative fragment involving an NH(2)-terminal deletion, and designated it as DNKu70. We generated this mutant construct, stably overexpressed it in Rat-1 cells, and showed that it has a dominant negative effect (i.e., DNKu70 overexpression results in decreased Ku-DNA end-binding activity, and increases radiosensitivity). We then constructed and generated recombinant replication-defective adenovirus, with DNKu70 controlled by the cytomegalovirus promoter, and infected human glioma U-87 MG cells and human colorectal tumor HCT-8 cells. We show that the infected cells significantly express DNKu70 and are greatly radiosensitized under both aerobic and hypoxic conditions. The functional ramification of DNKu70 was further shown in vivo: expression of DNKu70 inhibits radiation-induced DNA-PK catalytic subunit autophosphorylation and prolongs the persistence of gamma-H2AX foci. If radiation-resistant tumor cells could be sensitized by down-regulating the cellular level/activity of Ku/DNA-PK, this approach could be evaluated as an adjuvant to radiation therapy.
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PMID:Adenovirus-mediated expression of a dominant negative Ku70 fragment radiosensitizes human tumor cells under aerobic and hypoxic conditions. 1723 73

Here we show that the human BubR1 and MAD2 genes, which encode inhibitors of the anaphase promoting complex (APC/C), are directly activated by the oncogenic transcription factor c-MYC via E-box sequences in their first introns. In colorectal cancer biopsies elevated expression of c-MYC correlated with increased MAD2 levels. Activation of a conditional c-MYC allele delayed progression through mitosis in pro-metaphase in a MAD2- and BubR1-dependent manner. A fraction of the daughter cells derived from extended mitotic events underwent synchronous apoptosis, which was in part mediated by BubR1. Furthermore, c-MYC activation resulted in CIN (chromosomal instability) in the diploid MIN (microsatellite instability) cell line DLD-1 and further enhanced CIN in the aneuploid CIN-line MCF7. Unexpectedly, c-MYC-induced CIN was independent of c-MYC-induced BubR1/MAD2 expression and mitotic delay. Therefore, c-MYC-induced CIN may be caused be alternative pathways. We observed that activation of c-MYC induced DNA double-strand breaks, as evidenced by formation of gamma-H2AX foci, which colocalized with foci of active DNA replication. Furthermore, c-MYC activation resulted in mitotic chromosomes exhibiting DNA damage. Therefore, oncogenic deregulation of c-MYC prevents repair of replication-stress induced DNA lesions in the G(2)-phase. We suggest that the c-MYC-mediated persistence of DNA lesions throughout mitosis leads to chromosomal missegregation and underlies c-MYC-induced CIN. The effects of deregulated c-MYC on progression through mitosis described here may have important implications for the origin of chromosomal instability in many tumor types and the sensitivity towards cancer therapeutic agents targeting DNA or the mitotic spindle.
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PMID:c-MYC delays prometaphase by direct transactivation of MAD2 and BubR1: identification of mechanisms underlying c-MYC-induced DNA damage and chromosomal instability. 1729 7

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract and are caused by activating mutations of the KIT or platelet-derived growth factor receptor alpha (PDGFRA) tyrosine kinases. GISTs can be successfully treated with imatinib mesylate, a selective small-molecule protein kinase inhibitor that was first clinically approved to target the oncogenic BCR-ABL fusion protein kinase in chronic myelogenous leukemia, but which also potently inhibits KIT and PDGFR family members. The mechanistic events by which KIT/PDGFRA kinase inhibition leads to clinical responses in GIST patients are not known in detail. We report here that imatinib triggers GIST cell apoptosis in part through the up-regulation of soluble histone H2AX, a core histone H2A variant. We found that untreated GIST cells down-regulate H2AX in a pathway that involves KIT, phosphoinositide-3-kinase, and the ubiquitin/proteasome machinery, and that the imatinib-mediated H2AX up-regulation correlates with imatinib sensitivity. Depletion of H2AX attenuated the apoptotic response of GIST cells to imatinib. Soluble H2AX was found to sensitize GIST cells to apoptosis by aberrant chromatin aggregation and a transcriptional block. Our results underscore the importance of H2AX as a human tumor suppressor protein, provide mechanistic insights into imatinib-induced tumor cell apoptosis and establish H2AX as a novel target in cancer therapy.
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PMID:Histone H2AX is a mediator of gastrointestinal stromal tumor cell apoptosis following treatment with imatinib mesylate. 1736 89

Expression of the PTEN tumor suppressor gene is abnormal in many human cancers. Loss of PTEN expression leads to the activation of downstream signaling pathways that have been associated with resistance to radiation. In non-small cell lung carcinoma (NSCLC), suppressed expression of PTEN is frequently due to methylation of its promoter region. In this study, we tested whether gene transfer of wild-type PTEN into an NSCLC cell line with a known methylated PTEN promoter, H1299, would increase its sensitivity to ionizing radiation. Pretreating H1299 cells with an adenoviral-mediated PTEN (Ad-PTEN)-expressing vector sensitized H1299 cells to radiation. To determine the mechanism responsible for radiosensitization, we first examined radiation-induced apoptosis, which was enhanced but did not correlate with radiosensitizing effect of Ad-PTEN. Therefore, we next examined the ability of Ad-PTEN to modulate the repair of radiation-induced DNA double-strand breaks (DSBs) using the detection of repair foci positive for gamma-H2AX, a protein that becomes evident at the sites of each DSB and that can be visualized by immunofluorescent staining. Compared with controls, the repair of radiation-induced DSBs was retarded in H1299 cells pretreated with Ad-PTEN, consistent with the radiosensitizing effect of the vector. We conclude that signal transduction pathways residing primarily in the cytoplasm may intersect with DNA damage and repair pathways in the nucleus to modulate cellular responses to radiation. Elucidating the mechanisms responsible for this intersection may lead to novel strategies for improving therapy for cancers with defective PTEN.
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PMID:Adenoviral-mediated PTEN expression radiosensitizes non-small cell lung cancer cells by suppressing DNA repair capacity. 1743 3

SV40 virus has emerged as a potential cofactor with asbestos in the development of diffuse malignant mesothelioma, but its precise role in the pathogenesis of this tumor is unclear. SV40 large T antigen is known to inactivate cellular proteins involved in DNA damage and senescence, including p53 and pRb. We hypothesize that SV40 oncoproteins will sensitize mesothelial cells to DNA damage induced by asbestos or chemotherapeutic agents. SV40 oncoprotein expression in murine mesothelial cell lines enhanced spontaneous and asbestos-induced double-strand breaks, indicated by gamma-H2AX foci, and potentiated micronucleus formation. Mesothelial cells exposed to asbestos or bleomycin for 96 h acquired senescent-like morphology and displayed elevated senescence-associated beta-galactosidase activity, reduced bromodeoxyuridine (BrdUrd) incorporation, and reduced colony formation. SV40 oncoprotein expression abrogated the senescent phenotype, and transfected cell lines showed an increase in both BrdUrd incorporation and colony formation after prolonged DNA damage. Murine mesothelial cell lines lacking wild-type p53 due to a point mutation or gene rearrangement also failed to senesce in response to asbestos or chemotherapeutic agents. In addition, stress-induced senescence in human mesothelial cell lines was impaired by SV40 oncoprotein expression (MeT-5A), p53 small interfering RNA, or spontaneous p53 mutation (REN). These studies suggest that exposure to DNA-damaging agents can induce senescence in both murine and human mesothelioma cell lines and suggest a major, although not exclusive, role for p53 in this response. SV40 virus may contribute to mesothelioma progression by impairing stress-induced senescence, in part through p53 inactivation, thereby favoring survival and proliferation of mesothelial cells that have sustained DNA damage.
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PMID:SV40 oncoproteins enhance asbestos-induced DNA double-strand breaks and abrogate senescence in murine mesothelial cells. 1744 75


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