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)

Adenovirus infection activates cellular DNA damage response and repair pathways. Viral proteins that are synthesized before viral DNA replication prevent recognition of viral genomes as a substrate for DNA repair by targeting members of the sensor complex composed of Mre11/Rad50/NBS1 for degradation and relocalization, as well as targeting the effector protein DNA ligase IV. Despite inactivation of these cellular sensor and effector proteins, infection results in high levels of histone 2AX phosphorylation, or gammaH2AX. Although phosphorylated H2AX is a characteristic marker of double-stranded DNA breaks, this modification was widely distributed throughout the nucleus of infected cells and was coincident with the bulk of cellular DNA. H2AX phosphorylation occurred after the onset of viral DNA replication and after the degradation of Mre11. Experiments with inhibitors of the serine-threonine kinases ataxia telangiectasia mutated (ATM), AT- and Rad3-related (ATR), and DNA protein kinase (DNA-PK), the kinases responsible for H2AX phosphorylation, indicate that H2AX may be phosphorylated by ATR during a wild-type adenovirus infection, with some contribution from ATM and DNA-PK. Viral DNA replication appears to be the stimulus for this phosphorylation event, since infection with a nonreplicating virus did not elicit phosphorylation of H2AX. Infected cells also responded to high levels of input viral DNA by localized phosphorylation of H2AX. These results are consistent with a model in which adenovirus-infected cells sense and respond to both incoming viral DNA and viral DNA replication.
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PMID:Widespread phosphorylation of histone H2AX by species C adenovirus infection requires viral DNA replication. 1932 13

Long-term neurological deficiencies resulting from hippocampal cytotoxicity induced by cranial irradiation (IR) present a challenge in the treatment of primary and metastatic brain cancers, especially in children. Previously, we showed that lithium protected hippocampal neurons from IR-induced apoptosis and improved neurocognitive function in treated mice. Here, we demonstrate accelerated repair of IR-induced chromosomal double-strand breaks (DSBs) in lithium-treated neurons. Lithium treatment not only increased IR-induced DNA-dependent protein kinase (DNA-PK) threonine 2609 foci, a surrogate marker for activated nonhomologous end-joining (NHEJ) repair, but also enhanced double-strand DNA end-rejoining activity in hippocampal neurons. The increased NHEJ repair coincided with reduced numbers of IR-induced gamma-H2AX foci, well-characterized in situ markers of DSBs. These findings were confirmed in vivo in irradiated mice. Consistent with a role of NHEJ repair in lithium-mediated neuroprotection, attenuation of IR-induced apoptosis of hippocampal neurons by lithium was dramatically abrogated when DNA-PK function was abolished genetically in SCID mice or inhibited biochemically by the DNA-PK inhibitor IC86621. Importantly, none of these findings were evident in glioma cancer cells. These results support our hypothesis that lithium protects hippocampal neurons by promoting the NHEJ repair-mediated DNA repair pathway and warrant future investigation of lithium-mediated neuroprotection during cranial IR, especially in the pediatric population.
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PMID:Lithium-mediated protection of hippocampal cells involves enhancement of DNA-PK-dependent repair in mice. 1942 67

Understanding the molecular details associated with aberrant high mobility group A2 (HMGA2) gene expression is key to establishing the mechanism(s) underlying its oncogenic potential and effect on the development of therapeutic strategies. Here, we report the involvement of HMGA2 in impairing DNA-dependent protein kinase (DNA-PK) during the nonhomologous end joining (NHEJ) process. We showed that HMGA2-expressing cells displayed deficiency in overall and precise DNA end-joining repair and accumulated more endogenous DNA damage. Proper and timely activation of DNA-PK, consisting of Ku70, Ku80, and DNA-PKcs subunits, is essential for the repair of DNA double strand breaks (DSB) generated endogenously or by exposure to genotoxins. In cells overexpressing HMGA2, accumulation of histone 2A variant X phosphorylation at Ser-139 (gamma-H2AX) was associated with hyperphosphorylation of DNA-PKcs at Thr-2609 and Ser-2056 before and after the induction of DSBs. Also, the steady-state complex of Ku and DNA ends was altered by HMGA2. Microirradiation and real-time imaging in living cells revealed that HMGA2 delayed the release of DNA-PKcs from DSB sites, similar to observations found in DNA-PKcs mutants. Moreover, HMGA2 alone was sufficient to induce chromosomal aberrations, a hallmark of deficiency in NHEJ-mediated DNA repair. In summary, a novel role for HMGA2 to interfere with NHEJ processes was uncovered, implicating HMGA2 in the promotion of genome instability and tumorigenesis.
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PMID:Suppression of nonhomologous end joining repair by overexpression of HMGA2. 2014 17

Oxaliplatin, a chemotherapeutic drug, induces DNA double-strand breaks (DSBs) and apoptosis in colorectal cancer cells. It has been shown that gamma-H2AX acts as a marker of DSBs. However, the molecular events associated with oxaliplatin-mediated cell cycle arrest and cell death remain unclear. In this study, we investigated the roles of p53 and gamma-H2AX following oxaliplatin treatment, as they are important effector proteins for apoptosis and DSB repair, respectively. Both phosphorylated-p53 (Ser-15) and gamma-H2AX were up-regulated and accumulated in the nuclei of p53-wild type human colorectal cancer HCT116 cells after exposure to oxaliplatin. Concomitantly, oxaliplatin-induced G2/M arrest was associated with a reduction in both cyclin B1 expression and phosphorylated-CDC2 (Thr-161). Release of G2/M arrest by caffeine was accompanied by a decrease in the levels of p53/p21; however, gamma-H2AX levels were unchanged. Furthermore, inhibition of p53 phosphorylation by pifithrin-alpha was sufficient to reduce the oxaliplatin-induced up-regulation of gamma-H2AX and apoptosis. Oxaliplatin-induced gamma-H2AX via a p53-independent pathway but did not cause caspase-3 activation in p53-null HCT116 cells. Interestingly, no changes were observed in the H2AX gene knockdown with regards to oxaliplatin-induced G2/M arrest in p53-wild type and S phase arrest in p53-null HCT116 cells. Taken together, these data indicate that a molecular pathway involving p53, gamma-H2AX and cell cycle arrest plays a pivotal role in the cellular response to oxaliplatin.
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PMID:Oxaliplatin-induced gamma-H2AX activation via both p53-dependent and -independent pathways but is not associated with cell cycle arrest in human colorectal cancer cells. 1973 49

The H2AX core histone variant is phosphorylated in chromatin around DNA double strand breaks (DSBs) and functions through unknown mechanisms to suppress antigen receptor locus translocations during V(D)J recombination. Formation of chromosomal coding joins and suppression of translocations involves the ataxia telangiectasia mutated and DNA-dependent protein kinase catalytic subunit serine/threonine kinases, each of which phosphorylates H2AX along cleaved antigen receptor loci. Using Abelson transformed pre-B cell lines, we find that H2AX is not required for coding join formation within chromosomal V(D)J recombination substrates. Yet we show that H2AX is phosphorylated along cleaved Igkappa DNA strands and prevents their separation in G1 phase cells and their progression into chromosome breaks and translocations after cellular proliferation. We also show that H2AX prevents chromosome breaks emanating from unrepaired RAG endonuclease-generated TCR-alpha/delta locus coding ends in primary thymocytes. Our data indicate that histone H2AX suppresses translocations during V(D)J recombination by creating chromatin modifications that stabilize disrupted antigen receptor locus DNA strands to prevent their irreversible dissociation. We propose that such H2AX-dependent mechanisms could function at additional chromosomal locations to facilitate the joining of DNA ends generated by other types of DSBs.
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PMID:Histone H2AX stabilizes broken DNA strands to suppress chromosome breaks and translocations during V(D)J recombination. 1988 94

It has been reported that exposure to UV light triggers DNA damage response (DDR) seen as induction of gammaH2AX not only in S- but also in G(1)-phase cells. In the present study, in addition to gammaH2AX, we assessed other markers of DDR, namely phosphorylation of ATM on Ser1981, of ATM/ATR substrate on Ser/Thr at SQ/TQ cluster domains and of the tumor suppressor p53 on Ser15, in human pulmonary carcinoma A549 cells irradiated with 50 J/m(2) of UV-B. Phosphorylation of these proteins detected with phospho-specific Abs and measured by laser scanning cytometry in relation the cell cycle phase was found to be selective to S-phase cells. The kinetics of phosphorylation of ATM was strikingly similar to that of ATM/ATR substrate, peaking at 30 min after UV irradiation and followed by rapid dephosphorylation. The peak of H2AX phosphorylation was seen at 2 h and the peak of p53 phosphorylation at 4 h after exposure to UV light. Local high spatial density of these phospho-proteins reported by intensity of maximal pixel of immunofluorescence in the DDR nuclear foci was distinctly more pronounced in the early compared to late portion of S-phase. Exposure of cells to UV following 1 h pulse-labeling of their DNA with 5-ethynyl-2'deoxyuridine (EdU) made it possible to correlate the extent of DNA replication during the pulse with the extent of the UV-induced H2AX phosphorylation within the same cells. This correlation was very strong (R(2) = 0.98) and the cells that did not incorporate EdU showed no evidence of H2AX phosphorylation. The data are consistent with the mechanism in which stalling of DNA replication forks upon collision with the primary UV-induced DNA lesions and likely formation of double-strand DNA breaks triggers DDR. The prior reports (including our own) on induction of gammaH2AX in G(1) cells by UV may have erroneously identified cells initiating DNA replication following UV exposure as G(1) cells due to the fact that their DNA content did not significantly differ from that of G(1) cells that had not initiated DNA replication.
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PMID:Kinetics of the UV-induced DNA damage response in relation to cell cycle phase. Correlation with DNA replication. 2001 10

Recently, there have been many reports concerning proteins which can recognize DNA double strand break (DSBs), and such proteins include histone H2AX phosphorylated at serine 139 (gammaH2AX), ataxia telangiectasia mutated (ATM) phospho-serine 1981, DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phospho-threonine 2609, Nijmegen breakage syndrome 1 (NBS1) phospho-serine 343, checkpoint kinase 2 (CHK2), phospho-threonine 68, and structural maintenance of chromosomes 1 (SMC1) phospho-serine 966. Thus, it should be possible to follow the formation of DSBs and their repair using immunohistochemical methods with multiple antibodies to detect these proteins. When normal human fibroblasts (AG1522 cells) were exposed to 3 Gy of X-rays as a control, clearly discernable foci for these proteins were detected, and these foci localized with gammaH2AX foci. After heat treatment at 45.5 degrees C for 20 min, these proteins are partially localized with gammaH2AX foci. Here we show that there were slight differences in the localization pattern among these proteins, such as a disappearance from the nucleus (phospho-ATM) and translocation to the cytoplasm (phospho-NBS1) at 30 min after heat treatment, and some foci (phospho-DNA-PKcs and phospho-CHK2) appeared at 8 h after heat treatment. These results are discussed from perspectives of heat-induced denaturation of proteins and formation of DSBs.
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PMID:The foci of DNA double strand break-recognition proteins localize with gammaH2AX after heat treatment. 2017 16

Phosphorylation of the C-terminal end of histone H2A.X is the most characterized histone post-translational modification in DNA double-stranded breaks (DSB). DNA-dependent protein kinase (DNA-PK) is one of the three phosphatidylinositol 3 kinase-like family of kinase members that is known to phosphorylate histone H2A.X during DNA DSB repair. There is a growing body of evidence supporting a role for histone acetylation in DNA DSB repair, but the mechanism or the causative relation remains largely unknown. Using bacterially expressed recombinant mutants and stably and transiently transfected cell lines, we find that DNA-PK can phosphorylate Thr-136 in addition to Ser-139 both in vitro and in vivo. Furthermore, the phosphorylation reaction is not inhibited by the presence of H1, which in itself is a substrate of the reaction. We also show that, in contrast to previous reports, the ability of the enzyme to phosphorylate these residues is not affected by the extent of acetylation of the core histones. In vitro assembled nucleosomes and HeLa S3 native oligonucleosomes consisting of non-acetylated and acetylated histones are equally phosphorylated by DNA-PK. We demonstrate that the apparent differences in the extent of phosphorylation previously observed can be accounted for by the differential chromatin solubility under the MgCl(2) concentrations required for the phosphorylation reaction in vitro. Finally, we show that although H2A.X does not affect nucleosome conformation, it has a de-stabilizing effect that is enhanced by the DNA-PK-mediated phosphorylation and results in an impaired histone H1 binding.
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PMID:Phosphorylation of histone H2A.X by DNA-dependent protein kinase is not affected by core histone acetylation, but it alters nucleosome stability and histone H1 binding. 2035 35

The fact that eukaryotic DNA is packed into chromatin constitutes a physical barrier to enzymes and regulatory factors to reach the DNA molecule for replication, transcription, recombination and repair. Although most studies in this field have concentrated on how chromatin regulates transcription, there is a recent emphasis on studying the role of chromatin in the response to DNA damage. Two main chromatin-remodeling mechanisms have been identified, namely, ATP-dependent chromatin-remodeling complexes and histone post-translational modifications (PTMs). PTMs constitute reversible covalent modifications in aminoacidic residues, such as serine and threonine phosphorylation, lysine acetylation, lysine and arginine methylation and lysine ubiquitylation, among others. Moreover, nucleosome composition can be modified by the incorporation of histone variants, which are assembled into nucleosomes independently of DNA replication. The phosphorylation of the histone variant H2AX (gammaH2AX) is one of the best examples of histone PTMs in response to DNA damage induction, but many others have recently been revealed. In this review, we focus on and summarize the best-known histone PTMs observed in excision repair (base excision and nucleotide excision) and double-strand break (non-homologous end-joining and homologous recombination) repair pathways. In brief, the interplay between chromatin remodelers and DNA repair factors is discussed in relation to DNA damage response mechanisms.
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PMID:Histone post-translational modifications in DNA damage response. 2040 19

The checkpoint kinase Chk2 is an effector component of the ATM-dependent DNA damage response (DDR) pathway. The activation of Chk2 by genotoxic stress involves its phosphorylation on T68 by ATM and additional auto/transphosphorylations. Here we demonstrate that in unperturbed cells, chemical inhibition of Chk2 by VRX0466617 (VRX) enhances the phosphorylation of Chk2-T68 throughout the cell cycle phases. This event, dependent on the presence of ATM and catalytically functional Chk2, is not consequential to DNA damage, as neither gamma-H2AX nuclear foci nor increased ATM activation is detected in VRX-treated cells, suggesting the involvement of other regulatory proteins. As serine/threonine protein phosphatases (PPs) regulate the phosphorylation and deactivation of proteins of the DDR pathway, we analyzed their role in phospho-T68-Chk2 regulation. We found that intracellular inhibition of PP1 and PP2A-like activities by okadaic acid markedly raised the accumulation of Chk2-pT68 without DNA damage induction, and this phenomenon was also seen when PP1-C, PP2A-C, and Wip1/PPM1D were simultaneously knockdown by siRNA. Altogether, these data indicate a novel mechanism in undamaged cells where PPs function to maintain the balance between ATM and its direct substrate Chk2 through a regulatory circuit.
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PMID:A protein phosphatase feedback mechanism regulates the basal phosphorylation of Chk2 kinase in the absence of DNA damage. 2059 67


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