RAD52 function is required for virtually all DNA double-strand break repair and recombination events in Saccharomyces cerevisiae. To gain greater insight into the mechanism of RAD52-mediated repair, we screened for genes that suppress partially active alleles of RAD52 when mutant or overexpressed. Described here is the isolation of a phenotypic null allele of SRS2 that suppressed multiple alleles of RAD52 (rad52B, rad52D, rad52-1 and KlRAD52) and RAD51 (KlRAD51) but failed to suppress either a rad52 delta or a rad51 delta. These results indicate that SRS2 antagonizes RAD51 and RAD52 function in recombinational repair. The mechanism of suppression of RAD52 alleles by srs2 is distinct from that which has been previously described for RAD51 overexpression, as both conditions were shown to act additively with respect to the rad52B allele. Furthermore, overexpression of either RAD52 or RAD51 enhanced the recombination-dependent sensitivity of an srs2 delta RAD52 strain, suggesting that RAD52 and ...

TY - JOUR. T1 - Attenuating the DNA damage response to double-strand breaks restores function in models of CNS neurodegeneration. AU - Tuxworth , Richard AU - Taylor, Matthew AU - Anduaga, Ane Martin AU - Hussien-Ali, Al. AU - Chatzimatthaiou, Sotiroula AU - Longland, Joanne AU - Thompson, Adam. AU - Almutiri, Sharif AU - Alifragis, Pavlos. AU - Kyriacou, Charalambos AU - Kysela, Boris AU - Ahmed, Zubair PY - 2019/7/2. Y1 - 2019/7/2. N2 - DNA double-strand breaks are a feature of many acute and long-term neurological disorders, including neurodegeneration, following neurotrauma and after stroke. Persistent activation of the DNA damage response in response to double-strand breaks contributes to neural dysfunction and pathology as it can force post-mitotic neurons to re-enter the cell cycle leading to senescence or apoptosis. Mature, non-dividing neurons may tolerate low levels of DNA damage, in which case muting the DNA damage response might be neuroprotective. Here, we show that attenuating the ...

Quercetin suppresses DNA double-strand break repair and enhances the radiosensitivity of human ovarian cancer cells via p53-dependent endoplasmic reticulum stress pathway Cheng Gong,1 Zongyuan Yang,1 Lingyun Zhang,2 Yuehua Wang,2 Wei Gong,2 Yi Liu3 1Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 2Department of Oncology, XiangYang Central Hospital, Hubei University of Arts and Science, XiangYang, 3Department of Medicinal Chemistry, School of Pharmacy, Hubei University of Chinese Medicine, Wuhan, China Abstract: Quercetin is proven to have anticancer effects for many cancers. However, the role of tumor suppressor p53 on quercetin’s radiosensitization and regulation of endoplasmic reticulum (ER) stress response in this process remains obscure. Here, quercetin exposure resulted in ER stress, prolonged DNA repair, and the expression of p53 protein; phosphorylation on serine 15 and 20 increased in combination with

DNA double strand break (DSB) repair enzymes are thought to be necessary for retroviral infection, especially for the post-integration repair and circularization of viral cDNA. However, the detailed roles of DSB repair enzymes in retroviral infection remain to be elucidated. A GFP reporter assay showed that the infectivity of an HIV-based vector decreased in ATM- and DNA-PKcs-deficient cells when compared with their complemented cells, while that of an MLV-based vector was diminished in Mre11- and DNA-PKcs-deficient cells. By using a method based on inverse- and Alu-PCR, we analyzed sequences around 3 HIV-1 integration sites in ATM-, Mre11- and NBS1- deficient cells. Increased abnormal junctions between the HIV-1 provirus and the host DNA were found in these mutant cell lines compared to the complemented cell lines and control MRC5SV cells. The abnormal junctions contained two types of insertions: 1) GT dinucleotides, which are normally removed by integrase during integration, and 2) inserted

TY - JOUR. T1 - DNA-damage-induced degradation of EXO1 exonuclease limits DNA end resection to ensure accurate DNA repair. AU - Tomimatsu, Nozomi. AU - Mukherjee, Bipasha. AU - Harris, Janelle Louise. AU - Boffo, Francesca Ludovica. AU - Hardebeck, Molly Catherine. AU - Potts, Patrick Ryan. AU - Khanna, Kum Kum. AU - Burma, Sandeep. N1 - Publisher Copyright: © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.. PY - 2017/6/30. Y1 - 2017/6/30. N2 - End resection of DNA double-strand breaks (DSBs) to generate 3′-single-stranded DNA facilitates DSB repair via error-free homologous recombination (HR) while stymieing repair by the error-prone non-homologous end joining (NHEJ) pathway. Activation of DNA end resection involves phosphorylation of the 5′ to 3′ exonuclease EXO1 by the phosphoinositide 3-kinase-like kinases ATM (ataxia telangiectasia- mutated) and ATR (ATM and Rad3-related) and by the cyclin-dependent kinases 1 and 2. After activation, EXO1 must also be ...

Resection of DNA double-strand breaks (DSBs) is a pivotal step during which the choice between NHEJ and HR DNA repair pathways is made. Although CDKs are known to control initiation of resection, their role in regulating long-range resection remains elusive. Here we show that CDKs 1/2 phosphorylate the long-range resection nuclease EXO1 at four C-terminal S/TP sites during S/G2 phases of the cell cycle. Impairment of EXO1 phosphorylation attenuates resection, chromosomal integrity, cell survival and HR, but augments NHEJ upon DNA damage. In contrast, cells expressing phospho-mimic EXO1 are proficient in resection even after CDK inhibition and favour HR over NHEJ. Mutation of cyclin-binding sites on EXO1 attenuates CDK binding and EXO1 phosphorylation, causing a resection defect that can be rescued by phospho-mimic mutations. Mechanistically, phosphorylation of EXO1 augments its recruitment to DNA breaks possibly via interactions with BRCA1. In summary, phosphorylation of EXO1 by CDKs is a novel ...

DNA double-strand breaks (DSBs) containing unligatable termini are potent cytotoxic lesions leading to growth arrest or cell death. The Artemis nuclease and tyrosyl-DNA phosphodiesterase (TDP1) are each capable of resolving protruding 3′-phosphoglycolate (PG) termini of DNA double-strand breaks (DSBs). Consequently, a knockout of Artemis and a knockout/knockdown of TDP1 rendered cells sensitive to the radiomimetic agent neocarzinostatin (NCS), which induces 3′-PG-terminated DSBs. Unexpectedly, however, a knockdown or knockout of TDP1 in Artemis-null cells did not confer any greater sensitivity than either deficiency alone, indicating a strict epistasis between TDP1 and Artemis. Moreover, a deficiency in Artemis, but not TDP1, resulted in a fraction of unrepaired DSBs, which were assessed as 53BP1 foci. Conversely, a deficiency in TDP1, but not Artemis, resulted in a dramatic increase in dicentric chromosomes following NCS treatment. An inhibitor of DNA-dependent protein kinase, a key regulator

Rapid progress in the study on the association of histone modifications with chromatin remodeling factors has broadened our understanding of chromatin dynamics in DNA transactions. In DNA double-strand break (DSB) repair, the well-known mark of histones is the phosphorylation of the H2A variant, H2AX, which has been used as a surrogate marker of DSBs. The ubiquitylation of histone H2B by RNF20 E3 ligase was recently found to be a DNA damage-induced histone modification. This modification is required for DSB repair and regulated by a distinctive pathway from that of histone H2AX phosphorylation. Moreover, the connection between H2B ubiquitylation and the chromatin remodeling activity of SNF2H has been elucidated. In this review, we summarize the current knowledge of RNF20-mediated processes and the molecular link to H2AX-mediated processes during DSB repair.

Efficient and correct repair of DNA damage, especially DNA double-strand breaks (DSBs), is vital for the survival of individual cells and organisms. Defects in the DNA repair may lead to cell death or genomic instability and development of cancer. The repair of DSBs in cell lines with different DSB rejoining capabilities was studied after exposure to ionising radiation. A new cell lysis protocol performed at 0ºC, which prevents the inclusion of non-true DSBs in the quantification of DSBs by pulsed-field gel electrophoresis (PFGE), was developed. Results showed that when the standard protocol at 50ºC was used, 30-40% of the initial yield of DSBs corresponds to artifactual DSBs. The lesions transformed to DSBs during incubation at 50ºC were repaired within 60-90 minutes in vivo and the repair was independent of DNA-PK, XRCC1 and PARP-1.. Non-homologous end-joining (NHEJ) is the major DSB repair pathway in mammalian cells. We show that DSBs are processed into long single-stranded DNA (ssDNA) ...

The cohesin complex holds together newly-replicated chromatids and is involved in diverse pathways that preserve genome integrity. We show that in budding yeast, cohesin is transiently recruited to active replication origins and it spreads along DNA as forks progress. When DNA synthesis is impeded, cohesin accumulates at replication sites and is critical for the recovery of stalled forks. Cohesin enrichment at replication forks does not depend on H2A(X) formation, which differs from its loading requirements at DNA double-strand breaks (DSBs). However, cohesin localization is largely reduced in rad50delta mutants and cells lacking both Mec1 and Tel1 checkpoint kinases. Interestingly, cohesin loading at replication sites depends on the structural features of Rad50 that are important for bridging sister chromatids, including the CXXC hook domain and the length of the coiled-coil extensions. Together, these data reveal a novel function for cohesin in the maintenance of genome integrity during S phase. Scc1

Endonuclease that cooperates with the MRE11-RAD50-NBN (MRN) complex in DNA-end resection, the first step of double-strand break (DSB) repair through the homologous recombination (HR) pathway. HR is restricted to S and G2 phases of the cell cycle and preferentially repairs DSBs resulting from replication fork collapse. Key determinant of DSB repair pathway choice, as it commits cells to HR by preventing classical non-homologous end-joining (NHEJ). Functions downstream of the MRN complex and ATM, promotes ATR activation and its recruitment to DSBs in the S/G2 phase facilitating the generation of ssDNA. Component of the BRCA1-RBBP8 complex that regulates CHEK1 activation and controls cell cycle G2/M checkpoints on DNA damage (By similarity). During immunoglobulin heavy chain class-switch recombination, promotes microhomology-mediated alternative end joining (A-NHEJ) and plays an essential role in chromosomal translocations (By similarity).

Dr. Kathy Friedman is Associate Professor of Biological Sciences at Vanderbilt University in Nashville, TN. She received her Ph.D. from the University of Washington in the laboratory of Drs. Walt Fangman and Bonny Brewer studying the temporal regulation of DNA replication. During postdoctoral training with Dr. Thomas Cech at the University of Colorado at Boulder, Dr. Friedman initiated studies of the assembly and regulation of the yeast telomerase complex. Her laboratory at Vanderbilt University utilizes genetic and biochemical approaches in the model system Saccharomyces cerevisiae to elucidate mechanisms of genome maintenance. Specific interests include the regulation of telomere replication at normal chromosome ends and at DNA double-strand breaks. Dr. Friedman enjoys teaching genetics to undergraduate and graduate students at Vanderbilt and has been recognized for her achievements both as a laboratory mentor and classroom teacher.. ...

Mutations caused by DNA damage are a main driver of cancer. We discovered that recognition of newly synthesised histone H4 directs breast cancer type 1 susceptibility protein (BRCA1) to post-replicative chromatin. The switch from mutagenic to error-free DNA double strand break repair by homologous recombination is therefore controlled by chromatin. ...

The efficacy of treatment with a combination of radiotherapy and chemotherapy in the yeast Saccharomyces cerevisiae as suitable eukaryotic model is demonstrated by following the induction and repair of DNA double-strand breaks. These may be induced by ionizing radiation. Furthermore, the chemotherapeutic agent cisplatin may induce DNA double-strand breaks through cellular repair mechanisms. The aim of this study is to investigate the induction and repair of DNA double-strand breaks after a combination of treatment with cisplatin and radiation versus radiation alone under hypoxic conditions.The number of induced double-strand breaks caused by radiation under hypoxic conditions is dependent on dose. Following combined treatment with cisplatin and radiation, the radiation-induced double-strand breaks simply add to those induced by cisplatin. We identified no difference in the repair kinetics of double-strand breaks following radiation between cells treated with cisplatin and those not treated with ...

DNA double-strand breaks (DSBs) are among the most deleterious forms of DNA lesions in cells. Here we induced site-specific DSBs in yeast cells and monitored chromatin dynamics surrounding the DSB using Chromosome Conformation Capture (3C). We find that formation of a DSB within G1 cells is not sufficient to alter chromosome dynamics. However, DSBs formed within an asynchronous cell population result in large decreases in both intra- and interchromosomal interactions. Using live cell microscopy, we find that changes in chromosome dynamics correlate with relocalization of the DSB to the nuclear periphery. Sequestration to the periphery requires the nuclear envelope protein, Mps3p, and Mps3p-dependent tethering delays recombinational repair of a DSB and enhances gross chromosomal rearrangements. Furthermore, we show that components of the telomerase machinery are recruited to a DSB and that telomerase recruitment is required for its peripheral localization. Based on these findings, we propose that

Nonhomologous end joining (NHEJ) eliminates DNA double-strand breaks (DSBs) by direct ligation. NHEJ involves binding of the KU heterodimer to double-stranded DNA ends, recruitment of DNA-PKcs (MRX complex in yeast), processing of ends, and recruitment of the DNA ligase IV (LIG4)-XRCC4 complex, which brings about ligation. A recent study shows that bacteria accomplish NHEJ using just two proteins (Ku and DNA ligase), whereas eukaryotes require many factors. NHEJ repairs DSBs at all stages of the cell cycle, bringing about the ligation of two DNA DSBs without the need for sequence homology, and so is error-prone ...

Nonhomologous end joining (NHEJ) eliminates DNA double-strand breaks (DSBs) by direct ligation. NHEJ involves binding of the KU heterodimer to double-stranded DNA ends, recruitment of DNA-PKcs (MRX complex in yeast), processing of ends, and recruitment of the DNA ligase IV (LIG4)-XRCC4 complex, which brings about ligation. A recent study shows that bacteria accomplish NHEJ using just two proteins (Ku and DNA ligase), whereas eukaryotes require many factors. NHEJ repairs DSBs at all stages of the cell cycle, bringing about the ligation of two DNA DSBs without the need for sequence homology, and so is error-prone ...

Principal Investigator：IHARA Makoto, Project Period (FY)：1998 - 2000, Research Category：Grant-in-Aid for Scientific Research (C), Section：一般, Research Field：Radiation science

Metalloprotease that specifically cleaves Lys-63-linked polyubiquitin chains (PubMed:19214193, PubMed:20656690, PubMed:24075985, PubMed:26344097). Does not have activity toward Lys-48-linked polyubiquitin chains. Component of the BRCA1-A complex, a complex that specifically recognizes Lys-63-linked ubiquitinated histones H2A and H2AX at DNA lesions sites, leading to target the BRCA1-BARD1 heterodimer to sites of DNA damage at double-strand breaks (DSBs). In the BRCA1-A complex, it specifically removes Lys-63-linked ubiquitin on histones H2A and H2AX, antagonizing the RNF8-dependent ubiquitination at double-strand breaks (DSBs) (PubMed:20656690). Catalytic subunit of the BRISC complex, a multiprotein complex that specifically cleaves Lys-63-linked ubiquitin in various substrates (PubMed:20656690, PubMed:24075985, PubMed:26344097, PubMed:26195665). Mediates the specific Lys-63-specific deubiquitination associated with the COP9 signalosome complex (CSN), via the interaction of the BRISC complex

We have demonstrated that the DNA dependent protein kinase (a key protein in DNA double strand breaks repair) protects HIF-1alpha against nuclear degradation and favors the survival of tumors cells upon hypoxia. Furthermore, hypoxia induces the activation of DNA-PK in the absence of DNA double-strand breaks (DSBs) suggesting that the hypoxic stress is able to induce a DNA damage like response in the absence of DNA lesions (Bouquet et al. 2011 J Cell Sci). The catalytic sub-unit of DNA-PK belongs to the family of PI3KK a family that, in eukaryotes, initiates cellular stress responses when genome integrity, mRNA translation, or nutrient availability is compromised. We have found that the DNA DSBs signaling protein ATM is also involved in the regulation of HIF-1 (Bouquet et al. 2010 Cell Cycle). Importantly, we recently demonstrated that the ATR protein, which is activated in response to replication stress, also regulates HIF-1 biosynthesis upon hypoxia (Fallone et al. 2012 Oncogene). Taken ...

DSB repair pathways. There are three DNA DSB repair pathways: single-strand annealing (SSA), homologous recombination (HR), and nonhomologous end-joining (NHEJ) (51). In SSA, which requires homologous sequences to flank the break site, the nonhomologous DNA ends are removed, leaving overhangs that are aligned and annealed (45). After annealing, DNA synthesis and ligation occur. SSA results in deletion of the region between homologous sequences. In contrast, HR is a relatively error-free mechanism for DSB repair that relies on the homologous region on the sister chromatid as a template for DNA synthesis (Figure 6A) (1;16). HR is limited to replicating cells (including early embryonic developmental stages) and can be observed only after DNA replication in the S and G2 phases (52) . Finally, NHEJ is a rapid repair mechanism that simply ligates broken DNA ends after minimal or no processing (Figure 6B). NHEJ occurs in all phases of the cell cycle and is thought to be the primary DNA repair pathway ...

The involvement of SETD2 in an important DNA repair pathway could explain the high frequency of SETD2 mutations in several cancers and may provide an alternative mechanism to evade the p53-mediated checkpoint.

The Rad51 (also known as RecA) family of recombinases executes the critical step in homologous recombination: the search for homologous DNA to serve as a template during the repair of DNA double-strand breaks (DSBs). Although budding yeast Rad51 has been extensively characterized in vitro, the strin …

We have determined the precise location and structure of the double-strand DNA breaks (DSBs) formed during Saccharomyces cerevisiae meiosis. Breaks were examined at two recombination hot spots in both wild-type and rad50S mutant cells. At both loci, breaks occurred at multiple, irregularly spaced si …

A sequence variant of histone H2A called H2AX is one of the important components of chromatin involved in DNA damage response induced by different genotoxic tensions. loss in normal cells and tissues as well as in those deficient in ATM, DNA-PK, and DSB repair proteins activity. The results of the latest scientific research of the low-dose irradiation phenomenon are offered including the bystander effect and the adaptive Piboserod IC50 response estimated by H2AX detection in cells and tissues. Keywords: Phosphorylation, Histone H2AX, Dephoshorylation, DNA double-strand breaks Introduction DNA double-strand breaks (DSBs) are the most dangerous lesions induced by a variety of treatments including ionizing radiation (IR), radiomimetic drugs, and lasers action. DSB removal is usually determinated by DSB repair system efficiency and is usually crucial for cell survival. Unsuccessful DSB repair prospects to the appearance of chromosomal aberrations in mitosis and potentially could induce malignancy. ...

Modern light microscopical techniques were employed to follow dynamical nuclear processes during the cell cycle and during DNA-repair. Laser-UVA-microirradiation The protein Rad51 is essential for the repair of double-strand breaks (DSBs) via the conservative homologous recombination repair pathway. To test the hypothesis that Rad51 localizes to damaged sites during DSB repair, a laser-UVA-microirradiation system was established. With this system spots with sizes around 1 µm in nuclei of living cells can be irradiated with UVA-light. After sensitization of cells by incorporation of BrdU into nuclear DNA and staining with the live cell dye Hoechst 33258, the system can be used to introduce double-strand breaks and single-strand breaks in the irradiated spots. The response of Rad51 to microirradiation By use of laser-UVA microirradiation the localization of Rad51 at damaged sites containing DNA double-strand breaks could be demonstrated. The accumulation of Rad51 at microirradiated sites was ...

Protein kinases-mediated regulation of DNA double-strand breaks sensing and repairCell cycle progression is tightly regulated by a plethora of proteins that act in order to preserve the integrity of the genetic information to be transferred to daughter cells. A wide variety of DNA lesions leads to activation of various cell cycle checkpoints controlled by specific protein kinases that play an important role in the preservation of genomic integrity. Two major repair pathways are activated in the presence of DNA double-strand breaks (DSBs), i.e. homologous recombination (HR) and non-homologous end-joining (NHEJ). Studies on mammalian cells have revealed that one of the earliest events occurring upon induction of DSBs is the activation of ATM (ataxia-telangiectasia mutated) protein kinase whose downstream targets (e.g. 53BP1, MDC1, BRCA1) assembly to sites of DSBs promoting repairing of the DNA. We have recently identified novel components of this complex machinery and demonstrated that protein ...

DNA dsb represent a severe type of damage that can be provoked in cell nuclei by exposure to ionizing radiation and also to chemicals such as etoposide, methylmethane sulphonate and bleomycin. Both radiation and chemical agents induce a variety of other DNA lesions such as base and sugar modifications, and single-strand breaks mostly mediated by the formation of highly reactive oxygen radicals (Iliakis, 1991). Although dsb are perhaps the lesions that most strongly threaten cell survival, it is likely that damaged cells seek to repair all DNA lesions produced by the oxygen radicals, thus activating several DNA repair pathways (Wood, 1996). Therefore, by using these DNA damaging agents, it might be difficult to analyze exclusively the repair of DNA dsb. On the other hand, it is desirable to have an in vitro system to study the repair of dsb produced in an environment as close as possible to the cell nucleus.. We used prokaryotic restriction enzymes (REs) in order to introduce solely dsb in the ...

Mutations in this gene are associated with Nijmegen breakage syndrome, an autosomal recessive chromosomal instability syndrome characterized by microcephaly, growth retardation, immunodeficiency, and cancer predisposition. The encoded protein is a member of the MRE11/RAD50 double-strand break repair complex which consists of 5 proteins. This gene product is thought to be involved in DNA double-strand break repair and DNA damage-induced checkpoint activation. [provided by RefSeq, Jul 2008 ...

the double-strand has to happen by chance. So, you can imagine it will be a very rare event. And you have to use a lot of tricks to make up for the fact that its a very, very rare event, things like antibiotic selection, and so forth, to pull out just the cells that you want that have the event the question. By using a genome editing tool, by consciously going in and generating a double-strand break, youre jump-starting the process. The cell has multiple repair mechanisms by which it can repair that double-strand break. Depending on what you want to do, whether you want to knock out a gene, or knock in a DNA variant, or knock in a whole cassette, you take advantage of one or the other of these repair mechanisms. And, because theyre all started by a double-strand break, the efficiency becomes much, much higher if youre able to introduce that double-strand break at will. And so thats really the whole game of genome editing. Using a genome editing tool increases the efficiency by orders of ...

FUNCTION: [Summary is not available for the mouse gene. This summary is for the human ortholog.] This gene encodes the catalytic subunit of the DNA-dependent protein kinase (DNA-PK). It functions with the Ku70/Ku80 heterodimer protein in DNA double strand break repair and recombination. The protein encoded is a member of the PI3/PI4-kinase family.[provided by RefSeq, Jul 2010 ...

One of the major causes of carcinogenesis is loss of genome stability. RAD51 in process of homologous recombination (HR) played crucial role in maintenance integrity of genome through initiate of DNA double strand breaks repair. Presence of single nu

Single strand annealing was also characterized in several other respects. We have measured its homology dependence and showed that the product formation increases with increasing amounts of homology. Product formation plateaus at about 400 bp and the smallest amount of homology that we have tested was 29 bp which recombined at a level of 0.1%. Interestingly, if one repeat is immediately adjacent to the DSB, as little as 10 bp can be used at a very efficient frequency. We have also separated the repeats by as much as 25 kb and still obtain efficient repair by SSA deletion events. This process takes 6 hours during which time the cells arrest their cell cycle until the DSB can be repaired. This experiment suggests that the tails remain relatively stable during this time.. Genetic Requirements.. RAD52. SSA requires several gene products that are also required for other types of recombination assays. Rad52p, for example, is required for most if not all types of recombination processes including SSA. ...

The Rad51 (also known as RecA) family of recombinases executes the critical step in homologous recombination: the search for homologous DNA to serve as a template during the repair of DNA double-strand breaks (DSBs). Although budding yeast Rad51 has been extensively characterized in vitro, the stringency of its search and sensitivity to mismatched sequences in vivo remain poorly defined. Here, in Saccharomyces cerevisiae, we analysed Rad51-dependent break-induced replication in which the invading DSB end and its donor template share a 108-base-pair homology region and the donor carries different densities of single-base-pair mismatches. With every eighth base pair mismatched, repair was about 14% of that of completely homologous sequences. With every sixth base pair mismatched, repair was still more than 5%. Thus, completing break-induced replication in vivo overcomes the apparent requirement for at least 6-8 consecutive paired bases that has been inferred from in vitro studies. When ...

Gene Information Double-strand breaks in DNA result from genotoxic stresses and are among the most damaging of DNA lesions. This gene encodes a DNA repair factor essential for the nonhomologous end-joining pathway which preferentially mediates repair of double-stranded breaks. Mutations in this gene cause different kinds of severe combined immunodeficiency disorders. [provided by RefSeq Jul 2008]. ...

Here, we show for the first time that CDV, a U.S. Food and Drug Administration (FDA)-approved drug for the treatment of HCMV infection, possesses potent antitumor properties against glioblastoma. We found that CDV inhibits HCMV viral gene expression in HCMV-infected primary glioblastoma tumors derived from patient biopsy specimens, but that viral gene expression was not required for CDV antitumor activity. Our data indicate that CDV becomes incorporated into the DNA of proliferating tumor cells, which likely initiates DNA double-stranded breaks and/or stalling of the DNA replication fork, leading to tumor cell apoptosis. When CDV treatment was combined with ionizing radiation, we observed a dramatic increase in phosphorylation of histone H2AX, a sensitive indicator of DNA double-stranded breaks, thereby showing that the DNA-damaging effects of radiotherapy are exacerbated by CDV.. The current paradigm for the mechanism of action of CDV in treating various types of virus-associated diseases ...

The two methods of repair for DSBs are homologous recombination (HR) and non-homologous end-joining (NHEJ). In HR the nucleotide sequences of two sets of DNA are exchanged to repair the broken strands. For this to occur one of the sets of DNA must be undamaged. The 3 strand of the DSB invades the undamaged double stranded DNA and partially unwinds it for homologous pairing. The strands form branch migrations and Holiday junctions to restore the DNA with the DSB without any crossing over of the two sets 2. In NHEJ of a DSB, the broken pieces are ligated together (via the KU heterodimer) without the need of an undamaged, homologous set of DNA. However, there can be degradation at the ends of the break that can result in errors and the loss of DNA sequences2. DSBs caused from IR are usually repaired by this method4. DSBs can also be repaired by single-strand annealing (SSA) if the break occurs between two repeated sequences. The repeated sequences are extended on the ends and annealed to repair ...

The CRISPR-Cas9 system is an RNA-guided genome-editing tool that provides researchers a simple, easy, and quick way to modify the genomes of various organisms. Using this system, Cas9 is gudied to a target sequence where it cleaves the DNA to form a double-stranded DNA break (DSB). Cells repair the break through one of two approaches, non-homologous end joining (NHEJ) or by homology-directed repair (HDR). Most commonly, cells will utilize NHEJ, which has been shown to have high efficiencies in Cas9-mediated genome modification. However, NHEJ is notably imprecise, typically resutling in insertions or deletions (INDELS) that result in unpredictable outcomes. Many investigators desire precision targeting to modify exact DNA sequences (including SNPs, knock-ins, conditional knock-outs, etc) in a controlled setting. In these cases HDR from a provided DNA template is the preferred mechanism of repair; however, this pathway is utilized by the cell less frequently than NHEJ. A number of studies have ...

DNA double-strand breaks (DSBs) can be repaired by one of two major pathways--non-homologous end-joining (NHEJ) and homologous recombination (HR)--depending on

Tom Blundell maintains an active laboratory as Director of Research and Professor Emeritus in Biochemistry, University of Cambridge, where he was previously Sir William Dunn Professor and Head of Department (1996 - 2009) and Chair School of Biological Sciences (2003 - 2009). He previously held teaching and research positions in the Universities of London, Sussex and Oxford.. Tom researches on molecular, structural and computational biology of growth factors, receptor activation, signal transduction and DNA repair, important in cancer, tuberculosis and familial diseases. He has described complex assemblies of FGFR and Met receptor systems necessary for high signal to noise in cell signalling, and of DNA double-strand-break repair, both NHEJ including DNA-PK and HR (Rad51 and BRCA2).. Tom has produced many software packages for protein modelling and design including Modeller (~9500 citations) and Fugue (~1300 citations), and for predicting effects of mutations on stability and interactions (SDM & ...

The lethal effect of ionizing irradiation on tumour cells is mostly determined by the repair of DNA double-strand breaks (DSBs). Cells are able to repair most of the DSBs, but 1% to 3 % are either non- or mis-repaired, which will then give rise to lethal chromosomal aberrations. Cells have evolved complex DSB repair mechanisms with a stringent hierarchy to guarantee the genomic stability. However, in tumour cells both mechanisms as well as hierarchy are often disturbed. This knowledge is important for an understanding of the radiation response of tumours, but-most of all-for the establishment of new and specific targets for therapy.

8-12. A certain string processing language allows the programmer to break a string into two pieces. It costs $ n $ units of time to break a string of $ n $ characters into two pieces, since this involves copying the old string. A programmer wants to break a string into many pieces, and the order in which the breaks are made can affect the total amount of time used. For example, suppose we wish to break a 20-character string after characters 3, 8, and 10. If the breaks are made in left-right order, then the first break costs 20 units of time, the second break costs 17 units of time, and the third break costs 12 units of time, for a total of 49 steps. If the breaks are made in right-left order, the first break costs 20 units of time, the second break costs 10 units of time, and the third break costs 8 units of time, for a total of only 38 steps. Give a dynamic programming algorithm that takes a list of character positions after which to break and determines the cheapest break cost in $ O(n^3) $ ...

Double-strand DNA cleavage catalyzed by purified OsSpo11 and OsTopVIB proteins.Each purified OsSpo11 and OsTopVIB protein or a combination shown above the image

TSUBOUCHI Hideo , OGAWA Hideyuki Genetic material suffers various spontaneous or environmental damages, among which DNA double-strand breaks (DSBs) are fatal to the cell since they bring to the loss of genetic information. Cells prov … Seibutsu Butsuri 38(4), 156-161, 1998-07 J-STAGE Ichushi Web References (26) ...

Click on a genes description to view its network relationships with genes known to be involved in double strand break repair ...

A genomic interrogation of homosexuality turns up speculative links between genetic elements and sexual orientation, but researchers say the study is too small to be significant. 6 Comments. ...

TY - JOUR. T1 - Expression of ATM in ataxia telangiectasia fibroblasts rescues defects in DNA double-strand break repair in nuclear extracts. AU - Li, Yuling. AU - Carty, Michael P.. AU - Oakley, Gregory G.. AU - Seidman, Michael M.. AU - Medvedovic, Mario. AU - Dixon, Kathleen. PY - 2001/1/1. Y1 - 2001/1/1. N2 - Ataxia telangiectasia (A-T) is a human genetic disorder characterized by progressive cerebellar degeneration, hypersensitivity to ionizing radiation (IR), immunodeficiency, and high cancer risk. At the cellular level, IR sensitivity and increased frequency of spontaneous and IR-induced chromosomal breakage and rearrangements are the hallmarks of A-T. The ATM gene, mutated in this syndrome, has been cloned and codes for a protein sharing homology with DNA-PKcs, a protein kinase involved in DNA double-strand break (DSB) repair and DNA damage responses. The characteristics of the A-T cellular phenotypes and ATM gene suggest that ATM may play a role similar to that of DNA-PKcs in DSB repair ...

DSB repair pathway choice――――――. HR initiation at transcription active loci Yasuhara et al., Cell, 2018. DSB repair pathway choice in G1 Biehs et al., Mol Cell, 2017. 53BP1 dephosphorylation promotes pro-HR environment Isono et al., Cell Rep, 2017. MRE11-dependent initiation of resection Shibata et al., Mol Cell, 2014. DSB repair pathway choice in G2 Shibata et al., EMBO J, 2011. ―――DDR dependent immune ligands regulation―――. BER deficiency upregulates PD-L1 expression Permata et al., Oncogene, 2019. DSB-dependent ATR/Chk1 upregulats PD-L1 expression Sato et al., Nat Comm, 2017. ...

Meiotic recombination is initiated by DNA double-strand breaks (DSBs) created by the topoisomerase-like protein Spo11. During DSB formation, Spo11 becomes covalently attached to the 5 DSB ends. Removal of Spo11 is essential to repair the DSB by homologous recombination. Spo11 is removed endonucleolytically creating short-lived Spo11-oligonucleotide products. Here I demonstrate that: 1. Spo11-oligonucleotide products are not detected in recombination mutants believed to be defective in meiotic DSB formation. 2. When DSB repair is delayed, Spo11-oligonucleotides persist for longer. 3. Processing of Spo11-DSB ends to create Spo11-oligonucleotides is largely dependent on Mec1 and Tel1 activity. In the process of investigating Spo11-oligonucleotide degradation, it was observed that a mutant defective in both the meiotic recombination checkpoint and in DSB repair failed to accumulate the expected level of DSBs. Work described here leads to the proposal of a DSB feedback mechanism that functions ...

Ku70 is a protein that, in humans, is encoded by the XRCC6 gene. Together, Ku70 and Ku80 make up the Ku heterodimer, which binds to DNA double-strand break ends and is required for the non-homologous end joining (NHEJ) pathway of DNA repair. It is also required for V(D)J recombination, which utilizes the NHEJ pathway to promote antigen diversity in the mammalian immune system. In addition to its role in NHEJ, Ku is also required for telomere length maintenance and subtelomeric gene silencing. Ku was originally identified when patients with systemic lupus erythematosus were found to have high levels of autoantibodies to the protein. Mouse embryonic stem cells with homozygous Ku70 mutations, that is Ku70−/− cells, have markedly increased sensitivity to ionizing radiation compared to heterozygous Ku70+/− or wild-type Ku70+/+ embryonic stem cells. Mutant mice deficient in Ku70 exhibit early aging. Using several specific criteria of aging, the mutant mice were found to display the same aging ...

PROJECT SUMMARY Lung cancer continues to be the leading cause of cancer death in the U.S. One of the key strategies to combating it is to better understand its causes. Hexavalent chromium [Cr(VI)] is a human lung carcinogen of major public health concern because exposure to it is common in the workplace and in the general environment. Our study focuses on investigating the mechanisms of Cr(VI)-induced carcinogenesis, which are currently unknown. In particular, this work focuses on the particulate Cr(VI) compounds, because they are the most potent Cr(VI) carcinogens. Recent studies indicate particulate Cr(VI) induces chromosome instability and causes cells to evade DNA double strand break repair, which are hallmarks of human lung cancer. Thus, this research focuses on how particulate Cr(VI) induces cells to evade DNA double strand break repair leading to chromosome instability and carcinogenesis. Our data show prolonged exposure to particulate Cr(VI) specifically impacts the effector arm of ...

Nonhomologous end joining (NHEJ) and homologous recombination (HR) represent the two major pathways of DNA double-strand break (DSB) repair in eukaryotic cells. NHEJ repairs DSBs by ligation of cognate broken ends irrespective of homologous flanking sequences, whereas HR repairs DSBs using an undamaged homologous template. Although both NHEJ and HR have been clearly implicated in the maintenance of genome stability, how these apparently independent and mechanistically distinct pathways are coordinated remains largely unexplored. To investigate the relationship between HR and NHEJ modes of DSB repair, cells doubly deficient for the NHEJ factor DNA Ligase IV (Lig4) and the HR factor Rad54 were generated. Lig4 and Rad54 cooperate to support cellular proliferation, repair spontaneous DSBs, and prevent chromosome and single chromatid aberrations. These findings demonstrate a role for NHEJ in the repair of DSBs that occur spontaneously during or after DNA replication, and reveal overlapping functions ...

TY - JOUR. T1 - Reduced Rif2 and lack of Mec1 target short telomeres for elongation rather than double-strand break repair. AU - McGee, Jean S.. AU - Phillips, Jane A.. AU - Chan, Angela. AU - Sabourin, Michelle. AU - Paeschke, Katrin. AU - Zakian, Virginia A.. N1 - Funding Information: We thank D. Shore and A. Bianchi (University of Geneva) for strains and advice on the DSB assay, K. Runge for advice on telomere PCR, T. Petes (Duke University) for strains, M. Jayaram for discussions about FLP-induced DSBs and C. Webb and Y. Wu for comments on the manuscript. This work was supported by grants from the US National Institutes of Health (NIH; GM43265 to V.A.Z.), postdoctoral fellowships from the NIH (M.S.), Deutsche Forschungsgemeinschaft (K.P.) and NJCCR (K.P.), and predoctoral fellowships from NJCCR (J.A.P., J.S.M.) and NIH (J.S.M.).. PY - 2010/12. Y1 - 2010/12. N2 - Telomerase in Saccharomyces cerevisiae binds and preferentially elongates short telomeres, and this process requires the checkpoint ...

DNA double-strand breaks (DSBs) are mainly repaired either by homologous recombination (HR) or by nonhomologous end-joining (NHEJ) pathways. Here, we showed that myeloid cell leukemia sequence 1 (Mcl-1) acts as a functional switch in selecting between HR and NHEJ pathways. Mcl-1 was cell cycle-regulated during HR, with its expression peaking in S/G2 phase. While endogenous Mcl-1 depletion reduced HR and enhanced NHEJ, Mcl-1 overexpression resulted in a net increase in HR over NHEJ. Mcl-1 directly interacted with the dimeric Ku protein complex via its Bcl-2 homology 1 and 3 (BH1 and BH3) domains, which are required for Mcl-1 to inhibit Ku-mediated NHEJ. Mcl-1 also promoted DNA resection mediated by the Mre11 complex and HR-dependent DSB repair. Using the Mcl-1 BH1 domain as a docking site, we identified a small molecule, MI-223, that directly bound to BH1 and blocked Mcl-1-stimulated HR DNA repair, leading to sensitization of cancer cells to hydroxyurea- or olaparib-induced DNA replication ...

Efficient methodologies for recreating cancer-associated chromosome translocations are in high demand as tools for investigating how such events initiate cancer. The CRISPR/Cas9 system has been used to reconstruct the genetics of these complex rearrangements at native loci while maintaining the architecture and regulatory elements. However, the CRISPR system remains inefficient in human stem cells. Here, we compared three strategies aimed at enhancing the efficiency of the CRISPR-mediated t(11;22) translocation in human stem cells, including mesenchymal and induced pluripotent stem cells: (1) using end-joining DNA processing factors involved in repair mechanisms, or (2) ssODNs to guide the ligation of the double-strand break ends generated by CRISPR/Cas9; and (3) all-in-one plasmid or ribonucleoprotein complex-based approaches ...

TY - JOUR. T1 - The Ku80 carboxy terminus stimulates joining and artemis-mediated processing of DNA ends. AU - Weterings, Eric. AU - Verkaik, Nicole S.. AU - Keijzers, Guido. AU - Florea, Bogdan I.. AU - Wang, Shih Ya. AU - Ortega, Laura G.. AU - Uematsu, Naoya. AU - Chen, David J.. AU - Van Gent, Dik C.. PY - 2009/3/1. Y1 - 2009/3/1. N2 - Repair of DNA double-strand breaks (DSBs) is predominantly mediated by nonhomologous end joining (NHEJ) in mammalian cells. NHEJ requires binding of the Ku70-Ku80 heterodimer (Ku70/80) to the DNA ends and subsequent recruitment of the DNA-dependent protein kinase catalytic subunit (DNA-PKCS) and the XRCC4/ligase IV complex. Activation of the DNA-PKCS serine/threonine kinase requires an interaction with Ku70/80 and is essential for NHEJ-mediated DSB repair. In contrast to previous models, we found that the carboxy terminus of Ku80 is not absolutely required for the recruitment and activation of DNA-PKCS at DSBs, although cells that harbored a carboxy-terminal ...

Purpose: The clinical radiation responses of different organs vary widely and likely depend on the intrinsic radiosensitivities of their different cell populations. Double-strand breaks (DSBs) are the most deleterious form of DNA damage induced by ionizing radiation, and the cells capacity to rejoin radiation-induced DSBs is known to affect their intrinsic radiosensitivity. To date, only little is known about the induction and processing of radiation-induced DSBs in complex normal tissues. Using an in vivo model with repair-proficient mice, the highly sensitive {gamma}H2AX immunofluorescence was established to investigate whether differences in DSB rejoining could account for the substantial differences in clinical radiosensitivity observed among normal tissues. Methods and Materials: After whole body irradiation of C57BL/6 mice (0.1, 0.5, 1.0, and 2.0 Gy), the formation and rejoining of DSBs was analyzed by enumerating {gamma}H2AX foci in various organs representative of both early-responding ...

A sequence variant of histone H2A called H2AX is one of the important components of chromatin involved in DNA damage response induced by different genotoxic tensions. loss in normal cells and tissues as well as in those deficient in ATM, DNA-PK, and DSB repair proteins activity. The results of the latest scientific research of the low-dose irradiation phenomenon are offered including the bystander effect and the adaptive Piboserod IC50 response estimated by H2AX detection in cells and tissues. Keywords: Phosphorylation, Histone H2AX, Dephoshorylation, DNA double-strand breaks Introduction DNA double-strand breaks (DSBs) are the most dangerous lesions induced by a variety of treatments including ionizing radiation (IR), radiomimetic drugs, and lasers action. DSB removal is usually determinated by DSB repair system efficiency and is usually crucial for cell survival. Unsuccessful DSB repair prospects to the appearance of chromosomal aberrations in mitosis and potentially could induce malignancy. ...

Supplementary MaterialsTable_1. between healthy people and three types of familial breasts cancer situations: mutated (((group (situations (promoter is certainly associated with changed risk of breasts cancer in verified the current presence of hereditary predisposition for familial breasts cancers (2C4). These genes keep genome balance in regular cells by mending double-strand breaks generally through homologous recombination (HR) pathway; their mutated forms result in genome instability and elevated risk for breasts cancer advancement (5). A couple of two types of DNA double-strand break repair mechanisms: non-homologous end joining (NHEJ) and HR (6). Deficiency in the HR pathway, mainly caused by germline mutations, is well known to increase the risk of breast cancer (7); however, it is not equally obvious whether deficiency in NHEJ pathway can also increase breast malignancy risk (8). Ku is usually a heterodimer consisting of Ku80 encoded by and Ku70 encoded by (9C13). Deletion of in mice ...

Much of our current knowledge of DSB repair in eukaryotes is obtained from studies utilizing rare cutting endonucleases to generate site-specific DSBs in the genome. In Drosophila, P-element transposition has been widely employed as a system to generate site-specific chromosomal breaks (for a review see Lankenau 1995; Gloor and Lankenau 1998). In our study, excision of the 629-bp whd P-element was used to induce two staggered DSBs with identical 17-nucleotide 3′ overhangs at the site of integration in exon 6 of the white gene. Repair of these breaks is achieved through NHEJ and HR, the two main DSB-repair pathways in eukaryotes (Pastink et al. 2001). Since the P-element-specific endonuclease is constitutively expressed during development, the P-element is a target to continuous excision. The formation of DSBs is prevented when the breaks are resealed incorrectly via end joining or when in females the homologous chromosome, which does not contain the whd P-element, is used as a template for ...

Efficient and correct repair of DNA damage, especially DNA double-strand breaks (DSBs), is vital for the survival of individual cells and organisms. Defects in the DNA repair may lead to cell death or genomic instability and development of cancer. The repair of DSBs in cell lines with different DSB rejoining capabilities was studied after exposure to ionising radiation. A new cell lysis protocol performed at 0ºC, which prevents the inclusion of non-true DSBs in the quantification of DSBs by pulsed-field gel electrophoresis (PFGE), was developed. Results showed that when the standard protocol at 50ºC was used, 30-40% of the initial yield of DSBs corresponds to artifactual DSBs. The lesions transformed to DSBs during incubation at 50ºC were repaired within 60-90 minutes in vivo and the repair was independent of DNA-PK, XRCC1 and PARP-1.. Non-homologous end-joining (NHEJ) is the major DSB repair pathway in mammalian cells. We show that DSBs are processed into long single-stranded DNA (ssDNA) ...

The MRE11/RAD50/NBS1 (MRN) complex is a major sensor of DNA double strand breaks, whose role in controlling faithful DNA replication and preventing replication stress is also emerging. Inactivation of the MRN complex invariably leads to developmental and/or degenerative neuronal defects, the pathogenesis of which still remains poorly understood. In particular, NBS1 gene mutations are associated with microcephaly and strongly impaired cerebellar development, both in humans and in the mouse model. These phenotypes strikingly overlap those induced by inactivation of MYCN, an essential promoter of the expansion of neuronal stem and progenitor cells, suggesting that MYCN and the MRN complex might be connected on a unique pathway essential for the safe expansion of neuronal cells. Here, we show that MYCN transcriptionally controls the expression of each component of the MRN complex. By genetic and pharmacological inhibition of the MRN complex in a MYCN overexpression model and in the more ...

Late Radiation Morbidity Incidence in a South-East Scottish cohort and investigation into abnormalities in DNA double-strand break repair and damage ...

Kumaran, R. I., Li, J. J., Dynan, W. S., Porteus, M. H., Spector, D. L. (May 2016) Analysis of DNA Repair Pathway Choice Upon Induction of Double-Strand Breaks by Engineered Nuclease. Molecular Therapy, 24 (Supple). S225-S225. ISSN 1525-0016 Chen, Z., Jaafar, L., Agyekum, D. G., Xiao, H., Wade, M. F., Kumaran, R. I., Spector, D. L., Bao, G., Porteus, M. H., Dynan, W. S., Meiler, S. E. (August 2013) Receptor-mediated delivery of engineered nucleases for genome modification. Nucleic Acids Research, 41 (19). ISSN 03051048 (ISSN) Sehgal, P., Chaturvedi, P., Kumaran, R. I., Kumar, S., Parnaik, V. K. (February 2013) Lamin A/C Haploinsufficiency Modulates the Differentiation Potential of Mouse Embryonic Stem Cells. PLoS ONE, 8 (2). ISSN 1932-6203 (Electronic) Meiler, S. E., Dynan, W. S., Porteus, M., Spector, D. L., Bao, G., Chen, Z., Pandey, J., Goyal, D., Kumaran, R. I., Wade, M., Agyekum, D. G. (November 2011) A Biological Delivery Platform for Zinc Finger Nucleases Using Transferrin-Mediated ...

We also analyzed effects of low dose radiation on DNA double strand break repair using a transgenic mouse model in which homologous recombination (HR) events at a transgenic reporter give rise to fluorescent cells. To establish this model as a study tool for radiation effects we first exposed mice to 6 Gy radiation and analyzed the number of HR events. Treated animals displayed a significantly higher frequency of HR events than littermate controls (n=25; p=0.006). We then exposed transgenic mice to continuous very low dose radiation as described above and analyzed HR frequency but did not detect any difference in HR frequency as compared to littermate controls (n=60; p=0.8 ...

Endonucleolytic processing of covalent protein-linked DNA double-strand breaks.: DNA double-strand breaks (DSBs) with protein covalently attached to 5 strand t

Zimmer J, Tacconi EM, Folio C, Badie S, Porru M, Klare K, Tumiati M, Markkanen E, Halder S, Ryan A, Jackson SP, Ramadan K, Kuznetsov SG, Biroccio A, Sale JE, Tarsounas M. Molecular Cell 61, 449-460. G-quadruplex (G4)-forming genomic sequences, including telomeres, represent natural replication fork barriers. Stalled replication forks can be stabilized and restarted by homologous recombination (HR), which also repairs DNA double-strand breaks (DSBs) arising at collapsed forks.

The National Centre for Biological Sciences is delighted to welcome Anjana Badrinarayanan who joins the Centre as its newest faculty member.. Anjana is interested in understanding how cells maintain chromosome integrity, which is essential for survival. During her PhD with David Sherratt at the University of Oxford, she studied the role of the highly conserved Structural Maintenance of Chromosome (SMC) complex in organization and segregation of the bacterial chromosome. Her studies revealed the composition and dynamics of the SMC complex during chromosome organization and condensation and provided evidence for a novel loading mechanism of the complex on DNA. To further study the dynamics of chromosomes under the onslaught of DNA damage, she joined the laboratory of Michael Laub at the Massachusetts Institute of Technology as an HFSP Long-Term Postdoctoral Fellow. Here, she initiated a research project to visualize the process of DNA double-strand break repair inside living cells. Double-strand ...

Tumor cure with conventional fractionated radiotherapy is 65%, dependent on tumor cell-autonomous gradual buildup of DNA double-strand break (DSB) misrepair. Here we report that single-dose radiotherapy (SDRT), a disruptive technique that ablates more than 90% of human cancers, operates a distinct dual-target mechanism, linking acid sphingomyelinase-mediated (ASMase-mediated) microvascular perfusion defects to DNA unrepair in tumor cells to confer tumor cell lethality. ASMase-mediated microcirculatory vasoconstriction after SDRT conferred an ischemic stress response within parenchymal tumor cells, with ROS triggering the evolutionarily conserved SUMO stress response, specifically depleting chromatin-associated free SUMO3. Whereas SUMO3, but not SUMO2, was indispensable for homology-directed repair (HDR) of DSBs, HDR loss of function after SDRT yielded DSB unrepair, chromosomal aberrations, and tumor clonogen demise. Vasoconstriction blockade with the endothelin-1 inhibitor BQ-123, or ROS ...

Tumor cure with conventional fractionated radiotherapy is 65%, dependent on tumor cell-autonomous gradual buildup of DNA double-strand break (DSB) misrepair. Here we report that single-dose radiotherapy (SDRT), a disruptive technique that ablates more than 90% of human cancers, operates a distinct dual-target mechanism, linking acid sphingomyelinase-mediated (ASMase-mediated) microvascular perfusion defects to DNA unrepair in tumor cells to confer tumor cell lethality. ASMase-mediated microcirculatory vasoconstriction after SDRT conferred an ischemic stress response within parenchymal tumor cells, with ROS triggering the evolutionarily conserved SUMO stress response, specifically depleting chromatin-associated free SUMO3. Whereas SUMO3, but not SUMO2, was indispensable for homology-directed repair (HDR) of DSBs, HDR loss of function after SDRT yielded DSB unrepair, chromosomal aberrations, and tumor clonogen demise. Vasoconstriction blockade with the endothelin-1 inhibitor BQ-123, or ROS ...

Tumor cure with conventional fractionated radiotherapy is 65%, dependent on tumor cell-autonomous gradual buildup of DNA double-strand break (DSB) misrepair. Here we report that single-dose radiotherapy (SDRT), a disruptive technique that ablates more than 90% of human cancers, operates a distinct dual-target mechanism, linking acid sphingomyelinase-mediated (ASMase-mediated) microvascular perfusion defects to DNA unrepair in tumor cells to confer tumor cell lethality. ASMase-mediated microcirculatory vasoconstriction after SDRT conferred an ischemic stress response within parenchymal tumor cells, with ROS triggering the evolutionarily conserved SUMO stress response, specifically depleting chromatin-associated free SUMO3. Whereas SUMO3, but not SUMO2, was indispensable for homology-directed repair (HDR) of DSBs, HDR loss of function after SDRT yielded DSB unrepair, chromosomal aberrations, and tumor clonogen demise. Vasoconstriction blockade with the endothelin-1 inhibitor BQ-123, or ROS ...

Tumor cure with conventional fractionated radiotherapy is 65%, dependent on tumor cell-autonomous gradual buildup of DNA double-strand break (DSB) misrepair. Here we report that single-dose radiotherapy (SDRT), a disruptive technique that ablates more than 90% of human cancers, operates a distinct dual-target mechanism, linking acid sphingomyelinase-mediated (ASMase-mediated) microvascular perfusion defects to DNA unrepair in tumor cells to confer tumor cell lethality. ASMase-mediated microcirculatory vasoconstriction after SDRT conferred an ischemic stress response within parenchymal tumor cells, with ROS triggering the evolutionarily conserved SUMO stress response, specifically depleting chromatin-associated free SUMO3. Whereas SUMO3, but not SUMO2, was indispensable for homology-directed repair (HDR) of DSBs, HDR loss of function after SDRT yielded DSB unrepair, chromosomal aberrations, and tumor clonogen demise. Vasoconstriction blockade with the endothelin-1 inhibitor BQ-123, or ROS ...

Funding. Work in the Mailand group is generously supported by The Novo Nordisk Foundation, European Research Council (ERC), EMBO, The Danish Council for Independent Research, The Danish Cancer Society, and The Lundbeck Foundation.. For more information and inquiries about available positions, please contact Prof. Niels Mailand (email: [email protected]).. Selected recent publications:. Haahr P, Hoffmann S, Tollenaere M, Ho T, Toledo L, Mann M, Bekker-Jensen S, Raschle M, Mailand N. (2016). Activation of the ATR kinase by the RPA-binding protein ETAA1. Nature Cell Biol 18, 1196-1207.. Hansen RK, Mund A, Poulsen SL, Sandoval M, Tsouroula K, Klement K, Tollenaere MAX, Räschle M, Soria R, Offermanns S, Worzfeld T, Grosse R, Brandt DT, Rozell B, Mann M, Cole F, Soutoglou E, Goodarzi AA, Daniel J, Mailand N, Bekker-Jensen S. (2016). SCAI promotes DNA double-strand break repair in distinct chromosomal contexts. Nature Cell Biol 18, 1357-1366.. Saredi G, Huang H, Hammond CM, Alabert C, ...

Recently, I developed an alternate co-conversion marker: a temperature-sensitive lethal allele of pha-1. Oligo-templated repair of a CRISPR/Cas9 DNA double strand break at the pha-1 locus enriches for CRISPR/Cas9-catalyzed modifications at other loci and allows recovery of edits with minimal handling. Other than the injected parental animal, only pha-1(ts) rescued F1 are alive on a plate. Inactivation of non-homologous end-joining further increases knock-in efficiency. Homology arms of 35-80 bp are sufficient for efficient editing, and DSBs up to 54 bp away from the insertion point resulted in knock-ins. This method is described in detail in a publication (Ward, Genetics, 2015). Here, I provide a detailed guide to experimental design and practical considerations. A link to a printable PDF document is also provided below ...

BRCA1/2 are DNA repair enzymes that are the essential components of homologous repair (HR), which is invoked when DNA doubles strand breaks are encountered. HR works via consultation of sister chromatids during late S to early G2 of the cell cycle - without repair of double strand breaks, mitotic catastrophe ensues: Continue reading →. ...

Cells lacking ATM exhibited a slightly increased number of chromosomal breaks in untreated cells compared to VA13 . On the other hand, oxLDL SB-742457 cost kinase inhibitor drastically enhanced chromosomal breaks in each cell lines. In VA13 cells, the quantity of chromosomal breaks following 8 h greater up to thirty. In AT22 cells the quantity of chromosomal breaks improved as much as 42. Fig. 6B even further displays that the variety of oxLDL induced chromosomal breaks in AT22 cells are significantly higher when in contrast to VA13 cells. Treatment method of VA13 and AT22 cells with LDL was without results on chromosomal breaks when compared to untreated cells . three.5. PDTC scavenges oxLDL induced elevated ROS ranges in the T cells ATM deficient cells are inside a consistent state of oxidative pressure and might possibly exhibit diminished antioxidant capability . We demonstrate that AT22 cells exhibited approx. one.five fold higher ROS amounts when compared to VA13 cells . Incubation of ...

TY - JOUR. T1 - Inter-individual and inter-cell type variation in residual DNA damage after in vivo irradiation of human skin. AU - Chua, Melvin Lee Kiang. AU - Somaiah, Navita. AU - Bourne, Sara. AU - Daley, Frances. AU - AHern, Roger. AU - Nuta, Otilia. AU - Davies, Sue. AU - Herskind, Carsten. AU - Pearson, Ann. AU - Warrington, Jim. AU - Helyer, Sarah. AU - Owen, Roger. AU - Yarnold, John. AU - Rothkamm, Kai. PY - 2011/5. Y1 - 2011/5. N2 - Purpose: The aim of this study was to compare inter-individual and inter-cell type variation in DNA double-strand break (DSB) repair following in vivo irradiation of human skin. Materials and methods: Duplicate 4 mm core biopsies of irradiated and unirradiated skin were collected from 35 patients 24 h after 4 Gy exposure using 6 MeV electrons. Residual DSB were quantified by scoring 53BP1 foci in dermal fibroblasts, endothelial cells, superficial keratinocytes and basal epidermal cells. Results: Coefficients of inter-individual variation for levels of ...

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The measurement of -L2AX foci induction in cells provides a sensitive and reliable method for the quantitation of DNA harm responses in a variety of cell types. in the DNA-PKcs defective cells (XP14BRneo17), we noticed an more advanced preservation of foci in the nuclei a sign of incomplete fix of DNA DSB. In overview, the program of image resolution stream cytometry provides allowed an evaluation of foci in a huge amount of cells (20,000) for each cell series at each period stage. This provides a story technique to determine distinctions in fix kinetics between different cell types. We recommend that image resolution stream cytometry provides an choice system for accurate computerized high through-put evaluation of foci induction in a range of cell types. ? 2011 Cosmopolitan Culture for Advancement Troxacitabine of Cytometry gene which features in the control of cell routine criminal arrest and induction of DNA DSB fix outcomes in the tenacity of -L2AX foci in the nucleus of cells shown to IR ...

Clostridium botulinum produces a transcription factor that can aggregate and self-propagate a prion-like form, leading to genome-wide changes in gene expression in E. coli, according to a study.. 1 Comment. ...

Developmental biologist Maria Jasin focuses on double-strand break repair and genomic integrity in mammalian cells and the relationship to tumor suppression.

Every day, thousands of DNA damaging events occur within a cell which must be accurately repaired in order to maintain a stable genome. A key part of this process is how a cell can rapidly detect such damage and activate the appropriate cellular responses to facilitate accurate repair mechanisms. If the damage is too much for a cell to deal with, then alternative failsafe mechanisms trigger, leading to the destruction of the cell to prevent potentially pro-mutagenic lesions being passed on to progeny. These DNA repair and failsafe processes are often dysregulated in human cancers, and researchers within SInFoNiA are trying to understand how this occurs, and how it may offer a means to specifically kill cancer cells to improve current and future therapeutic regimes for cancer patients. These repair processes are also important during development and the attrition of genome integrity is a cause of cellular and organismal ageing. As, SInFoNiA researchers are also studying various aspects of these ...