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The mechanisms leading to brain tumor formation are poorly understood. Using Ptch1+/? mice as a medulloblastoma model, sequential mutations were found to shape tumor evolution. Initially, medulloblastoma preneoplastic lesions display loss of heterozygosity of the Ptch1 wild-type allele, an event associated with cell senescence in preneoplasia. Subsequently, p53 mutations lead to senescence evasion and progression from preneoplasia to medulloblastoma. These findings are consistent with a model where high levels of Hedgehog signaling caused by the loss of the tumor suppressor Ptch1 lead to oncogene-induced senescence and drive p53 mutations. Thus, cell senescence is an important characteristic of a subset of SHH medulloblastoma and might explain the acquisition of somatic TP53 mutations in human medulloblastoma. This mode of medulloblastoma formation contrasts with the one characterizing Li-Fraumeni patients with medulloblastoma, where TP53 germ-line mutations cause chromothriptic genomic instability and lead to mutations in Hedgehog signaling genes, which drive medulloblastoma growth. Here we discuss in detail these 2 alternative mechanisms leading to medulloblastoma tumorigenesis. 相似文献
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Haemi Lee Il-taeg Cho Chang-Hun Lee 《Biochemical and biophysical research communications》2009,378(1):32-329
53BP1 is phosphorylated by the protein kinase ATM upon DNA damage. Even though several ATM phosphorylation sites in 53BP1 have been reported, those sites have little functional implications in the DNA damage response. Here, we show that ATM phosphorylates the S1219 residue of 53BP1 in vitro and that the residue is phosphorylated in cells exposed to ionizing radiation (IR). Transfection with siRNA targeting ATM abolished IR-induced phosphorylation at this residue, supporting the theory that this process is mediated by the kinase. To determine the functional relevance of this phosphorylation event, a U2OS cell line expressing S1219A mutant 53BP1 was established. IR-induced foci formation of MDC1 and γH2AX, DNA damage signaling molecules, was reduced in this cell line, implying that S1219 phosphorylation is required for recruitment of these molecules to DNA damage sites. Furthermore, overexpression of the mutant protein impeded IR-induced G2 arrest. In conclusion, we have shown that S1219 phosphorylation by ATM is required for proper execution of DNA damage response. 相似文献
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Ian Gibbs‐Seymour Ewa Markiewicz Simon Bekker‐Jensen Niels Mailand Christopher J. Hutchison 《Aging cell》2015,14(2):162-169
Lamins A/C have been implicated in DNA damage response pathways. We show that the DNA repair protein 53BP1 is a lamin A/C binding protein. In undamaged human dermal fibroblasts (HDF), 53BP1 is a nucleoskeleton protein. 53BP1 binds to lamins A/C via its Tudor domain, and this is abrogated by DNA damage. Lamins A/C regulate 53BP1 levels and consequently lamin A/C‐null HDF display a 53BP1 null‐like phenotype. Our data favour a model in which lamins A/C maintain a nucleoplasmic pool of 53BP1 in order to facilitate its rapid recruitment to sites of DNA damage and could explain why an absence of lamin A/C accelerates aging. 相似文献
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BRCA1 is an important mediator of the DNA damage response pathway. Previous studies have identified a number of proteins that associate with BRCA1 at nuclear foci after ionizing radiation (IR)-induced DNA damage. However, the co-localization patterns of BRCA1 and various DNA damage response proteins have not yet been systematically quantified and compared within the same experimental system. In this study, a new inducible human cell line was established to allow unambiguous detection of YFP–BRCA1 at nuclear foci. Quantitative 2-D microscopic analysis was performed to compare the intranuclear co-localization of YFP–BRCA1 with 10 cellular proteins and 4 cellular domains before and after IR. Intriguingly, YFP–BRCA1 displayed significantly better focal co-localization with BARD1, RAP80 and Abraxas than with the upstream foci-initiating proteins γH2AX and MDC1. In contrast to previous reports, we found that the co-localization between YFP–BRCA1 and 53BP1 foci was surprisingly weak. Quantitative analyses of 3-D confocal images showed that ~ 60% of 53BP1 foci were unrelated to YFP–BRCA1 foci, ~ 35% of foci were abutting and only ~ 5% of foci co-localized. The YFP–BRCA1 and 53BP1 nuclear foci were distinctively separated within the first 3 h after IR. In addition, in situ nuclear retention analysis revealed YFP–BRCA1 and BARD1 are less mobile than 53BP1 at IR-induced nuclear foci. Our findings indicate that BRCA1–BARD1 and 53BP1 are proximal but not overlapping at DNA break sites and are consistent with recent evidence for distinct roles of these proteins in the DNA damage response pathway. 相似文献
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利用GFP示踪细胞内源性P53活性检测DNA损伤 总被引:1,自引:1,他引:1
DNA损伤的检测对预防癌症和遗传病等非常重要。采用分子克隆技术,将报告基因—绿色荧光蛋白(GFP)置于SV40基本启动子调控下,构建成对照载体pSV-GFP。在SV40基本启动子上游插入寡核苷酸P53RE,构建成示踪载体p53RE-GFP。转染NIH3T3细胞,以GFP示踪细胞内源性P53的转录激活活性。紫外线照射或H2O2处理转化细胞使DNA损伤,诱导细胞内源性P53的表达。用激光扫描共聚焦成像系统(LSCIS)对细胞进行红、绿、蓝三色光融合成像,并测定GFP经488nm激发后发出的绿色荧光光密度,验证GFP示踪P53的特异性。p53RE-GFP转化细胞3T3-REG经紫外线照射或H2O2处理后,GFP的表达增高,处理后1hr光密度即达到最高水平,随后逐渐降低。血清“饥饿”—非DNA损伤处理的3T3-REG细胞,以及经紫外和H2O2处理的对照载体pSV-GFP转化细胞3T3-SVG,GFP的表达无明显增强。实验表明:GFP示踪内源性P53转录激活活性用于检测DNA损伤有很高的灵敏度和特异性,适宜推广应用。 相似文献
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The radiation-induced lesions which trigger the bystander effect 总被引:7,自引:0,他引:7
Ward JF 《Mutation research》2002,499(2):151-154
The ionizing radiation induced bystander effect is initiated by damage to a cellular molecule which then gives rise to a signal exported to other cells. The nature of this damage is considered with the understanding that it may not be the same as that responsible for the traditional cellular effects of radiation. Consideration is give to amounts of endogenous damage and to radiation yields of the various candidate lesions. 相似文献
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Chunfang Wang Diana Jurk Mandy Maddick Glyn Nelson Carmen Martin-Ruiz Thomas von Zglinicki 《Aging cell》2009,8(3):311-323
The impact of cellular senescence onto aging of organisms is not fully clear, not at least because of the scarcity of reliable data on the mere frequency of senescent cells in aging tissues. Activation of a DNA damage response including formation of DNA damage foci containing activated H2A.X (γ-H2A.X) at either uncapped telomeres or persistent DNA strand breaks is the major trigger of cell senescence. Therefore, γ-H2A.X immunohistochemistry (IHC) was established by us as a reliable quantitative indicator of senescence in fibroblasts in vitro and in hepatocytes in vivo and the age dependency of DNA damage foci accumulation in ten organs of C57Bl6 mice was analysed over an age range from 12 to 42 months. There were significant increases with age in the frequency of foci-containing cells in lung, spleen, dermis, liver and gut epithelium. In liver, foci-positive cells were preferentially found in the centrilobular area, which is exposed to higher levels of oxidative stress. Foci formation in the intestine was restricted to the crypts. It was not associated with either apoptosis or hyperproliferation. That telomeres shortened with age in both crypt and villus enterocytes, but telomeres in the crypt epithelium were longer than those in villi at all ages were confirmed by us. Still, there was no more than random co-localization between γ-H2A.X foci and telomeres even in crypts from very old mice, indicating that senescence in the crypt enterocytes is telomere independent. The results suggest that stress-dependent cell senescence could play a causal role for aging of mice. 相似文献
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Rachele Cescutti Simona Negrini Masaoki Kohzaki Thanos D Halazonetis 《The EMBO journal》2010,29(21):3723-3732
TopBP1 is a checkpoint protein that colocalizes with ATR at sites of DNA replication stress. In this study, we show that TopBP1 also colocalizes with 53BP1 at sites of DNA double‐strand breaks (DSBs), but only in the G1‐phase of the cell cycle. Recruitment of TopBP1 to sites of DNA replication stress was dependent on BRCT domains 1–2 and 7–8, whereas recruitment to sites of DNA DSBs was dependent on BRCT domains 1–2 and 4–5. The BRCT domains 4–5 interacted with 53BP1 and recruitment of TopBP1 to sites of DNA DSBs in G1 was dependent on 53BP1. As TopBP1 contains a domain important for ATR activation, we examined whether it contributes to the G1 cell cycle checkpoint. By monitoring the entry of irradiated G1 cells into S‐phase, we observed a checkpoint defect after siRNA‐mediated depletion of TopBP1, 53BP1 or ATM. Thus, TopBP1 may mediate the checkpoint function of 53BP1 in G1. 相似文献
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Stavroula Tsaridou Georgia Velimezi Frances Willenbrock Maria Chatzifrangkeskou Waheba Elsayed Andreas Panagopoulos Dimitris Karamitros Vassilis Gorgoulis Zoi Lygerou Vassilis Roukos Eric O'Neill Dafni Eleftheria Pefani 《EMBO reports》2022,23(8)
DNA lesions occur across the genome and constitute a threat to cell viability; however, damage at specific genomic loci has a relatively greater impact on overall genome stability. The ribosomal RNA gene repeats (rDNA) are emerging fragile sites. Recent progress in understanding how the rDNA damage response is organized has highlighted a key role of adaptor proteins. Here, we show that the scaffold tumor suppressor RASSF1A is recruited to rDNA breaks. RASSF1A recruitment to double‐strand breaks is mediated by 53BP1 and depends on RASSF1A phosphorylation at Serine 131 by ATM kinase. Employing targeted rDNA damage, we uncover that RASSF1A recruitment promotes local ATM signaling. RASSF1A silencing, a common epigenetic event during malignant transformation, results in persistent breaks, rDNA copy number alterations and decreased cell viability. Overall, we identify a novel role for RASSF1A at rDNA break sites, provide mechanistic insight into how the DNA damage response is organized in a chromatin context, and provide further evidence for how silencing of the RASSF1A tumor suppressor contributes to genome instability. 相似文献
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《DNA Repair》2019
Maintenance of genome integrity and stability is a critical responsibility of the DNA damage response (DDR) within cells, such that any disruption in this kinase-based signaling pathway leads to development of various disorders, particularly cancer. The tumor suppressor P53-binding protein 1 (53BP1), as one of the main mediators of DDR, plays a pivotal role in orchestrating the choice of double-strand break (DSB) repair pathway and contains interaction surfaces for numerous DSB-responsive proteins. It has been extensively demonstrated that aberrant expression of 53BP1 contributes to tumor occurrence and development. 53BP1 loss of function in tumor tissues is also related to tumor progression and poor prognosis in human malignancies. Due to undeniable importance of this protein in various aspects of cancer initiation/progression, angiogenesis, metastasis and development of drug resistance, as well as its targeting in the treatment of cancer, this review focused on explaining the structure and function of 53BP1 and its contribution to cancer. 相似文献
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Aleksandra Lezaja 《Cell cycle (Georgetown, Tex.)》2018,17(1):24-32
Replication stress is a major source of DNA damage and an important driver of cancer development. Replication intermediates that occur upon mild forms of replication stress frequently escape cell cycle checkpoints and can be transmitted through mitosis into the next cell cycle. The consequences of such inherited DNA lesions for cell fate and survival are poorly understood. By using time-lapse microscopy and quantitative image-based cytometry to simultaneously monitor inherited DNA lesions marked by the genome caretaker protein 53BP1 and cell cycle progression, we show that inheritance of 53BP1-marked lesions from the previous S-phase is associated with a prolonged G1 duration in the next cell cycle. These results suggest that cell-to-cell variation in S-phase commitment is determined, at least partially, by the amount of replication-born inherited DNA damage in individual cells. We further show that loss of the tumor suppressor protein p53 overrides replication stress-induced G1 prolongation and allows S-phase entry with excessive amounts of inherited DNA lesions. Thus, replication stress and p53 loss may synergize during cancer development by promoting cell cycle re-entry with unrepaired mutagenic DNA lesions originating from the previous cell cycle. 相似文献
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Jan Benada Kamila Burdová Tomá? Lidak Patrick von Morgen Libor Macurek 《Cell cycle (Georgetown, Tex.)》2015,14(2):219-231
In response to genotoxic stress, cells protect their genome integrity by activation of a conserved DNA damage response (DDR) pathway that coordinates DNA repair and progression through the cell cycle. Extensive modification of the chromatin flanking the DNA lesion by ATM kinase and RNF8/RNF168 ubiquitin ligases enables recruitment of various repair factors. Among them BRCA1 and 53BP1 are required for homologous recombination and non-homologous end joining, respectively. Whereas mechanisms of DDR are relatively well understood in interphase cells, comparatively less is known about organization of DDR during mitosis. Although ATM can be activated in mitotic cells, 53BP1 is not recruited to the chromatin until cells exit mitosis. Here we report mitotic phosphorylation of 53BP1 by Plk1 and Cdk1 that impairs the ability of 53BP1 to bind the ubiquitinated H2A and to properly localize to the sites of DNA damage. Phosphorylation of 53BP1 at S1618 occurs at kinetochores and in cytosol and is restricted to mitotic cells. Interaction between 53BP1 and Plk1 depends on the activity of Cdk1. We propose that activity of Cdk1 and Plk1 allows spatiotemporally controlled suppression of 53BP1 function during mitosis. 相似文献
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Makoto R. Hara Benjamin D. Sachs Marc G. Caron Robert J. Lefkowitz 《Cell cycle (Georgetown, Tex.)》2013,12(2):219-231
In response to genotoxic stress, cells protect their genome integrity by activation of a conserved DNA damage response (DDR) pathway that coordinates DNA repair and progression through the cell cycle. Extensive modification of the chromatin flanking the DNA lesion by ATM kinase and RNF8/RNF168 ubiquitin ligases enables recruitment of various repair factors. Among them BRCA1 and 53BP1 are required for homologous recombination and non-homologous end joining, respectively. Whereas mechanisms of DDR are relatively well understood in interphase cells, comparatively less is known about organization of DDR during mitosis. Although ATM can be activated in mitotic cells, 53BP1 is not recruited to the chromatin until cells exit mitosis. Here we report mitotic phosphorylation of 53BP1 by Plk1 and Cdk1 that impairs the ability of 53BP1 to bind the ubiquitinated H2A and to properly localize to the sites of DNA damage. Phosphorylation of 53BP1 at S1618 occurs at kinetochores and in cytosol and is restricted to mitotic cells. Interaction between 53BP1 and Plk1 depends on the activity of Cdk1. We propose that activity of Cdk1 and Plk1 allows spatiotemporally controlled suppression of 53BP1 function during mitosis. 相似文献
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TRF2 is a component of shelterin, the protein complex that protects the ends of mammalian chromosomes. TRF2 is essential for telomere capping owing to its roles in suppressing an ATM-dependent DNA damage response (DDR) at chromosome ends and inhibiting end-to-end chromosome fusions. Mice deficient for TRF2 are early embryonic lethal. However, the role of TRF2 in later stages of development and in the adult organism remains largely unaddressed, with the exception of liver, where TRF2 was found to be dispensable for maintaining tissue function. Here, we study the impact of TRF2 conditional deletion in stratified epithelia by generating the TRF2∆/∆-K5-Cre mouse model, which targets TRF2 deletion to the skin from embryonic day E11.5. In marked contrast to TRF2 deletion in the liver, TRF2∆/∆-K5-Cre mice show lethality in utero reaching 100% lethality perinataly. At the molecular and cellular level, TRF2 deletion provokes induction of an acute DDR at telomeres, leading to activation of p53 signaling pathways and to programed cell death since the time of Cre expression at E11.5. Unexpectedly, neither inhibition of the NHEJ pathway by abrogation of 53BP1 nor inhibition of DDR by p53 deficiency rescued these severe phenotypes. Instead, TRF2 deletion provokes an extensive epidermal cell death accompanied by severe inflammation already at E16.5 embryos, which are independent of p53. These results are in contrast with conditional deletion of TRF1 and TPP1 in the skin, where p53 deficiency rescued the associated skin phenotypes, highlighting the comparatively more essential role of TRF2 in skin homeostasis. 相似文献
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R-M Laberge D Adler M DeMaria N Mechtouf R Teachenor G B Cardin P-Y Desprez J Campisi F Rodier 《Cell death & disease》2013,4(7):e727
Senescence is a cellular response to damage and stress. The senescence response prevents cancer by suppressing the proliferation of cells with a compromised genome and contributes to optimal wound healing in normal tissues. Persistent senescent cells are also thought to drive aging and age-associated pathologies through their secretion of inflammatory factors that modify the tissue microenvironment and alter the function of nearby normal or transformed cells. Understanding how senescent cells alter the microenvironment would be aided by the ability to induce or eliminate senescent cells at will in vivo. Here, we combine the use of the synthetic nucleoside analog ganciclovir (GCV) with herpes simplex virus thymidine kinase (HSVtk) activity to create or eliminate senescent human cells. We show that low concentrations of GCV induce senescence through the accumulation of nuclear DNA damage while higher concentrations of GCV, similar to those used in vivo, kill non-dividing senescent cells via mitochondrial DNA (mtDNA) damage and caspase-dependent apoptosis. Using this system, we effectively eliminated xenografted normal human senescent fibroblasts or induced senescence in human breast cancer cells in vivo. Thus, cellular senescence and mtDNA damage are outcomes of synthetic nucleoside analog treatment, indicating that the GCV–HSVtk combination can be used effectively to promote the targeted formation or eradication of senescent cells. 相似文献