SEARCH · PubMed Health

Results for “Checkpoint Kinase 2”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 37 records · Page 2Linked to original sources

PubMed2006

Trans-activation of the DNA-damage signalling protein kinase Chk2 by T-loop exchange.

The protein kinase Chk2 (checkpoint kinase 2) is a major effector of the replication checkpoint. Chk2 activation is initiated by phosphorylation of Thr68, in the serine-glutamine/threonine-glutamine cluster domain (SCD), by ATM. The phosphorylated SCD-segment binds to the FHA domain of a second Chk2 molecule, promoting dimerisation of the protein and triggering phosphorylation of the activation segment/T-loop in the kinase domain. We have now determined the structure of the kinase domain of human Chk2 in complexes with ADP and a small-molecule inhibitor debromohymenialdisine. The structure reveals a remarkable dimeric arrangement in which T-loops are exchanged between protomers, to form an active kinase conformation in trans. Biochemical data suggest that this dimer is the biologically active state promoted by ATM-phosphorylation, and also suggests a mechanism for dimerisation-driven activation of Chk2 by trans-phosphorylation.

Adenosine Diphosphate↗

PubMed1999

Mammalian Chk2 is a downstream effector of the ATM-dependent DNA damage checkpoint pathway.

In response to DNA damage and replication blocks, cells activate pathways that arrest the cell cycle and induce the transcription of genes that facilitate repair. In mammals, ATM (ataxia telangiectasia mutated) kinase together with other checkpoint kinases are important components in this response. We have cloned the rat and human homologs of Saccharomyces cerevisiae Rad 53 and Schizosaccharomyces pombe Cds1, called checkpoint kinase 2 (chk2). Complementation studies suggest that Chk2 can partially replace the function of the defective checkpoint kinase in the Cds1 deficient yeast strain. Chk2 was phosphorylated and activated in response to DNA damage in an ATM dependent manner. Its activation in response to replication blocks by hydroxyurea (HU) treatment, however, was independent of ATM. Using mass spectrometry, we found that, similar to Chk1, Chk2 can phosphorylate serine 216 in Cdc25C, a site known to be involved in negative regulation of Cdc25C. These results suggest that Chk2 is a downstream effector of the ATM-dependent DNA damage checkpoint pathway. Activation of Chk2 might not only delay mitotic entry, but also increase the capacity of cultured cells to survive after treatment with gamma-radiation or with the topoisomerase-I inhibitor topotecan.

Alkylating Agents↗

PubMed2004

Aberrations of the Chk2 tumour suppressor in advanced urinary bladder cancer.

Checkpoint kinase 2 (Chk2) is a tumour suppressor and signal transducer in genome integrity checkpoints that coordinate cell-cycle progression with DNA repair or cell death in response to DNA damage. Defects of Chk2 occur in subsets of diverse sporadic malignancies and predispose to several types of hereditary carcinomas. However, the status of Chk2 in tumours of the urinary bladder remains unknown. Here, we report that among 58 advanced (grade T2-T4) human bladder carcinomas, immunohistochemical analysis revealed tumour-specific reduction or lack of Chk2 protein in 6 (10.3%) cases. Genetic analysis of the latter subset showed that a Chk2-negative carcinoma #668 harboured a truncating mutation 1100delC, in one Chk2 allele and loss of the corresponding second allele. The 1100delC mutation was also found in the germ line of this patient. Sequencing of TP53 in tumour #668 identified two missense mutations. Furthermore, the vast majority of the tumours showed 'unscheduled' activatory phosphorylation on Thr68 of Chk2 in the absence of any DNA-damaging treatment. Our results indicate that the otherwise dormant DNA damage signal transducer Chk2 is aberrantly and constitutively activated in invasive urinary bladder carcinomas, and that such likely proapoptotic checkpoint signalling can be disabled by inactivation of Chk2 and/or p53 tumour suppressors in subsets of these tumours.

Carcinoma↗

PubMed2006

Gallic acid causes inactivating phosphorylation of cdc25A/cdc25C-cdc2 via ATM-Chk2 activation, leading to cell cycle arrest, and induces apoptosis in human prostate carcinoma DU145 cells.

We recently reported that gallic acid is a major active agent responsible for grape seed extract activity in DU145 human prostate carcinoma cells. The present study was conducted to examine its efficacy and associated mechanism. Gallic acid treatment of DU145 cells resulted in a strong cell growth inhibition, cell cycle arrest, and apoptotic death in a dose- and time-dependent manner, together with a decrease in cyclin-dependent kinases and cyclins but strong induction in Cip1/p21. Additional mechanistic studies showed that gallic acid induces an early Tyr(15) phosphorylation of cell division cycle 2 (cdc2). Further upstream, gallic acid also induced phosphorylation of both cdc25A and cdc25C via ataxia telangiectasia mutated (ATM)-checkpoint kinase 2 (Chk2) activation as a DNA damage response evidenced by increased phospho-histone 2AX (H2A.X) that is phosphorylated by ATM in response to DNA damage. Time kinetics of ATM phosphorylation, together with those of H2A.X and Chk2, was in accordance with an inactivating phosphorylation of cdc25A and cdc25C phosphatases and cdc2 kinase, suggesting that gallic acid increases cdc25A/C-cdc2 phosphorylation and thereby inactivation via ATM-Chk2 pathway following DNA damage that induces cell cycle arrest. Caffeine, an ATM/ataxia telangiectasia-rad3-related inhibitor, reversed gallic acid-caused ATM and H2A.X phosphorylation and cell cycle arrest, supporting the role of ATM pathway in gallic acid-induced cell cycle arrest. Additionally, gallic acid caused caspase-9, caspase-3, and poly(ADP)ribose polymerase cleavage, but pan-caspase inhibitor did not reverse apoptosis, suggesting an additional caspase-independent apoptotic mechanism. Together, this is the first report identifying gallic acid efficacy and associated mechanisms in an advanced and androgen-independent human prostate carcinoma DU145 cells, suggesting future in vivo efficacy studies with this agent in preclinical prostate cancer models.

Apoptosis↗

PubMed2006

Chk2 mediates stabilization of the FoxM1 transcription factor to stimulate expression of DNA repair genes.

The forkhead box M1 (FoxM1) transcription factor regulates expression of cell cycle genes essential for DNA replication and mitosis during organ repair and cancer progression. Here, we demonstrate that FoxM1-deficient (-/-) mouse embryonic fibroblasts and osteosarcoma U2OS cells depleted in FoxM1 levels by small interfering RNA transfection display increased DNA breaks, as evidenced by immunofluorescence focus staining for phosphospecific histone H2AX. FoxM1-deficient cells also exhibit stimulation of p53 transcriptional activity, as evidenced by increased expression of the p21(cip1) gene. FoxM1-deficient cells display reduced expression of the base excision repair factor X-ray cross-complementing group 1 (XRCC1) and breast cancer-associated gene 2 (BRCA2), the latter of which is involved in homologous recombination repair of DNA double-strand breaks. Furthermore, FoxM1 protein is phosphorylated by checkpoint kinase 2 (Chk2) in response to DNA damage. This phosphorylation of FoxM1 on serine residue 361 caused increased stability of the FoxM1 protein with corresponding increased transcription of XRCC1 and BRCA2 genes, both of which are required for repair of DNA damage. These results identify a novel role for FoxM1 in the transcriptional response during DNA damage/checkpoint signaling and show a novel mechanism by which Chk2 protein regulates expression of DNA repair enzymes.

Amino Acid Sequence↗

PubMed2026

TRIM28 regulates the G2/M transition via histone modification and DNA damage repair during mouse oocyte meiosis.

TRIM28, a member of the tripartite motif (TRIM) family, functions as a transcriptional coregulator involved in maintaining genome stability during mitosis. In this study, we explored the role of TRIM28 in mouse oocyte meiotic maturation, where transcriptional activity is barely detectable. We found that TRIM28 was constitutively expressed during the early stages of oocyte meiotic maturation, with predominant nuclear localization in germinal vesicle (GV)-stage oocytes. TRIM28 depletion caused defective germinal vesicle breakdown (GVBD), but oocytes that successfully underwent GVBD displayed unimpaired first polar body (PB1) extrusion. TRIM28 depletion impaired CDK1 activity and reduced cyclin B1 levels, leading to a delay in the G2/M transition. This delay might be caused by altered levels of HDAC2-mediated H4K12ac and H3K4me2-modulated H3K9me2 in nonsurrounded nucleolus (NSN)-type GV oocytes, which decreased transcription activity. Additionally, TRIM28-depleted oocytes exhibited elevated γ-H2A.X expression, accompanied by aberrant expression of CHK1 and CHK2, as well as dysregulated expression of RAD51, which collectively contributed to GVBD failure in mouse oocytes. In conclusion, our findings indicate that TRIM28 participates in the regulation of the G2/M transition during mouse oocyte meiotic maturation, acting through the modulation of histone modifications and DNA damage repair.

Animals↗

PubMed2005

Gemcitabine-induced activation of checkpoint signaling pathways that affect tumor cell survival.

Two signaling pathways are activated by antineoplastic therapies that damage DNA and stall replication. In one pathway, double-strand breaks activate ataxia-telangiectasia mutated kinase (ATM) and checkpoint kinase 2 (Chk2), two protein kinases that regulate apoptosis, cell-cycle arrest, and DNA repair. In the second pathway, other types of DNA lesions and replication stress activate the Rad9-Hus1-Rad1 complex and the protein kinases ataxia-telangiectasia mutated and Rad3-related kinase (ATR) and checkpoint kinase 1 (Chk1), leading to changes that block cell-cycle progression, stabilize stalled replication forks, and influence DNA repair. Gemcitabine and cytarabine are two highly active chemotherapeutic agents that disrupt DNA replication. Here, we examine the roles these pathways play in tumor cell survival after treatment with these agents. Cells lacking Rad9, Chk1, or ATR were more sensitive to gemcitabine and cytarabine, consistent with the fact that these agents stall replication forks, and this sensitization was independent of p53 status. Interestingly, ATM depletion sensitized cells to gemcitabine and ionizing radiation but not cytarabine. Together, these results demonstrate that 1) gemcitabine triggers both checkpoint signaling pathways, 2) both pathways contribute to cell survival after gemcitabine-induced replication stress, and 3) although gemcitabine and cytarabine both stall replication forks, ATM plays differential roles in cell survival after treatment with these agents.

Ataxia Telangiectasia Mutated Proteins↗

PubMed2005

Reduced p53 in peripheral blood mononuclear cells from patients with rheumatoid arthritis is associated with loss of radiation-induced apoptosis.

OBJECTIVE: Patients with autoimmune disorders exhibit highly reproducible gene expression profiles in their peripheral blood mononuclear cells. This profile includes, at least in part, a collection of underexpressed genes that encode proteins that inhibit cell cycle progression and stimulate apoptosis. We aimed to determine whether this gene expression profile confers functional liability on lymphocytes from patients with rheumatoid arthritis (RA). METHODS: Viability studies in response to a panel of proapoptotic stimuli revealed that T lymphocytes from patients with RA were resistant to gamma radiation-induced apoptosis, a process known to be dependent on p53. To assess p53 function in RA peripheral blood mononuclear cells, baseline levels of p53 protein and TP53 transcript were measured in patients with RA and controls. The cellular p53 response to gamma radiation was also assessed by immunoblotting. RESULTS: Lymphocytes from patients with RA had lower baseline levels of TP53 messenger RNA (mRNA) and p53 protein than did those from control subjects and were deficient in their ability to increase p53 after exposure to gamma radiation. A subgroup of patients with RA had a second biochemical defect characterized by expression of very low baseline levels of checkpoint kinase 2 mRNA and protein. CONCLUSION: We conclude that defects in the expression of TP53 mRNA and, in a subgroup, defects in expression of CHK2 mRNA, lead to severe defects in apoptosis in patients with RA. We hypothesize that this liability may contribute to autoimmunity.

Adult↗

PubMed2003

The promyelocytic leukemia protein protects p53 from Mdm2-mediated inhibition and degradation.

The p53 protein is kept labile under normal conditions. This regulation is governed largely by its major negative regulator, Mdm2. In response to stress however, p53 accumulates and becomes activated. For this to occur, the inhibitory effects of Mdm2 have to be neutralized. Here we investigated the role of the promyelocytic leukemia protein (PML) in the activation of p53 in response to stress. We found that PML is critical for the accumulation of p53 in response to DNA damage under physiological conditions. PML protects p53 from Mdm2-mediated ubiquitination and degradation, and from inhibition of apoptosis. PML neutralizes the inhibitory effects of Mdm2 by prolonging the stress-induced phosphorylation of p53 on serine 20, a site of the checkpoint kinase 2 (Chk2). PML recruits Chk2 and p53 into the PML nuclear bodies and enhances p53/Chk2 interaction. Our results provide a novel mechanistic explanation for the cooperation between PML and p53 in response to DNA damage.

Active Transport, Cell Nucleus↗

PubMed2005

Dual induction of apoptosis and senescence in cancer cells by Chk2 activation: checkpoint activation as a strategy against cancer.

The human checkpoint kinase 2 (Chk2) plays a central role in regulation of the cellular response to DNA damage, resulting in cell cycle arrest, DNA repair, or apoptosis depending on severity of DNA damage and the cellular context. Chk2 inhibitors are being developed as sensitizers for chemotherapeutic agents. In contrast, here we report that direct activation of Chk2 alone (without chemotherapeutic agents) led to potent inhibition of cancer cell proliferation. In the absence of de novo DNA damage, checkpoint activation was achieved by increased Chk2 expression, as evidenced by its phosphorylation at Thr68, resulting in senescence and apoptosis of cancer cells (DLD1 and HeLa). The Chk2-induced apoptosis was p53 independent and was mediated by caspase activation triggered by loss of mitochondrial potential. The Chk2-induced senescence was also p53 independent and was associated with induction of p21. These results suggest that direct activation of checkpoint kinases may be a novel approach for cancer therapy.

Apoptosis↗

PubMed2004

Inhibition of nuclear factor-kappaB and nitric oxide by curcumin induces G2/M cell cycle arrest and apoptosis in human melanoma cells.

Curcumin (diferuloylmethane) inhibits tumour cell growth by inducing apoptosis in many tumour types, including melanoma, via complex and ill-defined pathways. Recent studies have shown that curcumin is both a nitric oxide scavenger and an inhibitor of inducible nitric oxide synthase (iNOS) expression, low levels of which correlate with antiapoptotic function and poor survival and which may be regulated by inhibition of nuclear factor-kappaB (NFkappaB) activation. To elucidate the mechanisms by which curcumin inhibits melanoma proliferation, we tested the in vitro effects of curcumin on specific cell cycle pathways and melanoma cell survival, including NFkappaB activation. Curcumin induced melanoma cell apoptosis and cell cycle arrest, which is associated with the downregulation of NFkappaB activation, iNOS and DNA-dependent protein kinase catalytic subunit expression, and upregulation of p53, p21(Cip1), p27(Kip1) and checkpoint kinase 2. Curcumin also downregulated constitutive iNOS activity in melanoma cells. Our results demonstrate that curcumin arrested cell growth at the G(2)/M phase and induced apoptosis in human melanoma cells by inhibiting NFkappaB activation and thus depletion of endogenous nitric oxide. Therefore, curcumin should be considered further as a potential therapy for patients with melanoma.

Active Transport, Cell Nucleus↗

PubMedDate not supplied

The Chk2 protein kinase.

Checkpoint kinase 2 (Chk2) is a multifunctional enzyme whose functions are central to the induction of cell cycle arrest and apoptosis by DNA damage. Insight into Chk2 has derived from multiple approaches. Biochemical studies have addressed Chk2 structure, domain organization and regulation by phosphorylation. Extensive work has been done to identify factors that recognize and respond to DNA damage in order to activate Chk2. In turn a number of substrates and targets of Chk2 have been identified that play roles in the checkpoint response. The roles and regulation of Chk2 have been elucidated by studies in model genetic systems extending from worms and flies to mice and humans. The relationship of Chk2 to human cancer studies is developing rapidly with increasing evidence that Chk2 plays a role in tumor suppression.

Animals↗

PubMed2006

Augmented cancer resistance and DNA damage response phenotypes in PPM1D null mice.

The p53-induced serine/threonine phosphatase, protein phosphatase 1D magnesium-dependent, delta isoform (PPM1D) (or wild-type p53-induced phosphatase 1 (Wip1)), exhibits oncogenic activity in vitro and in vivo. It behaves as an oncogene in rodent fibroblast transformation assays and is amplified and overexpressed in several human tumor types. It may contribute to oncogenesis through functional inactivation of p53. Here, we show that the oncogenic function of PPM1D is associated with its phosphatase activity. While overexpressed PPM1D may be oncogenic, PPM1D null mice are resistant to spontaneous tumors over their entire lifespan. This cancer resistance may be based in part on an augmented stress response following DNA damage. PPM1D null mice treated with ionizing radiation display increased p53 protein levels and increased phosphorylation of p38 MAP kinase, p53, checkpoint kinase 1 (Chk1), and checkpoint kinase 2 (Chk2) in their tissues compared to their wild-type (WT) counterparts. Male PPM1D null mice show a modest reduction in longevity, reduced serum insulin-like growth factor 1 (IGF-1) levels, and reduced body weight compared to WT mice. The PPM1D null mouse phenotypes indicate that PPM1D has a homeostatic role in abrogating the DNA damage response and may regulate aspects of male longevity.

Aging↗

PubMed2003

Chk2 activates E2F-1 in response to DNA damage.

The E2F-1 transcription factor is regulated during cell cycle progression and induced by cellular stress, such as DNA damage. We report that checkpoint kinase 2 (Chk2) regulates E2F-1 activity in response to the DNA-damaging agent etoposide. A Chk2 consensus phosphorylation site in E2F-1 is phosphorylated in response to DNA damage, resulting in protein stabilization, increased half-life, transcriptional activation and localization of phosphorylated E2F-1 to discrete nuclear structures. Expression of a dominant-negative Chk2 mutant blocks induction of E2F-1 and prevents E2F-1-dependent apoptosis. Moreover, E2F-1 is resistant to induction by etoposide in tumour cells expressing mutant chk2. Therefore, Chk2 phosphorylates and activates E2F-1 in response to DNA damage, resulting in apoptosis. These results suggest a role for E2F-1 in checkpoint control and provide a plausible explanation for the tumour suppressor activity of E2F-1.

Apoptosis↗

PubMed2004

Statins enhance migratory capacity by upregulation of the telomere repeat-binding factor TRF2 in endothelial progenitor cells.

BACKGROUND: Cultivation of endothelial progenitor cells (EPCs) leads to premature replicative senescence, limiting ex vivo expansion for potential clinical cell therapy. Recent studies have linked senescence to the dysfunction of telomeres, the "ends" of chromosomes, via the so-called mitotic clock or culture-induced stress. The purpose of this study was to elucidate a possible role of telomere biology in the functional augmentation of EPCs by statins. METHODS AND RESULTS: Human EPCs were isolated from peripheral blood. Using flow cytometry after fluorescence in situ hybridization with a telomere-specific (C3TA2)3 peptide nucleic acid probe (Flow-FISH), we found mean telomere length in untreated EPCs from healthy subjects to range between 8.5+/-0.2 and 11.1+/-0.5 kb with no change over 6 days of culture, excluding telomere erosion as one cause for premature senescence. Although mean telomere length did not differ between statin-treated and untreated EPCs, atorvastatin (0.1 micromol/L) and mevastatin (1.0 micromol/L) both led to a more than 3-fold increase in the expression of the telomere capping protein TRF2 (telomere repeat-binding factor), as shown by immunoblotting, whereas quantitative reverse transcription-polymerase chain reaction demonstrated no increase in TRF2 mRNA. Telomere dysfunction of EPCs was also paralleled by a 4-fold increase in the DNA damage checkpoint-kinase 2 (Chk2). Conversely, statin cotreatment or overexpression of TRF2 completely suppressed Chk2 induction. Finally, overexpression of a dominant negative mutant of the TRF2 protein abrogated statin-induced enhancement of migratory activity down to baseline values. CONCLUSIONS: Ex vivo culturing of EPCs leads to "uncapping" of telomeres, indicated by the loss of TRF2. Statin cotreatment of EPCs prevents impairment of their functional capacity by a TRF2-dependent, posttranscriptional mechanism. This is the first time a beneficial effect of statins on telomere biology has been described.

Adenoviridae↗

PubMed2005

Mutations in proline 82 of p53 impair its activation by Pin1 and Chk2 in response to DNA damage.

Tumor suppression by the p53 protein largely depends on the elimination of damaged cells by apoptosis. Mutations in the polyproline region (PPR) of p53 impair its apoptotic function. Deletion of the PPR renders p53 more sensitive to inhibition by Mdm2 via an unknown mechanism. We have explored the mechanism by which the PPR modulates the p53/Mdm2 loop. Proline 82 of p53 was identified to be essential for its interaction with the checkpoint kinase 2 (Chk2) and consequent phosphorylation of p53 on serine 20, following DNA damage. These physical and functional interactions are regulated by Pin1 through cis-trans isomerization of proline 82. Our study unravels the pathway by which Pin1 activates p53 in response to DNA damage and explains how Pin1 protects p53 from Mdm2. Further, we propose a role for Pin1-dependent induction of p53 conformational change as a mechanism responsible for the enhanced interaction between p53 and Chk2 following DNA damage. Importantly, our findings elucidate the selection for mutations in the Pin1 target Thr81/Pro82 motif within the PPR of p53 in human cancer.

Animals↗

PubMed2005

The CHEK2 1100delC allele is not relevant for risk assessment in HNPCC and HBCC Spanish families.

The frame-shift mutation 1100delC in the cell-cycle-checkpoint kinase 2 gene (CHEK2) has been reported to be a low penetrance breast cancer gene in Northern European populations. However, the variant may be relevant for breast cancer risk in other populations, due to its low prevalence. Recent studies have proposed a role for the mutation in colorectal cancer, finding a strong association between the CHEK21100delC mutation and hereditary breast and colorectal cancer (HBCC). A previous study suggested that the CHEK21100delC variant was not of clinical relevance in Spanish breast/ovarian cancer families. Here, we demonstrate that this genetic variant is not of clinical relevance for HNPCC and HBCC Spanish families.

Adult↗

PubMed2001

CHK2 kinase--a busy messenger.

Checkpoint kinase 2 (Chk2) is emerging as a key mediator of diverse cellular responses to genotoxic stress, guarding the integrity of the genome throughout eukaryotic evolution. Recent studies show the fundamental role of Chk2 in the network of genome-surveillance pathways that coordinate cell-cycle progression with DNA repair and cell survival or death. Defects in Chk2 contribute to the development of both hereditary and sporadic human cancers, and earmark this kinase as a candidate tumour suppressor and an attractive target for drug discovery.

Animals↗