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Sample records for repair genes predispose

  1. Deficient DNA repair capacity, a predisposing factor in breast cancer.

    PubMed Central

    Parshad, R.; Price, F. M.; Bohr, V. A.; Cowans, K. H.; Zujewski, J. A.; Sanford, K. K.

    1996-01-01

    Women with breast cancer and a family history of breast cancer and some with sporadic breast cancer are deficient in the repair of radiation-induced DNA damage compared with normal donors with no family history of breast cancer. DNA repair was measured indirectly by quantifying chromatid breaks in phytohaemagglutinin (PHA)-stimulated blood lymphocytes after either X-irradiation or UV-C exposure, with or without post treatment with the DNA repair inhibitor, 1-beta-D-arabinofuranosylcytosine (ara-C). We have correlated chromatid breaks with unrepaired DNA strand breaks using responses to X-irradiation of cells from xeroderma pigmentosum patients with well-characterised DNA repair defects or responses of repair-deficient mutant Chinese hamster ovary (CHO) cells with or without transfected human DNA repair genes. Deficient DNA repair appears to be a predisposing factor in familial breast cancer and in some sporadic breast cancers. PMID:8679441

  2. Genomic scan for genes predisposing to schizophrenia

    SciTech Connect

    Coon, H.; Jensen. S.; Holik, J.

    1994-03-15

    We initiated a genome-wide search for genes predisposing to schizophrenia by ascertaining 9 families, each containing three to five cases of schizophrenia. The 9 pedigrees were initially genotyped with 329 polymorphic DNA loci distributed throughout the genome. Assuming either autosomal dominant or recessive inheritance, 254 DNA loci yielded lod scores less than -2.0 at {theta} = 0.0, 101 DNA markers gave lod scores less than -2.0 at {theta} = 0.05, while 5 DNA loci produced maximum lod scores greater than 1: D4S35, D14S17, D15S1, D22S84, and D22S55. Of the DNA markers yielding lod scores greater than 1, D4S35 and D22S55 also were suggestive of linkage when the Affected-Pedigree-Member method was used. The families were then genotyped with four highly polymorphic simple sequence repeat markers; possible linkage diminished with DNA markers mapping nearby D4S35, while suggestive evidence of linkage remained with loci in the region of D22S55. Although follow-up investigation of these chromosomal regions may be warranted, our linkage results should be viewed as preliminary observations, as 35 unaffected persons are not past the age of risk. 90 refs., 3 tabs.

  3. Human DNA repair genes.

    PubMed

    Wood, R D; Mitchell, M; Sgouros, J; Lindahl, T

    2001-02-16

    Cellular DNA is subjected to continual attack, both by reactive species inside cells and by environmental agents. Toxic and mutagenic consequences are minimized by distinct pathways of repair, and 130 known human DNA repair genes are described here. Notable features presently include four enzymes that can remove uracil from DNA, seven recombination genes related to RAD51, and many recently discovered DNA polymerases that bypass damage, but only one system to remove the main DNA lesions induced by ultraviolet light. More human DNA repair genes will be found by comparison with model organisms and as common folds in three-dimensional protein structures are determined. Modulation of DNA repair should lead to clinical applications including improvement of radiotherapy and treatment with anticancer drugs and an advanced understanding of the cellular aging process. PMID:11181991

  4. Gene Therapy for Cartilage Repair

    PubMed Central

    Madry, Henning; Orth, Patrick; Cucchiarini, Magali

    2011-01-01

    The concept of using gene transfer strategies for cartilage repair originates from the idea of transferring genes encoding therapeutic factors into the repair tissue, resulting in a temporarily and spatially defined delivery of therapeutic molecules to sites of cartilage damage. This review focuses on the potential benefits of using gene therapy approaches for the repair of articular cartilage and meniscal fibrocartilage, including articular cartilage defects resulting from acute trauma, osteochondritis dissecans, osteonecrosis, and osteoarthritis. Possible applications for meniscal repair comprise meniscal lesions, meniscal sutures, and meniscal transplantation. Recent studies in both small and large animal models have demonstrated the applicability of gene-based approaches for cartilage repair. Chondrogenic pathways were stimulated in the repair tissue and in osteoarthritic cartilage using genes for polypeptide growth factors and transcription factors. Although encouraging data have been generated, a successful translation of gene therapy for cartilage repair will require an ongoing combined effort of orthopedic surgeons and of basic scientists. PMID:26069580

  5. TNXB locus may be a candidate gene predisposing to schizophrenia.

    PubMed

    Wei, J; Hemmings, G P

    2004-02-15

    We report here on the detection of nine single nucleotide polymorphisms (SNPs) near to the NOTCH4 locus in the search for schizophrenia susceptibility genes in the class III region of the human major histocompatibility complex (MHC). We totally analyzed 122 family trios recruited in the UK. The TDT analysis demonstrated that of the nine SNPs, three were associated with schizophrenia, including rs1009382 (P = 0.00047), rs204887 (P = 0.007), and rs8283 (P = 0.015). Both rs1009382 and rs204887 are present in the TNXB locus. The rs1009382 is a non-synonymous SNP located in exon 23 of the gene and its A to G base change causes a Glu2578Gly substitution. The goodness-of-fit test showed that genotypic distribution of rs1009382 was deviated from Hardy-Weinberg equilibrium due to homozygote excess in the patient group (P = 0.01), suggesting that a double dose of a genetic risk may be involved. Possibly, rs1009382 is a candidate SNP predisposing to a schizophrenic illness. Moreover, the test for linkage disequilibrium (LD) between paired SNPs showed that the nine SNPs studied may be in the same LD block with an unexpected pattern as the strength of LD was not correlated with the distance between paired SNPs. The haplotype analysis suggested that there might be more than one disease-related allele located in the class III region of the MHC, and that these alleles possibly confer either susceptibility or resistance to schizophrenia. PMID:14755442

  6. Candidate colorectal cancer predisposing gene variants in Chinese early-onset and familial cases

    PubMed Central

    Zhang, Jun-Xiao; Fu, Lei; de Voer, Richarda M; Hahn, Marc-Manuel; Jin, Peng; Lv, Chen-Xi; Verwiel, Eugène TP; Ligtenberg, Marjolijn JL; Hoogerbrugge, Nicoline; Kuiper, Roland P; Sheng, Jian-Qiu; Geurts van Kessel, Ad

    2015-01-01

    AIM: To investigate whether whole-exome sequencing may serve as an efficient method to identify known or novel colorectal cancer (CRC) predisposing genes in early-onset or familial CRC cases. METHODS: We performed whole-exome sequencing in 23 Chinese patients from 21 families with non-polyposis CRC diagnosed at ≤ 40 years of age, or from multiple affected CRC families with at least 1 first-degree relative diagnosed with CRC at ≤ 55 years of age. Genomic DNA from blood was enriched for exome sequences using the SureSelect Human All Exon Kit, version 2 (Agilent Technologies) and sequencing was performed on an Illumina HiSeq 2000 platform. Data were processed through an analytical pipeline to search for rare germline variants in known or novel CRC predisposing genes. RESULTS: In total, 32 germline variants in 23 genes were identified and confirmed by Sanger sequencing. In 6 of the 21 families (29%), we identified 7 mutations in 3 known CRC predisposing genes including MLH1 (5 patients), MSH2 (1 patient), and MUTYH (biallelic, 1 patient), five of which were reported as pathogenic. In the remaining 15 families, we identified 20 rare and novel potentially deleterious variants in 19 genes, six of which were truncating mutations. One previously unreported variant identified in a conserved region of EIF2AK4 (p.Glu738_Asp739insArgArg) was found to represent a local Chinese variant, which was significantly enriched in our early-onset CRC patient cohort compared to a control cohort of 100 healthy Chinese individuals scored negative by colonoscopy (33.3% vs 7%, P < 0.001). CONCLUSION: Whole-exome sequencing of early-onset or familial CRC cases serves as an efficient method to identify known and potential pathogenic variants in established and novel candidate CRC predisposing genes. PMID:25892863

  7. Human DNA repair and recombination genes

    SciTech Connect

    Thompson, L.H.; Weber, C.A.; Jones, N.J.

    1988-09-01

    Several genes involved in mammalian DNA repair pathways were identified by complementation analysis and chromosomal mapping based on hybrid cells. Eight complementation groups of rodent mutants defective in the repair of uv radiation damage are now identified. At least seven of these genes are probably essential for repair and at least six of them control the incision step. The many genes required for repair of DNA cross-linking damage show overlap with those involved in the repair of uv damage, but some of these genes appear to be unique for cross-link repair. Two genes residing on human chromosome 19 were cloned from genomic transformants using a cosmid vector, and near full-length cDNA clones of each gene were isolated and sequenced. Gene ERCC2 efficiently corrects the defect in CHO UV5, a nucleotide excision repair mutant. Gene XRCC1 normalizes repair of strand breaks and the excessive sister chromatid exchange in CHO mutant EM9. ERCC2 shows a remarkable /approximately/52% overall homology at both the amino acid and nucleotide levels with the yeast RAD3 gene. Evidence based on mutation induction frequencies suggests that ERCC2, like RAD3, might also be an essential gene for viability. 100 refs., 4 tabs.

  8. Variation in Telangiectasia Predisposing Genes Is Associated With Overall Radiation Toxicity

    SciTech Connect

    Tanteles, George A.; Murray, Robert J.S.; Mills, Jamie; Barwell, Julian; Chakraborti, Prabir; Chan, Steve; Cheung, Kwok-Leung; Ennis, Dawn; Khurshid, Nazish; Lambert, Kelly; Machhar, Rohan; Meisuria, Mitul; Osman, Ahmed; Peat, Irene; Sahota, Harjinder; Woodings, Pamela; Talbot, Christopher J.; and others

    2012-11-15

    Purpose: In patients receiving radiotherapy for breast cancer where the heart is within the radiation field, cutaneous telangiectasiae could be a marker of potential radiation-induced heart disease. We hypothesized that single nucleotide polymorphisms (SNPs) in genes known to cause heritable telangiectasia-associated disorders could predispose to such late, normal tissue vascular damage. Methods and Materials: The relationship between cutaneous telangiectasia as a late normal tissue radiation injury phenotype in 633 breast cancer patients treated with radiotherapy was examined. Patients were clinically assessed for the presence of cutaneous telangiectasia and genotyped at nine SNPs in three candidate genes. Candidate SNPs were within the endoglin (ENG) and activin A receptor, type II-like 1 (ACVRL1) genes, mutations in which cause hereditary hemorrhagic telangiectasia and the ataxia-telangiectasia mutated (ATM) gene associated with ataxia-telangiectasia. Results: A total of 121 (19.1%) patients exhibited a degree of cutaneous telangiectasiae on clinical examination. Regression was used to examine the associations between the presence of telangiectasiae in patients who underwent breast-conserving surgery, controlling for the effects of boost and known brassiere size (n=388), and individual geno- or haplotypes. Inheritance of ACVRL1 SNPs marginally contributed to the risk of cutaneous telangiectasiae. Haplotypic analysis revealed a stronger association between inheritance of a ATM haplotype and the presence of cutaneous telangiectasiae, fibrosis and overall toxicity. No significant association was observed between telangiectasiae and the coinheritance of the candidate ENG SNPs. Conclusions: Genetic variation in the ATM gene influences reaction to radiotherapy through both vascular damage and increased fibrosis. The predisposing variation in the ATM gene will need to be better defined to optimize it as a predictive marker for assessing radiotherapy late effects.

  9. Preferential DNA repair in expressed genes.

    PubMed Central

    Hanawalt, P C

    1987-01-01

    Potentially deleterious alterations to DNA occur nonrandomly within the mammalian genome. These alterations include the adducts produced by many chemical carcinogens, but not the UV-induced cyclobutane pyrimidine dimer, which may be an exception. Recent studies in our laboratory have shown that the excision repair of pyrimidine dimers and certain other lesions is nonrandom in the mammalian genome, exhibiting a distinct preference for actively transcribed DNA sequences. An important consequence of this fact is that mutagenesis and carcinogenesis may be determined in part by the activities of the relevant genes. Repair may also be processive, and a model is proposed in which excision repair is coupled to transcription at the nuclear matrix. Similar but freely diffusing repair complexes may account for the lower overall repair efficiencies in the silent domains of the genome. Risk assessment in relation to chemical carcinogenesis requires assays that determine effective levels of DNA damage for producing malignancy. The existence of nonrandom repair in the genome casts into doubt the reliability of overall indicators of DNA binding and lesion repair for such determinations. Furthermore, some apparent differences between the intragenomic repair heterogeneity in rodent cells and that in human cells mandate a reevaluation of rodent test systems for human risk assessment. Tissue-specific and cell-specific differences in the coordinate regulation of gene expression and DNA repair may account for corresponding differences in the carcinogenic response. Images FIGURE 1. FIGURE 1. PMID:3447906

  10. Targeted gene repair – in the arena

    PubMed Central

    Kmiec, Eric B.

    2003-01-01

    The development of targeted gene repair is under way and, despite some setbacks, shows promise as an alternative form of gene therapy. This approach uses synthetic DNA molecules to activate and direct the cell’s inherent DNA repair systems to correct inborn errors. The progress of this technique and its therapeutic potential are discussed in relation to the treatment of genetic diseases. PMID:12952907

  11. Concepts in Gene Therapy for Cartilage Repair

    PubMed Central

    Steinert, Andre F.; Nöth, Ulrich; Tuan, Rocky S.

    2009-01-01

    Summary Once articular cartilage is injured, it has a very limited capacity for self-repair. Although current surgical therapeutic procedures to cartilage repair are clinically useful, they cannot restore a normal articular surface. Current research offers a growing number of bioactive reagents, including proteins and nucleic acids, that may be used to augment different aspects of the repair process. As these agents are difficult to administer effectively, gene transfer approaches are being developed to provide their sustained synthesis at sites of repair. To augment regeneration of articular cartilage, therapeutic genes can be delivered to the synovium, or directly to the cartilage lesion. Gene delivery to the cells of the synovial lining is generally considered more suitable for chondroprotective approaches, based on the expression of anti-inflammatory mediators. Gene transfer targeted to cartilage defects can be achieved by either direct vector administration to cells located at or surrounding the defects, or by transplantation of genetically modified chondrogenic cells into the defect. Several studies have shown that exogenous cDNAs encoding growth factors can be delivered locally to sites of cartilage damage, where they are expressed at therapeutically relevant levels. Furthermore, data is beginning to emerge indicating, that efficient delivery and expression of these genes is capable of influencing a repair response toward the synthesis of a more hyaline cartilage repair tissue in vivo. This review presents the current status of gene therapy for cartilage healing and highlights some of the remaining challenges. PMID:18313477

  12. A cancer-predisposing "hot spot" mutation of the fumarase gene creates a dominant negative protein.

    PubMed

    Lorenzato, Annalisa; Olivero, Martina; Perro, Mario; Brière, Jean Jacques; Rustin, Pierre; Di Renzo, Maria Flavia

    2008-02-15

    The Fumarase (Fumarate Hydratase, FH) is a tumor suppressor gene whose germline heterozygous mutations predispose to hereditary leiomyomatosis and renal cell cancer (HLRCC). The FH gene encodes an enzyme of the Krebs cycle, functioning as a homotetramer and catalyzing the hydration of fumarate to malate. Among the numerous FH mutations reported so far, the R190H missense mutation is the most frequent in HLRCC patients. Here we show the functional analyses of the R190H, in comparison to the better characterized E319Q mutation. We first expressed wild-type and mutated proteins in FH deficient human skin fibroblasts, using lentiviral vectors. The wild-type transgene was able to restore the FH enzymatic activity in cells, while the R190H- and E319Q-FH were not. More interestingly, when the same transgenes were expressed in normal, FH-proficient cells, only the R190H-FH reduced the endogenous FH enzymatic activity. By enforcing the expression of equal amount of wild-type and R190H-FH in the same cell, we showed that the mutated FH protein directly inhibited enzymatic activity by nearly abrogating the FH homotetramer formation. These data demonstrate the dominant negative effect of the R190H missense mutation in the FH gene and suggest that the FH tumor-suppressing activity might be impaired in cells carrying a heterozygous mutation. PMID:17960613

  13. A Common Variant in the FTO Gene Is Associated with Body Mass Index and Predisposes to Childhood and Adult Obesity

    PubMed Central

    Frayling, Timothy M.; Timpson, Nicholas J.; Weedon, Michael N.; Zeggini, Eleftheria; Freathy, Rachel M.; Lindgren, Cecilia M.; Perry, John R. B.; Elliott, Katherine S.; Lango, Hana; Rayner, Nigel W.; Shields, Beverley; Harries, Lorna W.; Barrett, Jeffrey C.; Ellard, Sian; Groves, Christopher J.; Knight, Bridget; Patch, Ann-Marie; Ness, Andrew R.; Ebrahim, Shah; Lawlor, Debbie A.; Ring, Susan M.; Ben-Shlomo, Yoav; Jarvelin, Marjo-Riitta; Sovio, Ulla; Bennett, Amanda J.; Melzer, David; Ferrucci, Luigi; Loos, Ruth J. F.; Barroso, Inês; Wareham, Nicholas J.; Karpe, Fredrik; Owen, Katharine R.; Cardon, Lon R.; Walker, Mark; Hitman, Graham A.; Palmer, Colin N. A.; Doney, Alex S. F.; Morris, Andrew D.; Smith, George Davey; Hattersley, Andrew T.; McCarthy, Mark I.

    2009-01-01

    Obesity is a serious international health problem that increases the risk of several common diseases. The genetic factors predisposing to obesity are poorly understood. A genome-wide search for type 2 diabetes–susceptibility genes identified a common variant in the FTO (fat mass and obesity associated) gene that predisposes to diabetes through an effect on body mass index (BMI). An additive association of the variant with BMI was replicated in 13 cohorts with 38,759 participants. The 16% of adults who are homozygous for the risk allele weighed about 3 kilograms more and had 1.67-fold increased odds of obesity when compared with those not inheriting a risk allele. This association was observed from age 7 years upward and reflects a specific increase in fat mass. PMID:17434869

  14. Stem Cells and Gene Therapy for Cartilage Repair

    PubMed Central

    Longo, Umile Giuseppe; Petrillo, Stefano; Franceschetti, Edoardo; Berton, Alessandra; Maffulli, Nicola; Denaro, Vincenzo

    2012-01-01

    Cartilage defects represent a common problem in orthopaedic practice. Predisposing factors include traumas, inflammatory conditions, and biomechanics alterations. Conservative management of cartilage defects often fails, and patients with this lesions may need surgical intervention. Several treatment strategies have been proposed, although only surgery has been proved to be predictably effective. Usually, in focal cartilage defects without a stable fibrocartilaginous repair tissue formed, surgeons try to promote a natural fibrocartilaginous response by using marrow stimulating techniques, such as microfracture, abrasion arthroplasty, and Pridie drilling, with the aim of reducing swelling and pain and improving joint function of the patients. These procedures have demonstrated to be clinically useful and are usually considered as first-line treatment for focal cartilage defects. However, fibrocartilage presents inferior mechanical and biochemical properties compared to normal hyaline articular cartilage, characterized by poor organization, significant amounts of collagen type I, and an increased susceptibility to injury, which ultimately leads to premature osteoarthritis (OA). Therefore, the aim of future therapeutic strategies for articular cartilage regeneration is to obtain a hyaline-like cartilage repair tissue by transplantation of tissues or cells. Further studies are required to clarify the role of gene therapy and mesenchimal stem cells for management of cartilage lesions. PMID:22481959

  15. SYN2 is an autism predisposing gene: loss-of-function mutations alter synaptic vesicle cycling and axon outgrowth.

    PubMed

    Corradi, Anna; Fadda, Manuela; Piton, Amélie; Patry, Lysanne; Marte, Antonella; Rossi, Pia; Cadieux-Dion, Maxime; Gauthier, Julie; Lapointe, Line; Mottron, Laurent; Valtorta, Flavia; Rouleau, Guy A; Fassio, Anna; Benfenati, Fabio; Cossette, Patrick

    2014-01-01

    An increasing number of genes predisposing to autism spectrum disorders (ASDs) has been identified, many of which are implicated in synaptic function. This 'synaptic autism pathway' notably includes disruption of SYN1 that is associated with epilepsy, autism and abnormal behavior in both human and mice models. Synapsins constitute a multigene family of neuron-specific phosphoproteins (SYN1-3) present in the majority of synapses where they are implicated in the regulation of neurotransmitter release and synaptogenesis. Synapsins I and II, the major Syn isoforms in the adult brain, display partially overlapping functions and defects in both isoforms are associated with epilepsy and autistic-like behavior in mice. In this study, we show that nonsense (A94fs199X) and missense (Y236S and G464R) mutations in SYN2 are associated with ASD in humans. The phenotype is apparent in males. Female carriers of SYN2 mutations are unaffected, suggesting that SYN2 is another example of autosomal sex-limited expression in ASD. When expressed in SYN2  knockout neurons, wild-type human Syn II fully rescues the SYN2 knockout phenotype, whereas the nonsense mutant is not expressed and the missense mutants are virtually unable to modify the SYN2 knockout phenotype. These results identify for the first time SYN2  as a novel predisposing gene for ASD and strengthen the hypothesis that a disturbance of synaptic homeostasis underlies ASD. PMID:23956174

  16. SYN2 is an autism predisposing gene: loss-of-function mutations alter synaptic vesicle cycling and axon outgrowth

    PubMed Central

    Corradi, Anna; Fadda, Manuela; Piton, Amélie; Patry, Lysanne; Marte, Antonella; Rossi, Pia; Cadieux-Dion, Maxime; Gauthier, Julie; Lapointe, Line; Mottron, Laurent; Valtorta, Flavia; Rouleau, Guy A.; Fassio, Anna; Benfenati, Fabio; Cossette, Patrick

    2014-01-01

    An increasing number of genes predisposing to autism spectrum disorders (ASDs) has been identified, many of which are implicated in synaptic function. This ‘synaptic autism pathway’ notably includes disruption of SYN1 that is associated with epilepsy, autism and abnormal behavior in both human and mice models. Synapsins constitute a multigene family of neuron-specific phosphoproteins (SYN1-3) present in the majority of synapses where they are implicated in the regulation of neurotransmitter release and synaptogenesis. Synapsins I and II, the major Syn isoforms in the adult brain, display partially overlapping functions and defects in both isoforms are associated with epilepsy and autistic-like behavior in mice. In this study, we show that nonsense (A94fs199X) and missense (Y236S and G464R) mutations in SYN2 are associated with ASD in humans. The phenotype is apparent in males. Female carriers of SYN2 mutations are unaffected, suggesting that SYN2 is another example of autosomal sex-limited expression in ASD. When expressed in SYN2  knockout neurons, wild-type human Syn II fully rescues the SYN2 knockout phenotype, whereas the nonsense mutant is not expressed and the missense mutants are virtually unable to modify the SYN2 knockout phenotype. These results identify for the first time SYN2  as a novel predisposing gene for ASD and strengthen the hypothesis that a disturbance of synaptic homeostasis underlies ASD. PMID:23956174

  17. The Two Main Forms of Histiocytic Sarcoma in the Predisposed Flatcoated Retriever Dog Display Variation in Gene Expression

    PubMed Central

    Boerkamp, Kim M.; van Steenbeek, Frank G.; Penning, Louis C.; Groot Koerkamp, Marian J. A.; van Leenen, Dik; Vos-Loohuis, Manon; Grinwis, Guy C. M.; Rutteman, Gerard R.

    2014-01-01

    Examination of gene functions in specific tumor types improves insight in tumorigenesis and helps design better treatments. Due to the rarity of histiocytic/dendritic cell sarcoma in humans, it is difficult to accrue such knowledge. Therefore, comparative research of these cancers in predisposed dog breeds, such as the Flatcoated retriever, can be of value. Histiocytic sarcoma in the dog can be grouped into a soft tissue- and visceral form. The soft tissue form at first is localized, while the visceral form progresses more quickly to a terminal state, which might be related to variations in gene expression. Microarray analyses were performed on fresh-frozen tissue from Flatcoated retrievers with either soft tissue- or visceral histiocytic sarcoma. Expression differences of ten most significantly differentially expressed genes were validated with quantitative real-time PCR (q PCR) analyses. Q PCR analyses confirmed the significantly aberrant expression of three of the selected genes: C6 was up-regulated; CLEC12A and CCL5 were down-regulated in the visceral histiocytic sarcoma compared to the soft tissue form. The findings of our study indicate that these two forms of histiocytic sarcoma in the dog display a variation in gene expression and warrant analysis of functional changes in the expression of those genes in these rare sarcomas in man. PMID:24886914

  18. Germline variants in the SEMA4A gene predispose to familial colorectal cancer type X

    PubMed Central

    Schulz, Eduard; Klampfl, Petra; Holzapfel, Stefanie; Janecke, Andreas R.; Ulz, Peter; Renner, Wilfried; Kashofer, Karl; Nojima, Satoshi; Leitner, Anita; Zebisch, Armin; Wölfler, Albert; Hofer, Sybille; Gerger, Armin; Lax, Sigurd; Beham-Schmid, Christine; Steinke, Verena; Heitzer, Ellen; Geigl, Jochen B.; Windpassinger, Christian; Hoefler, Gerald; Speicher, Michael R.; Richard Boland, C.; Kumanogoh, Atsushi; Sill, Heinz

    2014-01-01

    Familial colorectal cancer type X (FCCTX) is characterized by clinical features of hereditary non-polyposis colorectal cancer with a yet undefined genetic background. Here we identify the SEMA4A p.Val78Met germline mutation in an Austrian kindred with FCCTX, using an integrative genomics strategy. Compared with wild-type protein, SEMA4AV78M demonstrates significantly increased MAPK/Erk and PI3K/Akt signalling as well as cell cycle progression of SEMA4A-deficient HCT-116 colorectal cancer cells. In a cohort of 53 patients with FCCTX, we depict two further SEMA4A mutations, p.Gly484Ala and p.Ser326Phe and the single-nucleotide polymorphism (SNP) p.Pro682Ser. This SNP is highly associated with the FCCTX phenotype exhibiting increased risk for colorectal cancer (OR 6.79, 95% CI 2.63 to 17.52). Our study shows previously unidentified germline variants in SEMA4A predisposing to FCCTX, which has implications for surveillance strategies of patients and their families. PMID:25307848

  19. Isolating human DNA repair genes using rodent-cell mutants

    SciTech Connect

    Thompson, L.H.; Weber, C.A.; Brookman, K.W.; Salazar, E.P.; Stewart, S.A.; Mitchell, D.L.

    1987-03-23

    The DNA repair systems of rodent and human cells appear to be at least as complex genetically as those in lower eukaryotes and bacteria. The use of mutant lines of rodent cells as a means of identifying human repair genes by functional complementation offers a new approach toward studying the role of repair in mutagenesis and carcinogenesis. In each of six cases examined using hybrid cells, specific human chromosomes have been identified that correct CHO cell mutations affecting repair of damage from uv or ionizing radiations. This finding suggests that both the repair genes and proteins may be virtually interchangeable between rodent and human cells. Using cosmid vectors, human repair genes that map to chromosome 19 have cloned as functional sequences: ERCC2 and XRCC1. ERCC1 was found to have homology with the yeast excision repair gene RAD10. Transformants of repair-deficient cell lines carrying the corresponding human gene show efficient correction of repair capacity by all criteria examined. 39 refs., 1 fig., 1 tab.

  20. Low copy number of the salivary amylase gene predisposes to obesity.

    PubMed

    Falchi, Mario; El-Sayed Moustafa, Julia Sarah; Takousis, Petros; Pesce, Francesco; Bonnefond, Amélie; Andersson-Assarsson, Johanna C; Sudmant, Peter H; Dorajoo, Rajkumar; Al-Shafai, Mashael Nedham; Bottolo, Leonardo; Ozdemir, Erdal; So, Hon-Cheong; Davies, Robert W; Patrice, Alexandre; Dent, Robert; Mangino, Massimo; Hysi, Pirro G; Dechaume, Aurélie; Huyvaert, Marlène; Skinner, Jane; Pigeyre, Marie; Caiazzo, Robert; Raverdy, Violeta; Vaillant, Emmanuel; Field, Sarah; Balkau, Beverley; Marre, Michel; Visvikis-Siest, Sophie; Weill, Jacques; Poulain-Godefroy, Odile; Jacobson, Peter; Sjostrom, Lars; Hammond, Christopher J; Deloukas, Panos; Sham, Pak Chung; McPherson, Ruth; Lee, Jeannette; Tai, E Shyong; Sladek, Robert; Carlsson, Lena M S; Walley, Andrew; Eichler, Evan E; Pattou, Francois; Spector, Timothy D; Froguel, Philippe

    2014-05-01

    Common multi-allelic copy number variants (CNVs) appear enriched for phenotypic associations compared to their biallelic counterparts. Here we investigated the influence of gene dosage effects on adiposity through a CNV association study of gene expression levels in adipose tissue. We identified significant association of a multi-allelic CNV encompassing the salivary amylase gene (AMY1) with body mass index (BMI) and obesity, and we replicated this finding in 6,200 subjects. Increased AMY1 copy number was positively associated with both amylase gene expression (P = 2.31 × 10(-14)) and serum enzyme levels (P < 2.20 × 10(-16)), whereas reduced AMY1 copy number was associated with increased BMI (change in BMI per estimated copy = -0.15 (0.02) kg/m(2); P = 6.93 × 10(-10)) and obesity risk (odds ratio (OR) per estimated copy = 1.19, 95% confidence interval (CI) = 1.13-1.26; P = 1.46 × 10(-10)). The OR value of 1.19 per copy of AMY1 translates into about an eightfold difference in risk of obesity between subjects in the top (copy number > 9) and bottom (copy number < 4) 10% of the copy number distribution. Our study provides a first genetic link between carbohydrate metabolism and BMI and demonstrates the power of integrated genomic approaches beyond genome-wide association studies. PMID:24686848

  1. Copy number variations in the NF1 gene region are infrequent and do not predispose to recurrent type-1 deletions.

    PubMed

    Steinmann, Katharina; Kluwe, Lan; Cooper, David N; Brems, Hilde; De Raedt, Thomas; Legius, Eric; Mautner, Viktor-Felix; Kehrer-Sawatzki, Hildegard

    2008-05-01

    Gross deletions of the NF1 gene at 17q11.2 belong to the group of 'genomic disorders' characterized by local sequence architecture that predisposes to genomic rearrangements. Segmental duplications within regions associated with genomic disorders are prone to non-allelic homologous recombination (NAHR), which mediates gross rearrangements. Copy number variants (CNVs) without obvious phenotypic consequences also occur frequently in regions of genomic disorders. In the NF1 gene region, putative CNVs have been reportedly detected by array comparative genomic hybridization (array CGH). These variants include duplications and deletions within the NF1 gene itself (CNV1) and a duplication that encompasses the SUZ12 gene, the distal NF1-REPc repeat and the RHOT1 gene (CNV2). To explore the possibility that these CNVs could have played a role in promoting deletion mutagenesis in type-1 deletions (the most common type of gross NF1 deletion), non-affected transmitting parents of patients with type-1 NF1 deletions were investigated by multiplex ligation-dependent probe amplification (MLPA). However, neither CNV1 nor CNV2 were detected. This would appear to exclude these variants as frequent mediators of NAHR giving rise to type-1 deletions. Using MLPA, we were also unable to confirm CNV1 in healthy controls as previously reported. We conclude that locus-specific techniques should be used to independently confirm putative CNVs, originally detected by array CGH, to avoid false-positive results. In one patient with an atypical deletion, a duplication in the region of CNV2 was noted. This duplication could have occurred concomitantly with the deletion as part of a complex rearrangement or may alternatively have preceded the deletion. PMID:18212816

  2. Disturbance of cardiac gene expression and cardiomyocyte structure predisposes Mecp2-null mice to arrhythmias

    PubMed Central

    Hara, Munetsugu; Takahashi, Tomoyuki; Mitsumasu, Chiaki; Igata, Sachiyo; Takano, Makoto; Minami, Tomoko; Yasukawa, Hideo; Okayama, Satoko; Nakamura, Keiichiro; Okabe, Yasunori; Tanaka, Eiichiro; Takemura, Genzou; Kosai, Ken-ichiro; Yamashita, Yushiro; Matsuishi, Toyojiro

    2015-01-01

    Methyl-CpG-binding protein 2 (MeCP2) is an epigenetic regulator of gene expression that is essential for normal brain development. Mutations in MeCP2 lead to disrupted neuronal function and can cause Rett syndrome (RTT), a neurodevelopmental disorder. Previous studies reported cardiac dysfunction, including arrhythmias in both RTT patients and animal models of RTT. In addition, recent studies indicate that MeCP2 may be involved in cardiac development and dysfunction, but its role in the developing and adult heart remains unknown. In this study, we found that Mecp2-null ESCs could differentiate into cardiomyocytes, but the development and further differentiation of cardiovascular progenitors were significantly affected in MeCP2 deficiency. In addition, we revealed that loss of MeCP2 led to dysregulation of endogenous cardiac genes and myocardial structural alterations, although Mecp2-null mice did not exhibit obvious cardiac functional abnormalities. Furthermore, we detected methylation of the CpG islands in the Tbx5 locus, and showed that MeCP2 could target these sequences. Taken together, these results suggest that MeCP2 is an important regulator of the gene-expression program responsible for maintaining normal cardiac development and cardiomyocyte structure. PMID:26073556

  3. Disturbance of cardiac gene expression and cardiomyocyte structure predisposes Mecp2-null mice to arrhythmias.

    PubMed

    Hara, Munetsugu; Takahashi, Tomoyuki; Mitsumasu, Chiaki; Igata, Sachiyo; Takano, Makoto; Minami, Tomoko; Yasukawa, Hideo; Okayama, Satoko; Nakamura, Keiichiro; Okabe, Yasunori; Tanaka, Eiichiro; Takemura, Genzou; Kosai, Ken-ichiro; Yamashita, Yushiro; Matsuishi, Toyojiro

    2015-01-01

    Methyl-CpG-binding protein 2 (MeCP2) is an epigenetic regulator of gene expression that is essential for normal brain development. Mutations in MeCP2 lead to disrupted neuronal function and can cause Rett syndrome (RTT), a neurodevelopmental disorder. Previous studies reported cardiac dysfunction, including arrhythmias in both RTT patients and animal models of RTT. In addition, recent studies indicate that MeCP2 may be involved in cardiac development and dysfunction, but its role in the developing and adult heart remains unknown. In this study, we found that Mecp2-null ESCs could differentiate into cardiomyocytes, but the development and further differentiation of cardiovascular progenitors were significantly affected in MeCP2 deficiency. In addition, we revealed that loss of MeCP2 led to dysregulation of endogenous cardiac genes and myocardial structural alterations, although Mecp2-null mice did not exhibit obvious cardiac functional abnormalities. Furthermore, we detected methylation of the CpG islands in the Tbx5 locus, and showed that MeCP2 could target these sequences. Taken together, these results suggest that MeCP2 is an important regulator of the gene-expression program responsible for maintaining normal cardiac development and cardiomyocyte structure. PMID:26073556

  4. Detection of a Tumor Suppressor Gene Variant Predisposing to Colorectal Cancer in an 18th Century Hungarian Mummy

    PubMed Central

    Feldman, Michal; Hershkovitz, Israel; Sklan, Ella H.; Kahila Bar-Gal, Gila; Pap, Ildikó; Szikossy, Ildikó; Rosin-Arbesfeld, Rina

    2016-01-01

    Mutations of the Adenomatous polyposis coli (APC) gene are common and strongly associated with the development of colorectal adenomas and carcinomas. While extensively studied in modern populations, reports on visceral tumors in ancient populations are scarce. To the best of our knowledge, genetic characterization of mutations associated with colorectal cancer in ancient specimens has not yet been described. In this study we have sequenced hotspots for mutations in the APC gene isolated from 18th century naturally preserved human Hungarian mummies. While wild type APC sequences were found in two mummies, we discovered the E1317Q missense mutation, known to be a colorectal cancer predisposing mutation, in a large intestine tissue of an 18th century mummy. Our data suggests that this genetic predisposition to cancer already existed in the pre-industrialization era. This study calls for similar investigations of ancient specimens from different periods and geographical locations to be conducted and shared for the purpose of obtaining a larger scale analysis that will shed light on past cancer epidemiology and on cancer evolution. PMID:26863316

  5. Knockout of the TauT Gene Predisposes C57BL/6 Mice to Streptozotocin-Induced Diabetic Nephropathy

    PubMed Central

    Han, Xiaobin; Patters, Andrea B.; Ito, Takashi; Schaffer, Stephen W.; Chesney, Russell W.

    2015-01-01

    Diabetic nephropathy is the leading cause of end stage renal disease in the world. Although tremendous efforts have been made, scientists have yet to identify an ideal animal model that can reproduce the characteristics of human diabetic nephropathy. In this study, we hypothesize that taurine insufficiency is a critical risk factor for development of diabetic nephropathy associated with diabetes mellitus. This hypothesis was tested in vivo in TauT heterozygous (TauT+/-) and homozygous (TauT-/-) knockout in C57BL/6 background mice. We have shown that alteration of the TauT gene (also known as SLC6A6) has a substantial effect on the susceptibility to development of extensive diabetic kidney disease in both TauT+/- and TauT-/-mouse models of diabetes. These animals developed histological changes characteristic of human diabetic nephropathy that included glomerulosclerosis, nodular lesions, arteriosclerosis, arteriolar dilation, and tubulointerstitial fibrosis. Immunohistochemical staining of molecular markers of smooth muscle actin, CD34, Ki67 and collagen IV further confirmed these observations. Our results demonstrated that both homozygous and heterozygous TauT gene deletion predispose C57BL/6 mice to develop end-stage diabetic kidney disease, which closely replicates the pathological features of diabetic nephropathy in human diabetic patients. PMID:25629817

  6. Low-copy repeats at the human VIPR2 gene predispose to recurrent and nonrecurrent rearrangements

    PubMed Central

    Beri, Silvana; Bonaglia, Maria Clara; Giorda, Roberto

    2013-01-01

    Submicroscopic structural variations, including deletions, duplications, inversions and more complex rearrangements, are widespread in normal human genomes. Inverted segmental duplications or highly identical low-copy repeat (LCR) sequences can mediate the formation of inversions and more complex structural rearrangements through non-allelic homologous recombination. In a patient with 7q36 inverted duplication/terminal deletion, we demonstrated the central role of a pair of short inverted LCRs in the vasoactive intestinal peptide receptor gene (VIPR2)-LCRs in generating the rearrangement. We also revealed a relatively common VIPR2-LCR-associated inversion polymorphism disrupting the gene in almost 1% of healthy subjects, and a small number of complex duplications/triplications. In genome-wide studies of several thousand patients, a significant association of rare microduplications with variable size, all involving VIPR2, with schizophrenia was recently described, suggesting that altered vasoactive intestinal peptide signaling is likely implicated in the pathogenesis of schizophrenia. Genetic testing for VIPR2-LCR-associated inversions should be performed on available cohorts of psychiatric patients to evaluate their potential pathogenic role. PMID:23073313

  7. Reelin gene alleles and haplotypes as a factor predisposing to autistic disorder.

    PubMed

    Persico, A M; D'Agruma, L; Maiorano, N; Totaro, A; Militerni, R; Bravaccio, C; Wassink, T H; Schneider, C; Melmed, R; Trillo, S; Montecchi, F; Palermo, M; Pascucci, T; Puglisi-Allegra, S; Reichelt, K L; Conciatori, M; Marino, R; Quattrocchi, C C; Baldi, A; Zelante, L; Gasparini, P; Keller, F

    2001-03-01

    Autistic disorder (MIM 209850) is currently viewed as a neurodevelopmental disease. Reelin plays a pivotal role in the development of laminar structures including the cerebral cortex, hippocampus, cerebellum and of several brainstem nuclei. Neuroanatomical evidence is consistent with Reelin involvement in autistic disorder. In this study, we describe several polymorphisms identified using RNA-SSCP and DNA sequencing. Association and linkage were assessed comparing 95 Italian patients to 186 ethnically-matched controls, and using the transmission/disequilibrium test and haplotype-based haplotype relative risk in 172 complete trios from 165 families collected in Italy and in the USA. Both case-control and family-based analyses yield a significant association between autistic disorder and a polymorphic GGC repeat located immediately 5' of the reelin gene (RELN) ATG initiator codon, as well as with specific haplotypes formed by this polymorphism with two single-base substitutions located in a splice junction in exon 6 and within exon 50. Triplet repeats located in 5' untranslated regions (5'UTRs) are indicative of strong transcriptional regulation. Our findings suggest that longer triplet repeats in the 5'UTR of the RELN gene confer vulnerability to autistic disorder. PMID:11317216

  8. A genome-wide search for genes predisposing to manic-depression, assuming autosomal dominant inheritance

    SciTech Connect

    Coon, H.; Jensen, S.; Hoff, M.; Holik, J.; Plaetke, R.; Reimherr, F.; Wender, P.; Leppert, M.; Byerley, W. )

    1993-06-01

    Manic-depressive illness (MDI), also known as [open quotes]bipolar affective disorder[close quotes], is a common and devastating neuropsychiatric illness. Although pivotal biochemical alterations underlying the disease are unknown, results of family, twin, and adoption studies consistently implicate genetic transmission in the pathogenesis of MDI. In order to carry out linkage analysis, the authors ascertained eight moderately sized pedigrees containing multiple cases of the disease. For a four-allele marker mapping at 5 cM from the disease gene, the pedigree sample has >97% power to detect a dominant allele under genetic homogeneity and has >73% power under 20% heterogeneity. To date, the eight pedigrees have been genotyped with 328 polymorphic DNA loci throughout the genome. When autosomal dominant inheritance was assumed, 273 DNA markers gave lod scores <[minus]2.0 at [theta] = .05, and 4 DNA marker loci yielded lod scores >1 (chromosome 5 -- D5S39, D5S43, and D5S62; chromosome 11 -- D11S85). Of the markers giving lod scores >1, only D5S62 continued to show evidence for linkage when the affected-pedigree-member method was used. The D5S62 locus maps to distal 5q, a region containing neurotransmitter-receptor genes for dopamine, norepinephrine, glutamate, and gamma-aminobutyric acid. Although additional work in this region may be warranted, the linkage results should be interpreted as preliminary data, as 68 unaffected individuals are not past the age of risk. 72 refs., 2 tabs.

  9. A genome-wide search for genes predisposing to manic-depression, assuming autosomal dominant inheritance.

    PubMed Central

    Coon, H; Jensen, S; Hoff, M; Holik, J; Plaetke, R; Reimherr, F; Wender, P; Leppert, M; Byerley, W

    1993-01-01

    Manic-depressive illness (MDI), also known as "bipolar affective disorder," is a common and devastating neuropsychiatric illness. Although pivotal biochemical alterations underlying the disease are unknown, results of family, twin, and adoption studies consistently implicate genetic transmission in the pathogenesis of MDI. In order to carry out linkage analysis, we ascertained eight moderately sized pedigrees containing multiple cases of the disease. For a four-allele marker mapping 5 cM from the disease gene, the pedigree sample has > 97% power to detect a dominant allele under genetic homogeneity and has > 73% power under 20% heterogeneity. To date, the eight pedigrees have been genotyped with 328 polymorphic DNA loci throughout the genome. When autosomal dominant inheritance was assumed, 273 DNA markers gave lod scores < -2.0 at recombination fraction (theta) = .0, 174 DNA loci produced lod scores < -2.0 at theta = .05, and 4 DNA marker loci yielded lod scores > 1 (chromosome 5--D5S39, D5S43, and D5S62; chromosome 11--D11S85). Of the markers giving lod scores > 1, only D5S62 continued to show evidence for linkage when the affected-pedigree-member method was used. The D5S62 locus maps to distal 5q, a region containing neurotransmitter-receptor genes for dopamine, norepinephrine, glutamate, and gamma-aminobutyric acid. Although additional work in this region may be warranted, our linkage results should be interpreted as preliminary data, as 68 unaffected individuals are not past the age of risk. PMID:8503452

  10. Polymorphism in the Serotonin Receptor 2a (HTR2A) Gene as Possible Predisposal Factor for Aggressive Traits

    PubMed Central

    Banlaki, Zsofia; Elek, Zsuzsanna; Nanasi, Tibor; Szekely, Anna; Nemoda, Zsofia; Sasvari-Szekely, Maria; Ronai, Zsolt

    2015-01-01

    Aggressive manifestations and their consequences are a major issue of mankind, highlighting the need for understanding the contributory factors. Still, aggression-related genetic analyses have so far mainly been conducted on small population subsets such as individuals suffering from a certain psychiatric disorder or a narrow-range age cohort, but no data on the general population is yet available. In the present study, our aim was to identify polymorphisms in genes affecting neurobiological processes that might explain some of the inter-individual variation between aggression levels in the non-clinical Caucasian adult population. 55 single nucleotide polymorphisms (SNP) were simultaneously determined in 887 subjects who also filled out the self-report Buss-Perry Aggression Questionnaire (BPAQ). Single marker association analyses between genotypes and aggression scores indicated a significant role of rs7322347 located in the HTR2A gene encoding serotonin receptor 2a following Bonferroni correction for multiple testing (p = 0.0007) both for males and females. Taking the four BPAQ subscales individually, scores for Hostility, Anger and Physical Aggression showed significant association with rs7322347 T allele in themselves, while no association was found with Verbal Aggression. Of the subscales, relationship with rs7322347 was strongest in the case of Hostility, where statistical significance virtually equaled that observed with the whole BPAQ. In conclusion, this is the first study to our knowledge analyzing SNPs in a wide variety of genes in terms of aggression in a large sample-size non-clinical adult population, also describing a novel candidate polymorphism as predisposal to aggressive traits. PMID:25658328

  11. The c.657del5 variant in the NBN gene predisposes to pancreatic cancer.

    PubMed

    Borecka, Marianna; Zemankova, Petra; Lhota, Filip; Soukupova, Jana; Kleiblova, Petra; Vocka, Michal; Soucek, Pavel; Ticha, Ivana; Kleibl, Zdenek; Janatova, Marketa

    2016-08-10

    Pancreatic ductal adenocarcinoma (PDAC) is the sixth most frequent cancer type in the Czech Republic with a poor prognosis that could be improved by an early detection and subsequent surgical treatment combined with chemotherapy. Genetic factors play an important role in PDAC risk. We previously identified one PDAC patient harboring the Slavic founder deleterious mutation c.657del5 in the NBN gene, using a panel next-generation sequencing (NGS). A subsequent analysis of 241 unselected PDAC patients revealed other mutation carriers. The overall frequency of c.657del5 in unselected PDAC patients (5/241; 2.07%) significantly differed from that in non-cancer controls (2/915; 0.2%; P=0.006). The result indicates that the NBN c.657del5 variant represents a novel PDAC-susceptibility allele increasing PDAC risk (OR=9.7; 95% CI: 1.9 to 50.2). The increased risk of PDAC in follow-up recommendations for NBN mutation carriers should be considered if other studies also confirm an increased frequency of c.657del5 carriers in PDAC patients from other populations. PMID:27150568

  12. Significant Evidence of Linkage for a Gene Predisposing to Colorectal Cancer and Multiple Primary Cancers on 22q11

    PubMed Central

    Teerlink, Craig; Nelson, Quentin; Burt, Randall; Cannon-Albright, Lisa

    2014-01-01

    Objectives: The genetic basis of colorectal cancer (CRC) is not completely specified. Part of the difficulty in mapping predisposition genes for CRC may be because of phenotypic heterogeneity. Using data from a population genealogy of Utah record linked to a statewide cancer registry, we identified a subset of CRC cases that exhibited familial clustering in excess of that expected for all CRC cases in general, which may represent a genetically homogeneous subset of CRC. Methods: Using a new familial aggregation method referred to as the subset genealogic index of familiality (subsetGIF), combined with detailed information from a statewide tumor registry, we identified a subset of CRC cases that exhibited excess familial clustering above that expected for CRC: CRC cases who had at least one other primary tumor at a different site. A genome-wide linkage analysis was performed on a set of high-risk CRC pedigrees that included multiple CRC cases with additional primaries to identify evidence for predisposition loci. Results: A total of 13 high-risk CRC pedigrees with multiple CRC cases with other primary cancers were identified. Linkage analysis identified one pedigree with a significant linkage signal at 22q11 (LOD (logarithm (base 10) of odds)=3.39). Conclusions: A predisposition gene or variant for CRC that also predisposes to other primary cancers likely resides on chromosome 22q11. The ability to use statewide population genealogy and tumor registry data was critical to identify an informative subset of CRC cases that is possibly more genetically homogeneous than CRC in general, and may have improved statistical power for predisposition locus identification in this study. PMID:24572700

  13. Cloning a Eukaryotic DNA Glycosylase Repair Gene by the Suppression of a DNA Repair Defect in Escherichia coli

    NASA Astrophysics Data System (ADS)

    Chen, Jin; Derfler, Bruce; Maskati, Azmat; Samson, Leona

    1989-10-01

    If eukaryotic genes could protect bacteria with defects in DNA repair, this effect could be exploited for the isolation of eukaryotic DNA repair genes. We have thus cloned a DNA repair gene from Saccharomyces cerevisiae that directs the synthesis of a DNA glycosylase that specifically releases 3-methyladenine from alkylated DNA and in so doing protects alkylation-sensitive Escherichia coli from killing by methylating agents. The cloned yeast gene was then used to generate a mutant strain of S. cerevisiae that carries a defect in the glycosylase gene and is extremely sensitive to DNA methylation. This approach may allow the isolation of a large number of eukaryotic DNA repair genes.

  14. Polymorphism of Alcohol Metabolizing Gene ADH3 Predisposes to Development of Alcoholic Pancreatitis in North Indian Population

    PubMed Central

    Singh, Divya; Negi, Tajwar S.; Upadhyay, Ghanshyam; Choudhuri, Gourdas

    2015-01-01

    Background and aim: Genetic factors regulating alcohol metabolism could predispose in developing alcoholic pancreatitis (ACP). Studies revealed that alcohol could be metabolized by both ways, oxidative and non-oxidative. The main oxidative pathway includes alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), and cytochrome P450 enzyme. We investigated the association of polymorphisms in these enzymes with the alcoholic pancreatitis in the north Indian population. Method: Patients with alcoholic pancreatitis (ACP; n = 72), tropical calcific pancreatitis (TCP; n = 75), alcoholic controls (AC; n = 40), and healthy controls (HC; n = 100) were included in the study. Blood samples were collected from the subjects in EDTA coated vials. DNA was extracted and genotyping for ADH3, ALDH2, and CYP2E1 was done by PCR-RFLP (polymerase chain reaction—restriction fragment length polymorphism). The products were analyzed by gel electrophoresis. Result: The frequency distribution of ADH3*1/*1 genotype was significantly higher in ACP group (59.7%) compared with TCP (38.7%), HC (42%), and AC (37.5%) and was found to be associated with increased risk of alcoholic pancreatitis. There was no statistically significant difference between the frequency distribution of ADH3*1/*1, ADH3*1/*2, and ADH3*2/*2 genotypes between TCP and HC or healthy alcoholics. ALDH2 gene was monomorphic in our population, and the frequencies for CYP2E1 intron 6 Dra I polymorphism were comparable in all the four groups. Conclusion: This study shows that carriers of ADH3*1/*1 individuals consuming alcohol are at higher risk for alcoholic pancreatitis than those with other genotypes such as ADH3*1/*2 and ADH3*2/*2. PMID:26734614

  15. A Chromosome 8 Gene-Cluster Polymorphism with Low Human Beta-Defensin 2 Gene Copy Number Predisposes to Crohn Disease of the Colon

    PubMed Central

    Fellermann, Klaus; Stange, Daniel E.; Schaeffeler, Elke; Schmalzl, Hartmut; Wehkamp, Jan; Bevins, Charles L.; Reinisch, Walter; Teml, Alexander; Schwab, Matthias; Lichter, Peter; Radlwimmer, Bernhard; Stange, Eduard F.

    2006-01-01

    Defensins are endogenous antimicrobial peptides that protect the intestinal mucosa against bacterial invasion. It has been suggested that deficient defensin expression may underlie the chronic inflammation of Crohn disease (CD). The DNA copy number of the beta-defensin gene cluster on chromosome 8p23.1 is highly polymorphic within the healthy population, which suggests that the defective beta-defensin induction in colonic CD could be due to low beta-defensin–gene copy number. Here, we tested this hypothesis, using genomewide DNA copy number profiling by array-based comparative genomic hybridization and quantitative polymerase-chain-reaction analysis of the human beta-defensin 2 (HBD-2) gene. We showed that healthy individuals, as well as patients with ulcerative colitis, have a median of 4 (range 2–10) HBD-2 gene copies per genome. In a surgical cohort with ileal or colonic CD and in a second large cohort with inflammatory bowel diseases, those with ileal resections/disease exhibited a normal median HBD-2 copy number of 4, whereas those with colonic CD had a median of only 3 copies per genome (P=.008 for the surgical cohort; P=.032 for the second cohort). Overall, the copy number distribution in colonic CD was shifted to lower numbers compared with controls (P=.002 for both the surgical cohort and the cohort with inflammatory bowel diseases). Individuals with ⩽3 copies have a significantly higher risk of developing colonic CD than did individuals with ⩾4 copies (odds ratio 3.06; 95% confidence interval 1.46–6.45). An HBD-2 gene copy number of <4 was associated with diminished mucosal HBD-2 mRNA expression (P=.033). In conclusion, a lower HBD-2 gene copy number in the beta-defensin locus predisposes to colonic CD, most likely through diminished beta-defensin expression. PMID:16909382

  16. Non-functional genes repaired at the RNA level.

    PubMed

    Burger, Gertraud

    2016-01-01

    Genomes and genes continuously evolve. Gene sequences undergo substitutions, deletions or nucleotide insertions; mobile genetic elements invade genomes and interleave in genes; chromosomes break, even within genes, and pieces reseal in reshuffled order. To maintain functional gene products and assure an organism's survival, two principal strategies are used - either repair of the gene itself or of its product. I will introduce common types of gene aberrations and how gene function is restored secondarily, and then focus on systematically fragmented genes found in a poorly studied protist group, the diplonemids. Expression of their broken genes involves restitching of pieces at the RNA-level, and substantial RNA editing, to compensate for point mutations. I will conclude with thoughts on how such a grotesquely unorthodox system may have evolved, and why this group of organisms persists and thrives since tens of millions of years. PMID:27180109

  17. Understanding the Pathogenicity of Noncoding Mismatch Repair Gene Promoter Variants in Lynch Syndrome.

    PubMed

    Liu, Qing; Thompson, Bryony A; Ward, Robyn L; Hesson, Luke B; Sloane, Mathew A

    2016-05-01

    Lynch syndrome is the most common familial cancer condition that mainly predisposes to tumors of the colon and endometrium. Cancer susceptibility is caused by the autosomal dominant inheritance of a loss-of-function mutation or epimutation in one of the DNA mismatch repair (MMR) genes. Cancer risk assessment is often possible with nonsynonymous coding region mutations, but in many cases patients present with DNA sequence changes within noncoding regions, including the promoters, of MMR genes. The pathogenic role of promoter variants, and hence clinical significance, is unclear and this hinders the clinical management of carriers. In this review, we provide an overview of the classification of MMR gene variants, outline the laboratory assays and online resources that can be used to assess the causality of promoter variants in Lynch syndrome, and highlight some of the practical challenges of demonstrating the pathogenicity of these variants. In conclusion, we propose a guide that could be integrated into the current InSiGHT classification scheme to help determine if a MMR gene promoter variant is pathogenic. PMID:26888055

  18. Polymorphisms in DNA repair genes, smoking, and pancreatic adenocarcinoma risk.

    PubMed

    McWilliams, Robert R; Bamlet, William R; Cunningham, Julie M; Goode, Ellen L; de Andrade, Mariza; Boardman, Lisa A; Petersen, Gloria M

    2008-06-15

    Base excision repair and nucleotide excision repair are vital responses to multiple types of DNA damage, including damage from tobacco exposure. Single-nucleotide polymorphisms (SNP) in these pathways may affect DNA repair capacity and therefore influence risk for cancer development. We performed a clinic-based, case-control study comprising 481 consecutive patients with confirmed pancreatic adenocarcinoma and 625 healthy controls. Allele and genotype frequencies for 16 SNPs in DNA repair genes ERCC1, XPD/ERCC2, XPC, XPF/ERCC4, OGG1, and XRCC1 were compared after adjusting for age, sex, and smoking history. Subgroup analysis by sex and smoking history was performed. Carriers of one or two XPF/ERCC4 minor alleles at R415Q had decreased risk of pancreatic adenocarcinoma compared with those who had two major alleles [odds ratio (OR), 0.59; 95% confidence interval (95% CI), 0.40-0.85]. Heavy smokers (>40 pack-years) had increased risk for cancer if they were carriers of at least one minor allele for XPD/ERCC2 at D312N (OR, 2.78; 95% CI, 1.28-6.04) or D711D (OR, 2.19; 95% CI, 1.01-4.73). No other significant differences in risk were identified. Minor alleles in DNA repair genes XPF/ERCC4 and XPD/ERCC2 were associated with altered risk for pancreatic cancer. PMID:18544627

  19. Hodgkin Lymphoma Risk: Role of Genetic Polymorphisms and Gene-Gene Interactions in DNA repair pathways

    PubMed Central

    Monroy, Claudia M.; Cortes, Andrea C.; Lopez, Mirtha; Rourke, Elizabeth; Etzel, Carol J.; Younes, Anas; Strom, Sara S.; El-Zein, Randa

    2011-01-01

    DNA repair variants may play a potentially important role in an individual’s susceptibility to developing cancer. Numerous studies have reported the association between genetic single nucleotide polymorphisms (SNPs) in DNA repair genes and different types of hematologic cancers. However, to date, the effects of such SNPs on modulating Hodgkin Lymphoma (HL) risk have not yet been investigated. We hypothesized that gene-gene interaction between candidate genes in Direct Reversal, Nucleotide excision repair (NER), Base excision repair (BER) and Double strand break (DSB) pathways may contribute to susceptibility to HL. To test this hypothesis, we conducted a study on 200 HL cases and 220 controls to assess associations between HL risk and 21 functional SNPs in DNA repair genes. We evaluated potential gene-gene interactions and the association of multiple polymorphisms in a chromosome region using a multi-analytic strategy combining logistic regression, multi-factor dimensionality reduction and classification and regression tree approaches. We observed that, in combination, allelic variants in the XPC Ala499Val, NBN Glu185Gln, XRCC3 Thr241Me, XRCC1 Arg194Trp and XRCC1 399Gln polymorphisms modify the risk for developing HL. Moreover, the cumulative genetic risk score revealed a significant trend where the risk for developing HL increases as the number of adverse alleles in BER and DSB genes increase. These findings suggest that DNA repair variants in BER and DSB pathways may play an important role in the development of HL. PMID:21374732

  20. DNA Repair Pathway Gene Expression Score Correlates with Repair Proficiency and Tumor Sensitivity to Chemotherapy

    PubMed Central

    Pitroda, Sean. P.; Pashtan, Itai. M.; Logan, Hillary. L.; Budke, Brian; Darga, Thomas E.; Weichselbaum, Ralph. R.; Connell, Philip. P.

    2016-01-01

    Homologous recombination (HR) and non-homologous end joining (NHEJ) are alternative pathways of double-strand DNA break repair. We developed a method to quantify the efficiency of DNA repair pathways in the context of cancer therapy. The Recombination Proficiency Score (RPS) utilizes the expression levels for four genes involved in DNA repair pathway preference (RIF1, PARI, RAD51, and Ku80), such that high expression of these genes yields a low RPS. Carcinoma cells with low RPS exhibit HR suppression and frequent DNA copy number alterations, which are characteristic of error-prone repair processes that arise in HR-deficient backgrounds. The RPS system was clinically validated in patients with breast or non-small cell lung carcinomas (NSCLC). Tumors with low RPS were associated with greater mutagenesis, adverse clinical features, and inferior patient survival rates, suggesting that HR suppression plays a central role in promoting the genomic instability that fuels malignant progression. This adverse prognosis associated with low RPS was diminished if NSCLC patients received adjuvant chemotherapy, suggesting that HR suppression and associated sensitivity to platinum-based drugs counteracts the adverse prognosis associated with low RPS. Therefore, RPS may predict which therapies will be effective for individual patients, thereby enabling more personalized oncology care. PMID:24670686

  1. Mismatch-mediated error prone repair at the Immunoglobulin genes

    PubMed Central

    Chahwan, Richard; Edelmann, Winfried; Scharff, Matthew D; Roa, Sergio

    2011-01-01

    The generation of effective antibodies depends upon somatic hypermutation (SHM) and class-switch recombination (CSR) of antibody genes by activation induced cytidine deaminase (AID) and the subsequent recruitment of error prone base excision and mismatch repair. While AID initiates and is required for SHM, more than half of the base changes that accumulate in V regions are not due to the direct deamination of dC to dU by AID, but rather arise through the recruitment of the mismatch repair complex (MMR) to the U:G mismatch created by AID and the subsequent perversion of mismatch repair from a high fidelity process to one that is very error prone. In addition, the generation of double-strand breaks (DSBs) is essential during CSR, and the resolution of AID-generated mismatches by MMR to promote such DSBs is critical for the efficiency of the process. While a great deal has been learned about how AID and MMR cause hypermutations and DSBs, it is still unclear how the error prone aspect of these processes is largely restricted to antibody genes. The use of knockout models and mice expressing mismatch repair proteins with separation-of-function point mutations have been decisive in gaining a better understanding of the roles of each of the major MMR proteins and providing further insight into how mutation and repair are coordinated. Here, we review the cascade of MMR factors and repair signals that are diverted from their canonical error free role and hijacked by B cells to promote genetic diversification of the Ig locus. This error prone process involves AID as the inducer of enzymatically-mediated DNA mismatches, and a plethora of downstream MMR factors acting as sensors, adaptors and effectors of a complex and tightly regulated process from much of which is not yet well understood. PMID:22100214

  2. N-acetyltransferase 2 (NAT2) gene polymorphism as a predisposing factor for phenytoin intoxication in tuberculous meningitis or tuberculoma patients having seizures - A pilot study

    PubMed Central

    Adole, Prashant S.; Kharbanda, Parampreet S.; Sharma, Sadhna

    2016-01-01

    Background & objectives: Simultaneous administration of phenytoin and isoniazid (INH) in tuberculous meningitis (TBM) or tuberculoma patients with seizures results in higher plasma phenytoin level and thus phenytoin intoxication. N-acetyltransferase 2 (NAT2) enzyme catalyses two acetylation reactions in INH metabolism and NAT2 gene polymorphism leads to slow and rapid acetylators. The present study was aimed to evaluate the effect of allelic variants of N-acetyltransferase 2 (NAT2) gene as a predisposing factor for phenytoin toxicity in patients with TBM or tuberculoma having seizures, and taking INH and phenytoin simultaneously. Methods: Sixty patients with TBM or tuberculoma with seizures and taking INH and phenytoin simultaneously for a minimum period of seven days were included in study. Plasma phenytoin was measured by high performance liquid chromatography. NAT2 gene polymorphism was studied using restriction fragment length polymorphism and allele specific PCR. Results: The patients were grouped into those having phenytoin intoxication and those with normal phenytoin level, and also classified as rapid or slow acetylators by NAT2 genotyping. Genotypic analysis showed that of the seven SNPs (single nucleotide polymorphisms) of NAT2 gene studied, six mutations were found to be associated with phenytoin intoxication. For rs1041983 (C282T), rs1799929 (C481T), rs1799931 (G857A), rs1799930 (G590A), rs1208 (A803G) and rs1801280 (T341C) allelic variants, the proportion of homozygous mutant was higher in phenytoin intoxicated group than in phenytoin non-intoxicated group. Interpretation & conclusions: Homozygous mutant allele of NAT2 gene at 481site may act as a predisposing factor for phenytoin intoxication among TBM or tuberculoma patients having seizures. PMID:27488001

  3. Associations between polymorphisms in DNA repair genes and glioblastoma.

    PubMed

    McKean-Cowdin, Roberta; Barnholtz-Sloan, Jill; Inskip, Peter D; Ruder, Avima M; Butler, Maryann; Rajaraman, Preetha; Razavi, Pedram; Patoka, Joe; Wiencke, John K; Bondy, Melissa L; Wrensch, Margaret

    2009-04-01

    A pooled analysis was conducted to examine the association between select variants in DNA repair genes and glioblastoma multiforme, the most common and deadliest form of adult brain tumors. Genetic data for approximately 1,000 glioblastoma multiforme cases and 2,000 controls were combined from four centers in the United States that have conducted case-control studies on adult glioblastoma multiforme, including the National Cancer Institute, the National Institute for Occupational Safety and Health, the University of Texas M. D. Anderson Cancer Center, and the University of California at San Francisco. Twelve DNA repair single-nucleotide polymorphisms were selected for investigation in the pilot collaborative project. The C allele of the PARP1 rs1136410 variant was associated with a 20% reduction in risk for glioblastoma multiforme (odds ratio(CT or CC), 0.80; 95% confidence interval, 0.67-0.95). A 44% increase in risk for glioblastoma multiforme was found for individuals homozygous for the G allele of the PRKDC rs7003908 variant (odds ratio(GG), 1.44; 95% confidence interval, 1.13-1.84); there was a statistically significant trend (P = 0.009) with increasing number of G alleles. A significant, protective effect was found when three single-nucleotide polymorphisms (ERCC2 rs13181, ERCC1 rs3212986, and GLTSCR1 rs1035938) located near each other on chromosome 19 were modeled as a haplotype. The most common haplotype (AGC) was associated with a 23% reduction in risk (P = 0.03) compared with all other haplotypes combined. Few studies have reported on the associations between variants in DNA repair genes and brain tumors, and few specifically have examined their impact on glioblastoma multiforme. Our results suggest that common variation in DNA repair genes may be associated with risk for glioblastoma multiforme. PMID:19318434

  4. ASSOCIATIONS BETWEEN POLYMORPHISMS IN DNA REPAIR GENES AND GLIOBLASTOMA

    PubMed Central

    McKean-Cowdin, Roberta; Barnholtz-Sloan, Jill; Inskip, Peter; Ruder, Avima; Butler, MaryAnn; Rajaraman, Preetha; Razavi, Pedram; Patoka, Joe; Wiencke, John; Bondy, Melissa; Wrensch, Margaret

    2009-01-01

    A pooled analysis was conducted to examine the association between select variants in DNA repair genes and glioblastoma multiforme (GBM), the most common and deadliest form of adult brain tumors. Genetic data for approximately 1,000 GBM cases and 2,000 controls were combined from four centers in the United States that have conducted case-control studies of adult GBM including the National Cancer Institute, the National Institute for Occupational Safety and Health, the University of Texas M.D. Anderson Cancer Center, and the University of California at San Francisco. Twelve DNA repair SNPs were selected for investigation in the pilot collaborative project. The C allele of the PARP1 rs1136410 variant was associated with a 20% reduction in risk of GBM (ORCT or CC =0.80; 95%CI 0.67–0.95). A 44% increase in risk of GBM was found for individuals homozygous for the G allele of the PRKDC rs7003908 variant (ORGG 1.44; 95%CI 1.13–1.84); there was a statistically significant trend (p=0.009) with increasing number of G alleles. A significant, protective effect was found when 3 SNPs (ERCC2 rs13181, ERCC1 rs3212986, and GLTSCR1 rs1035938) located near each other on chromosome 19 were modeled as a haplotype. The most common haplotype (AGC) was associated with a 23% reduction in risk (p=0.03) compared to all other haplotypes combined. Few studies have reported on the associations between variants in DNA repair genes and brain tumors, and few specifically have examined their impact on GBMs. Our results suggest that common variation in DNA repair genes may be associated with risk of GBMs. PMID:19318434

  5. Non-DBS DNA Repair Genes Regulate Radiation-induced Cytogenetic Damage Repair and Cell Cycle Progression

    NASA Technical Reports Server (NTRS)

    Zhang, Ye; Rohde, Larry H.; Emami, Kamal; Casey, Rachael; Wu, Honglu

    2008-01-01

    Changes of gene expression profile are one of the most important biological responses in living cells after ionizing radiation (IR) exposure. Although some studies have shown that genes up-regulated by IR may play important roles in DNA damage repair, the relationship between the regulation of gene expression by IR, particularly genes not known for their roles in DSB repair, and its impact on cytogenetic responses has not been systematically studied. In the present study, the expression of 25 genes selected on the basis of their transcriptional changes in response to IR was individually knocked down by transfection with small interfering RNA in human fibroblast cells. The purpose of this study is to identify new roles of these selected genes on regulating DSB repair and cell cycle progression , as measured in the micronuclei formation and chromosome aberration. In response to IR, the formation of MN was significantly increased by suppressed expression of 5 genes: Ku70 in the DSB repair pathway, XPA in the NER pathway, RPA1 in the MMR pathway, and RAD17 and RBBP8 in cell cycle control. Knocked-down expression of 4 genes (MRE11A, RAD51 in the DSB pathway, SESN1, and SUMO1) significantly inhibited cell cycle progression, possibly because of severe impairment of DNA damage repair. Furthermore, loss of XPA, P21, or MLH1 expression resulted in both significantly enhanced cell cycle progression and increased yields of chromosome aberrations, indicating that these gene products modulate both cell cycle control and DNA damage repair. Most of the 11 genes that affected cytogenetic responses are not known to have clear roles influencing DBS repair. Nine of these 11 genes were up-regulated in cells exposed to gamma radiation, suggesting that genes transcriptionally modulated by IR were critical to regulate the biological consequences after IR.

  6. A Stem Cell-Like Chromatin Pattern May Predispose Tumor Suppressor Genes to DNA Hypermethylation and Silencing in Adult Cancers

    PubMed Central

    Ohm, Joyce E.; McGarvey, Kelly M.; Yu, Xiaobing; Cheng, Linzhao; Schuebel, Kornel E.; Cope, Leslie; Mohammad, Helai P.; Chen, Wei; Daniel, Vincent C.; Yu, Wayne; Berman, David M.; Jenuwein, Thomas; Pruitt, Kevin; Sharkis, Saul J.; Watkins, D. Neil; Herman, James G.; Baylin, Stephen B.

    2009-01-01

    Adult cancers may derive from stem or early progenitor cells1,2. Epigenetic modulation of gene expression is essential for normal function of these early cells, but is highly abnormal in cancers, which often exhibit aberrant promoter CpG island hypermethylation and transcriptional silencing of tumor suppressor genes and pro-differentiation factors3-5. We find that, for such genes, both normal and malignant embryonic cells generally lack the gene DNA hypermethylation found in adult cancers. In embryonic stem (ES) cells, these genes are held in a “transcription ready” state mediated by a “bivalent” promoter chromatin pattern consisting of the repressive polycomb group (PcG) H3K27me mark plus the active mark, H3K4me. However, embryonic carcinoma (EC) cells add two key repressive marks, H3K9me2 and H3K9me3, both associated with DNA hypermethylated genes in adult cancers6-8. We hypothesize that cell chromatin patterns and transient silencing of these important growth regulatory genes in stem or progenitor cells of origin for cancer may leave these genes vulnerable to aberrant DNA hypermethylation and heritable gene silencing in adult tumors. PMID:17211412

  7. Sensitized phenotypic screening identifies gene dosage sensitive region on chromosome 11 that predisposes to disease in mice

    PubMed Central

    Ermakova, Olga; Piszczek, Lukasz; Luciani, Luisa; Cavalli, Florence M G; Ferreira, Tiago; Farley, Dominika; Rizzo, Stefania; Paolicelli, Rosa Chiara; Al-Banchaabouchi, Mumna; Nerlov, Claus; Moriggl, Richard; Luscombe, Nicholas M; Gross, Cornelius

    2011-01-01

    The identification of susceptibility genes for human disease is a major goal of current biomedical research. Both sequence and structural variation have emerged as major genetic sources of phenotypic variability and growing evidence points to copy number variation as a particularly important source of susceptibility for disease. Here we propose and validate a strategy to identify genes in which changes in dosage alter susceptibility to disease-relevant phenotypes in the mouse. Our approach relies on sensitized phenotypic screening of megabase-sized chromosomal deletion and deficiency lines carrying altered copy numbers of ∼30 linked genes. This approach offers several advantages as a method to systematically identify genes involved in disease susceptibility. To examine the feasibility of such a screen, we performed sensitized phenotyping in five therapeutic areas (metabolic syndrome, immune dysfunction, atherosclerosis, cancer and behaviour) of a 0.8 Mb reciprocal chromosomal duplication and deficiency on chromosome 11 containing 27 genes. Gene dosage in the region significantly affected risk for high-fat diet-induced metabolic syndrome, antigen-induced immune hypersensitivity, ApoE-induced atherosclerosis, and home cage activity. Follow up studies on individual gene knockouts for two candidates in the region showed that copy number variation in Stat5 was responsible for the phenotypic variation in antigen-induced immune hypersensitivity and metabolic syndrome. These data demonstrate the power of sensitized phenotypic screening of segmental aneuploidy lines to identify disease susceptibility genes. PMID:21204268

  8. Approaches to diagnose DNA mismatch repair gene defects in cancer.

    PubMed

    Peña-Diaz, Javier; Rasmussen, Lene Juel

    2016-02-01

    The DNA repair pathway mismatch repair (MMR) is responsible for the recognition and correction of DNA biosynthetic errors caused by inaccurate nucleotide incorporation during replication. Faulty MMR leads to failure to address the mispairs or insertion deletion loops (IDLs) left behind by the replicative polymerases and results in increased mutation load at the genome. The realization that defective MMR leads to a hypermutation phenotype and increased risk of tumorigenesis highlights the relevance of this pathway for human disease. The association of MMR defects with increased risk of cancer development was first observed in colorectal cancer patients that carried inactivating germline mutations in MMR genes and the disease was named as hereditary non-polyposis colorectal cancer (HNPCC). Currently, a growing list of cancers is found to be MMR defective and HNPCC has been renamed Lynch syndrome (LS) partly to include the associated risk of developing extra-colonic cancers. In addition, a number of non-hereditary, mostly epigenetic, alterations of MMR genes have been described in sporadic tumors. Besides conferring a strong cancer predisposition, genetic or epigenetic inactivation of MMR genes also renders cells resistant to some chemotherapeutic agents. Therefore, diagnosis of MMR deficiency has important implications for the management of the patients, the surveillance of their relatives in the case of LS and for the choice of treatment. Some of the alterations found in MMR genes have already been well defined and their pathogenicity assessed. Despite this substantial wealth of knowledge, the effects of a large number of alterations remain uncharacterized (variants of uncertain significance, VUSs). The advent of personalized genomics is likely to increase the list of VUSs found in MMR genes and anticipates the need of diagnostic tools for rapid assessment of their pathogenicity. This review describes current tools and future strategies for addressing the relevance

  9. Regulation of the Saccharomyces cerevisiae DNA repair gene RAD16.

    PubMed Central

    Bang, D D; Timmermans, V; Verhage, R; Zeeman, A M; van de Putte, P; Brouwer, J

    1995-01-01

    The RAD16 gene product has been shown to be essential for the repair of the silenced mating type loci [Bang et al. (1992) Nucleic Acids Res. 20, 3925-3931]. More recently we demonstrated that the RAD16 and RAD7 proteins are also required for repair of non-transcribed strands of active genes in Saccharomyces cerevisiae [Waters et al. (1993) Mol. Gen. Genet. 239, 28-32]. We have studied the regulation of the RAD16 gene and found that the RAD16 transcript levels increased up to 7-fold upon UV irradiation. Heat shock at 42 degrees C also results in elevated levels of RAD16 mRNA. In sporulating MAT alpha/MATa diploid cells RAD16 mRNA is also induced. The basal level of the RAD16 transcript is constant during the mitotic cell cycle. G1-arrested cells show normal induction of RAD16 mRNA upon UV irradiation demonstrating that the induction is not a secondary consequence of G2 cell cycle arrest following UV irradiation. However, in cells arrested in G1 the induction of RAD16 mRNA after UV irradiation is not followed by a rapid decline as occurs in normal growing cells suggesting that the down regulation of RAD16 transcription is dependent on progression into the cell cycle. Images PMID:7784171

  10. Review: Clinical aspects of hereditary DNA Mismatch repair gene mutations.

    PubMed

    Sijmons, Rolf H; Hofstra, Robert M W

    2016-02-01

    Inherited mutations of the DNA Mismatch repair genes MLH1, MSH2, MSH6 and PMS2 can result in two hereditary tumor syndromes: the adult-onset autosomal dominant Lynch syndrome, previously referred to as Hereditary Non-Polyposis Colorectal Cancer (HNPCC) and the childhood-onset autosomal recessive Constitutional Mismatch Repair Deficiency syndrome. Both conditions are important to recognize clinically as their identification has direct consequences for clinical management and allows targeted preventive actions in mutation carriers. Lynch syndrome is one of the more common adult-onset hereditary tumor syndromes, with thousands of patients reported to date. Its tumor spectrum is well established and includes colorectal cancer, endometrial cancer and a range of other cancer types. However, surveillance for cancers other than colorectal cancer is still of uncertain value. Prophylactic surgery, especially for the uterus and its adnexa is an option in female mutation carriers. Chemoprevention of colorectal cancer with aspirin is actively being investigated in this syndrome and shows promising results. In contrast, the Constitutional Mismatch Repair Deficiency syndrome is rare, features a wide spectrum of childhood onset cancers, many of which are brain tumors with high mortality rates. Future studies are very much needed to improve the care for patients with this severe disorder. PMID:26746812

  11. Germ line variation in nucleotide excision repair genes and lung cancer risk in smokers.

    PubMed

    Sakoda, Lori C; Loomis, Melissa M; Doherty, Jennifer A; Julianto, Liberto; Barnett, Matt J; Neuhouser, Marian L; Thornquist, Mark D; Weiss, Noel S; Goodman, Gary E; Chen, Chu

    2012-01-01

    Since nucleotide excision repair (NER) is primarily responsible for detecting and removing bulky DNA lesions induced by tobacco smoke in the respiratory tract, single nucleotide polymorphisms (SNPs) in NER protein-encoding genes may influence lung cancer risk, particularly in smokers. Studies testing this hypothesis have produced inconsistent results, with most analyzing a few SNPs in relatively small population samples. In a study nested in the Beta- Carotene and Retinol Efficacy Trial, we examined 79 tag and previously reported risk-associated SNPs in the ERCC1, ERCC2, ERCC3, ERCC4, ERCC5, LIG1, POLE, XPA, and XPC genes in 744 lung cancer cases and 1,477 controls, all of whom were non-Hispanic white smokers. Using logistic regression, odds ratios (OR) and 95% confidence intervals (95% CI) were calculated to estimate lung cancer risk associated with SNP genotypes and haplotypes, adjusting for case-control matching factors. Lung cancer risk was modestly associated with LIG1 rs156640 (OR per G allele, 1.23; 95% CI, 1.08-1.40), rs156641 (OR per A allele, 1.23; 95% CI, 1.08-1.40), and rs8100261 (OR per A allele, 0.83; 95% CI, 0.76-0.98); XPA rs3176658 (OR per A allele, 0.83; 95% CI, 0.69-1.00); and ERCC2 rs50871 (OR per C allele, 1.15; 95% CI: 1.01-1.30). Associations with LIG1 and XPA, but not ERCC2, haplotypes were found. The results of this study and others suggest that inherited variants in LIG1 and possibly other NER genes may predispose to smoking-related lung cancer. Given that chance likely accounts for one or more of the associations observed, replication of our findings is needed. PMID:22493747

  12. Polymorphism of the DNA Base Excision Repair Genes in Keratoconus

    PubMed Central

    Wojcik, Katarzyna A.; Synowiec, Ewelina; Sobierajczyk, Katarzyna; Izdebska, Justyna; Blasiak, Janusz; Szaflik, Jerzy; Szaflik, Jacek P.

    2014-01-01

    Keratoconus (KC) is a degenerative corneal disorder for which the exact pathogenesis is not yet known. Oxidative stress is reported to be associated with this disease. The stress may damage corneal biomolecules, including DNA, and such damage is primarily removed by base excision repair (BER). Variation in genes encoding BER components may influence the effectiveness of corneal cells to cope with oxidative stress. In the present work we genotyped 5 polymorphisms of 4 BER genes in 284 patients and 353 controls. The A/A genotype of the c.–1370T>A polymorphism of the DNA polymerase γ (POLG) gene was associated with increased occurrence of KC, while the A/T genotype was associated with decreased occurrence of KC. The A/G genotype and the A allele of the c.1196A>G polymorphism of the X-ray repair cross-complementing group 1 (XRCC1) were associated with increased, and the G/G genotype and the G allele, with decreased KC occurrence. Also, the C/T and T as well as C/C genotypes and alleles of the c.580C>T polymorphism of the same gene displayed relationship with KC occurrence. Neither the g.46438521G>C polymorphism of the Nei endonuclease VIII-like 1 (NEIL1) nor the c.2285T>C polymorphism of the poly(ADP-ribose) polymerase-1 (PARP-1) was associated with KC. In conclusion, the variability of the XRCC1 and POLG genes may play a role in KC pathogenesis and determine the risk of this disease. PMID:25356504

  13. Simulated microgravity influenced the expression of DNA damage repair genes

    NASA Astrophysics Data System (ADS)

    Zhang, Meng; Sun, Yeqing; Jiawei, Liu; Wang, Ting

    2016-07-01

    Ionizing radiation and microgravity were considered to be the most important stress factors of space environmental the respective study of the biological effects of the radiation and microgravity carried out earlier, but the interaction of the effects of radiation with microgravity started later, and due to difference of the materials and methods the result of this experiment were not consistent. To further investigate the influence of microgravity on the expression of the radiation damage repair genes, the seed of Arabidopsis (Col) and its gravity-insensitive mutant (PIN2) were exposed to 0.1Gy of the dose of energetic carbon-ion beam radiation (LET = 30KeV / μm), and the germinated seed were than fixed in the 3D random positioning apparatus immediately for a 10-day simulated microgravity. By measuring the deflection angle of root tip and the changes of the expression of Ku70 and RAD51 protein, we investigated the impact of microgravity effect on radiation damage repair systems. The results shown that radiation, microgravity and microgravity with radiation could increase the angle of the root of the Col significantly, but no obvious effect on PIN2 type. The radiation could increase the expression of Ku70 significantly in both Col and PIN2, microgravity does not affect the expression, but the microgravity with radiation could decrease the expression of Ku70. This result shown that the microgravity could influence the radiation damage repair systems in molecular level. Moreover, our findings were important to understand the molecular mechanism of the impact of microgravity effect on radiation damage repair systems in vivo.

  14. Effect of GDNF on depressive-like behavior, spatial learning and key genes of the brain dopamine system in genetically predisposed to behavioral disorders mouse strains.

    PubMed

    Naumenko, Vladimir S; Kondaurova, Elena M; Bazovkina, Daria V; Tsybko, Anton S; Ilchibaeva, Tatyana V; Khotskin, Nikita V; Semenova, Alina A; Popova, Nina K

    2014-11-01

    The effect of glial cell line-derived neurotrophic factor (GDNF) on behavior and brain dopamine system in predisposed to depressive-like behavior ASC (Antidepressant Sensitive Cataleptics) mice in comparison with the parental "nondepressive" CBA mice was studied. In 7days after administration (800ng, i.c.v.) GDNF decreased escape latency time and the path traveled to reach hidden platform in Morris water maze in ASC mice. GDNF enhanced depressive-like behavioral traits in both "nondepressive" CBA and "depressive" ASC mice. In CBA mice, GDNF decreased functional response to agonists of D1 (chloro-APB hydrobromide) and D2 (sumanirole maleate) receptors in tail suspension test, reduced D2 receptor gene expression in the substantia nigra and increased monoamine oxydase A (MAO A) gene expression in the striatum. GDNF increased D1 and D2 receptor genes expression in the nucleus accumbens of ASC mice but failed to alter expression of catechol-O-methyltransferase, dopamine transporter, MAO B and tyrosine hydroxylase genes in both investigated mouse strains. Thus, GDNF produced long-term genotype-dependent effect on behavior and the brain dopamine system. GDNF pretreatment (1) reduced D1 and D2 receptors functional responses and D2 receptor gene expression in s. nigra of CBA mice; (2) increased D1 and D2 receptor genes expression in n. accumbens of ASC mice and (3) improved spatial learning in ASC mice. GDNF enhanced depressive-like behavior both in CBA and ASC mice. The data suggest that genetically defined variance in the cross-talk between GDNF and brain dopamine system contributes to the variability of GDNF-induced responses and might be responsible for controversial GDNF effects. PMID:25101543

  15. Hirudo medicinalis: a platform for investigating genes in neural repair.

    PubMed

    Wang, W Z; Emes, R D; Christoffers, K; Verrall, J; Blackshaw, S E

    2005-03-01

    We have used the nervous system of the medicinal leech as a preparation to study the molecular basis of neural repair. The leech central nervous system, unlike mammalian CNS, can regenerate to restore function, and contains identified nerve cells of known function and connectivity. We have constructed subtractive cDNA probes from whole and regenerating ganglia of the ventral nerve cord and have used these to screen a serotonergic Retzius neuron library. This identifies genes that are regulated as a result of axotomy, and are expressed by the Retzius cell. This approach identifies many genes, both novel and known. Many of the known genes identified have homologues in vertebrates, including man. For example, genes encoding thioredoxin (TRX), Rough Endoplasmic Reticulum Protein 1 (RER-1) and ATP synthase are upregulated at 24 h postinjury in leech nerve cord. To investigate the functional role of regulated genes in neuron regrowth we are using microinjection of antisense oligonucleotides in combination with horseradish peroxidase to knock down expression of a chosen gene and to assess regeneration in single neurons in 3-D ganglion culture. As an example of this approach we describe experiments to microinject antisense oligonucleotide to a leech isoform of the structural protein, Protein 4.1. Our approach thus identifies genes regulated at different times after injury that may underpin the intrinsic ability of leech neurons to survive damage, to initiate regrowth programs and to remake functional connections. It enables us to determine the time course of gene expression in the regenerating nerve cord, and to study the effects of gene knockdown in identified neurons regenerating in defined conditions in culture. PMID:16047550

  16. POLE mutations in families predisposed to cutaneous melanoma.

    PubMed

    Aoude, Lauren G; Heitzer, Ellen; Johansson, Peter; Gartside, Michael; Wadt, Karin; Pritchard, Antonia L; Palmer, Jane M; Symmons, Judith; Gerdes, Anne-Marie; Montgomery, Grant W; Martin, Nicholas G; Tomlinson, Ian; Kearsey, Stephen; Hayward, Nicholas K

    2015-12-01

    Germline mutations in the exonuclease domain of POLE have been shown to predispose to colorectal cancers and adenomas. POLE is an enzyme involved in DNA repair and chromosomal DNA replication. In order to assess whether such mutations might also predispose to cutaneous melanoma, we interrogated whole-genome and exome data from probands of 34 melanoma families lacking pathogenic mutations in known high penetrance melanoma susceptibility genes: CDKN2A, CDK4, BAP1, TERT, POT1, ACD and TERF2IP. We found a novel germline mutation, POLE p.(Trp347Cys), in a 7-case cutaneous melanoma family. Functional assays in S. pombe showed that this mutation led to an increased DNA mutation rate comparable to that seen with a Pol ε mutant with no exonuclease activity. We then performed targeted sequencing of POLE in 1243 cutaneous melanoma cases and found that a further ten probands had novel or rare variants in the exonuclease domain of POLE. Although this frequency is not significantly higher than that in unselected Caucasian controls, we observed multiple cancer types in the melanoma families, suggesting that some germline POLE mutations may predispose to a broad spectrum of cancers, including melanoma. In addition, we found the first mutation outside the exonuclease domain, p.(Gln520Arg), in a family with an extensive history of colorectal cancer. PMID:26251183

  17. An Obesity-Predisposing Variant of the FTO Gene Regulates D2R-Dependent Reward Learning.

    PubMed

    Sevgi, Meltem; Rigoux, Lionel; Kühn, Anne B; Mauer, Jan; Schilbach, Leonhard; Hess, Martin E; Gruendler, Theo O J; Ullsperger, Markus; Stephan, Klaas Enno; Brüning, Jens C; Tittgemeyer, Marc

    2015-09-01

    Variations in the fat mass and obesity-associated (FTO) gene are linked to obesity. However, the underlying neurobiological mechanisms by which these genetic variants influence obesity, behavior, and brain are unknown. Given that Fto regulates D2/3R signaling in mice, we tested in humans whether variants in FTO would interact with a variant in the ANKK1 gene, which alters D2R signaling and is also associated with obesity. In a behavioral and fMRI study, we demonstrate that gene variants of FTO affect dopamine (D2)-dependent midbrain brain responses to reward learning and behavioral responses associated with learning from negative outcome in humans. Furthermore, dynamic causal modeling confirmed that FTO variants modulate the connectivity in a basic reward circuit of meso-striato-prefrontal regions, suggesting a mechanism by which genetic predisposition alters reward processing not only in obesity, but also in other disorders with altered D2R-dependent impulse control, such as addiction. Significance statement: Variations in the fat mass and obesity-associated (FTO) gene are associated with obesity. Here we demonstrate that variants of FTO affect dopamine-dependent midbrain brain responses and learning from negative outcomes in humans during a reward learning task. Furthermore, FTO variants modulate the connectivity in a basic reward circuit of meso-striato-prefrontal regions, suggesting a mechanism by which genetic vulnerability in reward processing can increase predisposition to obesity. PMID:26354923

  18. Strain and cocaine-induced differential opioid gene expression may predispose Lewis but not Fischer rats to escalate cocaine self-administration.

    PubMed

    Valenza, Marta; Picetti, Roberto; Yuferov, Vadim; Butelman, Eduardo R; Kreek, Mary Jeanne

    2016-06-01

    The aim of the present study was to investigate alterations in gene expression of opioid system components induced by extended access (18 h) cocaine self-administration and to determine the impact of genetic background in the vulnerability to escalate cocaine intake. Comparing two inbred rat strains, we previously reported that Lewis rats progressively escalated cocaine consumption compared to Fischer rats, in a new translational model of intravenous cocaine self-administration, which included 14 sessions of 18-h operant sessions in which rats were allowed to select the cocaine unit dose to self-administer. We compare here Fischer and Lewis rats in the gene expression of endogenous opioid peptides (Pomc, Penk, Pdyn) and cognate receptors (Oprm, Oprk and Oprd) in reward-related brain regions, after exposure to either cocaine self-administration or yoked-saline, in the aforementioned translational paradigm. We performed a correlation analysis between the mRNA level, found in the Dorsal Striatum (DS), Nucleus accumbens (NAcc) shell and core respectively, and individual cocaine intake. Our findings show that the gene expression of all the aforementioned opioid genes exhibit strain-dependent differences in the DS, in absence of cocaine exposure. Also, different strain-specific cocaine-induced mRNA expression of Oprm and Oprk was found in DS. Only few differences were found in the ventral parts of the striatum. Moreover, gene expression level of Pdyn, Penk, Oprk, and Oprm in the DS was significantly correlated with cocaine intake only in Fischer rats. Overall, these data shed light on potential genetic differences which may predispose of subjects to initiate and escalate cocaine consumption. PMID:26777278

  19. Peutz-Jeghers families unlinked to STK11/LKB1 gene mutations are highly predisposed to primitive biliary adenocarcinoma

    PubMed Central

    Olschwang, S.; Boisson, C.; Thomas, G.

    2001-01-01

    INTRODUCTION—Germline mutations of the STK11/LKB1 tumour suppressor gene (19p13.3) are responsible for Peutz-Jeghers syndrome (PJS), a rare genetic disorder, which is dominantly inherited. In addition to the typical hamartomatous gastrointestinal polyps and perioral pigmented lesions, PJS is also associated with the development of tumours in various sites. No specific follow up has yet been evaluated for gene carriers. Furthermore, genetic heterogeneity has been reported, which makes genetic counselling difficult.
METHODS—We report here the analysis of the STK11/LKB1 locus in a series of 34 PJS families, combining the search for mutations and rearrangements in the coding sequence, allele specific expression tests, and linkage studies.
RESULTS—Germline deleterious mutation of the STK11/LKB1 gene were identified in 70% of cases. The hypothesis of a second PJS locus was reinforced and PJS families could be divided into two groups on the basis of the presence or absence of an identified STK11/LKB1 alteration. Analysis of clinical data indicates that the cancer associated risk is markedly different in the two groups. PJS patients with no identified STK11/LKB1 mutation are at major risk for proximal biliary adenocarcinoma, an infrequent tumour in the general population.
CONCLUSION—Up to 30% of PJS patients are caused by mutation in an unidentified gene that confers high susceptibility to cancer development.


Keywords: Peutz-Jeghers disease; genetic heterogeneity; cancer predisposition; risk estimation PMID:11389158

  20. The Contribution of Whole Gene Deletions and Large Rearrangements to the Mutation Spectrum in Inherited Tumor Predisposing Syndromes.

    PubMed

    Smith, Miriam J; Urquhart, Jill E; Harkness, Elaine F; Miles, Emma K; Bowers, Naomi L; Byers, Helen J; Bulman, Michael; Gokhale, Carolyn; Wallace, Andrew J; Newman, William G; Evans, D Gareth

    2016-03-01

    Heterozygous whole gene deletions (WGDs), and intragenic microdeletions, account for a significant proportion of mutations underlying cancer predisposition syndromes. We analyzed the frequency and genotype-phenotype correlations of microdeletions in 12 genes (BRCA1, BRCA2, TP53, MSH2, MLH1, MSH6, PMS2, NF1, NF2, APC, PTCH1, and VHL) representing seven tumor predisposition syndromes in 5,897 individuals (2,611 families) from our center. Overall, microdeletions accounted for 14% of identified mutations. As expected, smaller deletions or duplications were more common (12%) than WGDs (2.2%). Where a WGD was identified in the germline in NF2, the mechanism of somatic second hit was not deletion, as previously described for NF1. For neurofibromatosis type 1 and 2, we compared the mechanism of germline deletion. Unlike NF1, where three specific deletion sizes account for most germline WGDs, NF2 deletion breakpoints were different across seven samples tested. One of these deletions was 3.93 Mb and conferred a severe phenotype, thus refining the region for a potential NF2 modifier gene to a 2.04-Mb region on chromosome 22. The milder phenotype of NF2 WGDs may be due to the apparent absence of chromosome 22 loss as the second hit. These observations of WGD phenotypes will be helpful for interpreting incidental findings from microarray analysis and next-generation sequencing. PMID:26615784

  1. Muir-Torre Syndrome and founder mismatch repair gene mutations: A long gone historical genetic challenge.

    PubMed

    Ponti, G; Manfredini, M; Tomasi, A; Pellacani, G

    2016-09-10

    A "cancer predisposing syndrome" later labeled as Hereditary Non-Polyposis Colorectal Cancer (HNPCC) or Lynch Syndrome, was firstly described by Warthin, about one century ago. An increased predisposition to the development of multiple familial tumors is described as characteristic of this syndrome where visceral and cutaneous malignancies may appear at an early age namely endometrial, gastric, small bowel, ureteral and renal pelvis, ovarian, hepatobiliary tract, pancreatic, brain (Turcot Syndrome) and sebaceous glands (Muir-Torre Syndrome). The latter, a variant of Lynch Syndrome, is characterized by the presence of sebaceous skin adenomas, carcinomas and/or keratoacanthomas associated with visceral malignancies. Both Lynch Syndrome and Muir-Torre Syndrome have been recognized due to germline mutations in mismatch repair genes MLH1, MSH2 and MSH6. To date, 56 Lynch Syndrome founder mutations dependent on MLH1, MSH2 and, although less frequently found, MSH6 and PMS2 are described. Some of these founder mutations, principally of MSH2 gene, have been described to cause Muir-Torre phenotype and have been traced in large and outbreed Muir-Torre Syndrome families living in different US and European territories. Due to the evidences of highly specific Muir-Torre phenotypes related to the presence of widespread MSH2 founder mutations, preliminary search for these MSH2 common mutations in individuals carrying sebaceous tumors and/or keratoacanthomas, at early age or in association to visceral and familial tumors, permits cost-effective and time-saving diagnostic strategies for Lynch/Muir-Torre Syndromes. PMID:26143115

  2. Whole exome sequencing in families at high risk for Hodgkin lymphoma: identification of a predisposing mutation in the KDR gene.

    PubMed

    Rotunno, Melissa; McMaster, Mary L; Boland, Joseph; Bass, Sara; Zhang, Xijun; Burdett, Laurie; Hicks, Belynda; Ravichandran, Sarangan; Luke, Brian T; Yeager, Meredith; Fontaine, Laura; Hyland, Paula L; Goldstein, Alisa M; Chanock, Stephen J; Caporaso, Neil E; Tucker, Margaret A; Goldin, Lynn R

    2016-07-01

    Hodgkin lymphoma shows strong familial aggregation but no major susceptibility genes have been identified to date. The goal of this study was to identify high-penetrance variants using whole exome sequencing in 17 Hodgkin lymphoma prone families with three or more affected cases or obligate carriers (69 individuals), followed by targeted sequencing in an additional 48 smaller HL families (80 individuals). Alignment and variant calling were performed using standard methods. Dominantly segregating, rare, coding or potentially functional variants were further prioritized based on predicted deleteriousness, conservation, and potential importance in lymphoid malignancy pathways. We selected 23 genes for targeted sequencing. Only the p.A1065T variant in KDR (kinase insert domain receptor) also known as VEGFR2 (vascular endothelial growth factor receptor 2) was replicated in two independent Hodgkin lymphoma families. KDR is a type III receptor tyrosine kinase, the main mediator of vascular endothelial growth factor induced proliferation, survival, and migration. Its activity is associated with several diseases including lymphoma. Functional experiments have shown that p.A1065T, located in the activation loop, can promote constitutive autophosphorylation on tyrosine in the absence of vascular endothelial growth factor and that the kinase activity was abrogated after exposure to kinase inhibitors. A few other promising mutations were identified but appear to be "private". In conclusion, in the largest sequenced cohort of Hodgkin lymphoma families to date, we identified a causal mutation in the KDR gene. While independent validation is needed, this mutation may increase downstream tumor cell proliferation activity and might be a candidate for targeted therapy. PMID:27365461

  3. Targeted DNA methylation by homology-directed repair in mammalian cells. Transcription reshapes methylation on the repaired gene

    PubMed Central

    Morano, Annalisa; Angrisano, Tiziana; Russo, Giusi; Landi, Rosaria; Pezone, Antonio; Bartollino, Silvia; Zuchegna, Candida; Babbio, Federica; Bonapace, Ian Marc; Allen, Brittany; Muller, Mark T.; Chiariotti, Lorenzo; Gottesman, Max E.; Porcellini, Antonio; Avvedimento, Enrico V.

    2014-01-01

    We report that homology-directed repair of a DNA double-strand break within a single copy Green Fluorescent Protein (GFP) gene in HeLa cells alters the methylation pattern at the site of recombination. DNA methyl transferase (DNMT)1, DNMT3a and two proteins that regulate methylation, Np95 and GADD45A, are recruited to the site of repair and are responsible for selective methylation of the promoter-distal segment of the repaired DNA. The initial methylation pattern of the locus is modified in a transcription-dependent fashion during the 15–20 days following repair, at which time no further changes in the methylation pattern occur. The variation in DNA modification generates stable clones with wide ranges of GFP expression. Collectively, our data indicate that somatic DNA methylation follows homologous repair and is subjected to remodeling by local transcription in a discrete time window during and after the damage. We propose that DNA methylation of repaired genes represents a DNA damage code and is source of variation of gene expression. PMID:24137009

  4. ApcMin, A Mutation in the Murine Apc Gene, Predisposes to Mammary Carcinomas and Focal Alveolar Hyperplasias

    NASA Astrophysics Data System (ADS)

    Moser, Amy Rapaich; Mattes, Ellen M.; Dove, William F.; Lindstrom, Mary J.; Haag, Jill D.; Gould, Michael N.

    1993-10-01

    ApcMin (Min, multiple intestinal neoplasia) is a point mutation in the murine homolog of the APC gene. Min/+ mice develop multiple intestinal adenomas, as do humans carrying germ-line mutations in APC. Female mice carrying Min are also prone to develop mammary tumors. Min/+ mammary glands are more sensitive to chemical carcinogenesis than are +/+ mammary glands. Transplantation of mammary cells from Min/+ or +/+ donors into +/+ hosts demonstrates that the propensity to develop mammary tumors is intrinsic to the Min/+ mammary cells. Long-term grafts of Min/+ mammary glands also gave rise to focal alveolar hyperplasias, indicating that the presence of the Min mutation also has a role in the development of these lesions.

  5. Glucocorticoid-regulated gene expression during cutaneous wound repair.

    PubMed

    Beer, H D; Fässler, R; Werner, S

    2000-01-01

    Glucocorticoids exert a deleterious effect on the wound healing process, which has been suggested to result from the anti-inflammatory action of these steroids. In addition, recent studies have demonstrated that glucocorticoids regulate the expression of various genes at the wound site which are likely to encode key players in the wound repair process. Using a murine full-thickness excisional wound healing model, we analyzed the effect of dexamethasone on the expression of various cytokines, growth factors, enzymes, and extracellular matrix molecules in normal and wounded skin. We demonstrate that the proinflammatory cytokines interleukin-1 alpha and -beta, tumor necrosis factor alpha, keratinocyte growth factor, transforming growth factors beta 1, beta 2, and beta 3 and their receptors, platelet-derived growth factors and their receptors, tenascin-C, stromelysin-2, macrophage metalloelastase, and enzymes involved in the generation of nitric oxide are targets of glucocorticoid action in wounded skin. These results indicate that anti-inflammatory steroids inhibit wound repair at least in part by influencing the expression of these key regulatory molecules. PMID:10714241

  6. Energy and Technology Review: Unlocking the mysteries of DNA repair

    SciTech Connect

    Quirk, W.A.

    1993-04-01

    DNA, the genetic blueprint, has the remarkable property of encoding its own repair following diverse types of structural damage induced by external agents or normal metabolism. We are studying the interplay of DNA damaging agents, repair genes, and their protein products to decipher the complex biochemical pathways that mediate such repair. Our research focuses on repair processes that correct DNA damage produced by chemical mutagens and radiation, both ionizing and ultraviolet. The most important type of DNA repair in human cells is called excision repair. This multistep process removes damaged or inappropriate pieces of DNA -- often as a string of 29 nucleotides containing the damage -- and replaces them with intact ones. We have isolated, cloned, and mapped several human repair genes associated with the nucleotide excision repair pathway and involved in the repair of DNA damage after exposure to ultraviolet light or mutagens in cooked food. We have shown that a defect in one of these repair genes, ERCC2, is responsible for the repair deficiency in one of the groups of patients with the recessive genetic disorder xeroderma pigmentosum (XP group D). We are exploring ways to purify sufficient quantities (milligrams) of the protein products of these and other repair genes so that we can understand their functions. Our long-term goals are to link defective repair proteins to human DNA repair disorders that predispose to cancer, and to produce DNA-repair-deficient mice that can serve as models for the human disorders.

  7. Lifespan and Stress Resistance in Drosophila with Overexpressed DNA Repair Genes.

    PubMed

    Shaposhnikov, Mikhail; Proshkina, Ekaterina; Shilova, Lyubov; Zhavoronkov, Alex; Moskalev, Alexey

    2015-01-01

    DNA repair declines with age and correlates with longevity in many animal species. In this study, we investigated the effects of GAL4-induced overexpression of genes implicated in DNA repair on lifespan and resistance to stress factors in Drosophila melanogaster. Stress factors included hyperthermia, oxidative stress, and starvation. Overexpression was either constitutive or conditional and either ubiquitous or tissue-specific (nervous system). Overexpressed genes included those involved in recognition of DNA damage (homologs of HUS1, CHK2), nucleotide and base excision repair (homologs of XPF, XPC and AP-endonuclease-1), and repair of double-stranded DNA breaks (homologs of BRCA2, XRCC3, KU80 and WRNexo). The overexpression of different DNA repair genes led to both positive and negative effects on lifespan and stress resistance. Effects were dependent on GAL4 driver, stage of induction, sex, and role of the gene in the DNA repair process. While the constitutive/neuron-specific and conditional/ubiquitous overexpression of DNA repair genes negatively impacted lifespan and stress resistance, the constitutive/ubiquitous and conditional/neuron-specific overexpression of Hus1, mnk, mei-9, mus210, and WRNexo had beneficial effects. This study demonstrates for the first time the effects of overexpression of these DNA repair genes on both lifespan and stress resistance in D. melanogaster. PMID:26477511

  8. Lifespan and Stress Resistance in Drosophila with Overexpressed DNA Repair Genes

    PubMed Central

    Shaposhnikov, Mikhail; Proshkina, Ekaterina; Shilova, Lyubov; Zhavoronkov, Alex; Moskalev, Alexey

    2015-01-01

    DNA repair declines with age and correlates with longevity in many animal species. In this study, we investigated the effects of GAL4-induced overexpression of genes implicated in DNA repair on lifespan and resistance to stress factors in Drosophila melanogaster. Stress factors included hyperthermia, oxidative stress, and starvation. Overexpression was either constitutive or conditional and either ubiquitous or tissue-specific (nervous system). Overexpressed genes included those involved in recognition of DNA damage (homologs of HUS1, CHK2), nucleotide and base excision repair (homologs of XPF, XPC and AP-endonuclease-1), and repair of double-stranded DNA breaks (homologs of BRCA2, XRCC3, KU80 and WRNexo). The overexpression of different DNA repair genes led to both positive and negative effects on lifespan and stress resistance. Effects were dependent on GAL4 driver, stage of induction, sex, and role of the gene in the DNA repair process. While the constitutive/neuron-specific and conditional/ubiquitous overexpression of DNA repair genes negatively impacted lifespan and stress resistance, the constitutive/ubiquitous and conditional/neuron-specific overexpression of Hus1, mnk, mei-9, mus210, and WRNexo had beneficial effects. This study demonstrates for the first time the effects of overexpression of these DNA repair genes on both lifespan and stress resistance in D. melanogaster. PMID:26477511

  9. Dystrophin Gene Replacement and Gene Repair Therapy for Duchenne Muscular Dystrophy in 2016: An Interview.

    PubMed

    Duan, Dongsheng

    2016-03-01

    After years of relentless efforts, gene therapy has now begun to deliver its therapeutic promise in several diseases. A number of gene therapy products have received regulatory approval in Europe and Asia. Duchenne muscular dystrophy (DMD) is an X-linked inherited lethal muscle disease. It is caused by mutations in the dystrophin gene. Replacing and/or repairing the mutated dystrophin gene holds great promises to treated DMD at the genetic level. Last several years have evidenced significant developments in preclinical experimentations in murine and canine models of DMD. There has been a strong interest in moving these promising findings to clinical trials. In light of rapid progress in this field, the Parent Project Muscular Dystrophy (PPMD) recently interviewed me on the current status of DMD gene therapy and readiness for clinical trials. Here I summarized the interview with PPMD. PMID:27003751

  10. Polymorphisms of nucleotide excision repair genes predict melanoma survival.

    PubMed

    Li, Chunying; Yin, Ming; Wang, Li-E; Amos, Christopher I; Zhu, Dakai; Lee, Jeffrey E; Gershenwald, Jeffrey E; Grimm, Elizabeth A; Wei, Qingyi

    2013-07-01

    Melanoma is the most highly malignant skin cancer, and nucleotide excision repair (NER) is involved in melanoma susceptibility. In this analysis of 1,042 melanoma patients, we evaluated whether genetic variants of NER genes may predict survival outcome of melanoma patients. We used genotyping data of 74 tagging single-nucleotide polymorphisms (tagSNPs) in eight core NER genes from our genome-wide association study (including two in XPA, 14 in XPC, three in XPE, four in ERCC1, 10 in ERCC2, eight in ERCC3, 14 in ERCC4, and 19 in ERCC5) and evaluated their associations with prognosis of melanoma patients. Using the Cox proportional hazards model and Kaplan-Meier analysis, we found a predictive role of XPE rs28720291, ERCC5 rs4150314, XPC rs2470458, and ERCC2 rs50871 SNPs in the prognosis of melanoma patients (rs28720291: AG vs. GG, adjusted hazard ratio (adjHR)=11.2, 95% confidence interval (CI) 3.04-40.9, P=0.0003; rs4150314: AG vs. GG, adjHR=4.76, 95% CI 1.09-20.8, P=0.038; rs2470458: AA vs. AG/GG, adjHR=2.11, 95% CI 1.03-4.33, P=0.040; and rs50871: AA vs. AC/CC adjHR=2.27, 95% CI 1.18-4.35, P=0.015). Patients with an increasing number of unfavorable genotypes had markedly increased death risk. Genetic variants of NER genes, particularly XPE rs28720291, ERCC5 rs4150314, XPC rs2470458, and ERCC2 rs50871, may independently or jointly modulate survival outcome of melanoma patients. Because our results were based on a median follow-up of 3 years without multiple test corrections, additional large prospective studies are needed to confirm our findings. PMID:23407396

  11. Candidate DNA repair susceptibility genes identified by exome sequencing in high-risk pancreatic cancer.

    PubMed

    Smith, Alyssa L; Alirezaie, Najmeh; Connor, Ashton; Chan-Seng-Yue, Michelle; Grant, Robert; Selander, Iris; Bascuñana, Claire; Borgida, Ayelet; Hall, Anita; Whelan, Thomas; Holter, Spring; McPherson, Treasa; Cleary, Sean; Petersen, Gloria M; Omeroglu, Atilla; Saloustros, Emmanouil; McPherson, John; Stein, Lincoln D; Foulkes, William D; Majewski, Jacek; Gallinger, Steven; Zogopoulos, George

    2016-01-28

    The genetic basis underlying the majority of hereditary pancreatic adenocarcinoma (PC) is unknown. Since DNA repair genes are widely implicated in gastrointestinal malignancies, including PC, we hypothesized that there are novel DNA repair PC susceptibility genes. As germline DNA repair gene mutations may lead to PC subtypes with selective therapeutic responses, we also hypothesized that there is an overall survival (OS) difference in mutation carriers versus non-carriers. We therefore interrogated the germline exomes of 109 high-risk PC cases for rare protein-truncating variants (PTVs) in 513 putative DNA repair genes. We identified PTVs in 41 novel genes among 36 kindred. Additional genetic evidence for causality was obtained for 17 genes, with FAN1, NEK1 and RHNO1 emerging as the strongest candidates. An OS difference was observed for carriers versus non-carriers of PTVs with early stage (≤IIB) disease. This adverse survival trend in carriers with early stage disease was also observed in an independent series of 130 PC cases. We identified candidate DNA repair PC susceptibility genes and suggest that carriers of a germline PTV in a DNA repair gene with early stage disease have worse survival. PMID:26546047

  12. Expression profiles of DNA repair-related genes in rat target organs under subchronic cadmium exposure.

    PubMed

    Lei, Y X; Lu, Q; Shao, C; He, C C; Lei, Z N; Lian, Y Y

    2015-01-01

    We aimed to evaluate the toxicity of long-term exposure to different cadmium (Cd) doses in rats and expression profiles of DNA repair-related genes. The model rats were exposed to different concentrations of CdCl2 for 3 months, and 5 DNA repair-related genes - hMSH2, MLH1, XRCC1, hOGG1, ERCC1 - were cloned in different tissues, including the liver, kidney, heart, and lung. Accumulated amounts of Cd were detected in the tissues. Gene and protein detections were conducted via fluorescence quantitative real-time polymerase chain reaction and Western blotting, respectively. Methylated sequences of the 5 DNA repair-related gene promoters were used to investigate whether the low expression levels of the genes were related to methylation of the promoter. In the Cd-exposed group, 3 DNA repair genes (i.e., XRCC1, hOGG1, and ERCC1) significantly decreased in the rat liver, kidney, heart, and lung according to the β-actin internal standard (P < 0.01). Western blotting indicated the same trend for the different tissues. Each of the DNA repair genes had special characteristics; for example, hOGG1 gene expression decreased by 75% in the kidney, and XRCC1 gene expression decreased by 5% in the liver and heart when compared to the control group (P < 0.01). A negative correlation between the DNA repair gene expression levels and the cumulative levels of Cd was also suggested by malignancy pathology. The expression levels of 3 DNA repair genes (i.e., ERCC1, XRCC1, and hOGG1) played an important role in the rat response to Cd exposure but not DNA methylated protection. PMID:25729986

  13. Genes and Junk in Plant Mitochondria—Repair Mechanisms and Selection

    PubMed Central

    Christensen, Alan C.

    2014-01-01

    Plant mitochondrial genomes have very low mutation rates. In contrast, they also rearrange and expand frequently. This is easily understood if DNA repair in genes is accomplished by accurate mechanisms, whereas less accurate mechanisms including nonhomologous end joining or break-induced replication are used in nongenes. An important question is how different mechanisms of repair predominate in coding and noncoding DNA, although one possible mechanism is transcription-coupled repair (TCR). This work tests the predictions of TCR and finds no support for it. Examination of the mutation spectra and rates in genes and junk reveals what DNA repair mechanisms are available to plant mitochondria, and what selective forces act on the repair products. A model is proposed that mismatches and other DNA damages are repaired by converting them into double-strand breaks (DSBs). These can then be repaired by any of the DSB repair mechanisms, both accurate and inaccurate. Natural selection will eliminate coding regions repaired by inaccurate mechanisms, accounting for the low mutation rates in genes, whereas mutations, rearrangements, and expansions generated by inaccurate repair in noncoding regions will persist. Support for this model includes the structure of the mitochondrial mutS homolog in plants, which is fused to a double-strand endonuclease. The model proposes that plant mitochondria do not distinguish a damaged or mismatched DNA strand from the undamaged strand, they simply cut both strands and perform homology-based DSB repair. This plant-specific strategy for protecting future generations from mitochondrial DNA damage has the side effect of genome expansions and rearrangements. PMID:24904012

  14. Genomic survey and expression analysis of DNA repair genes in the genus Leptospira.

    PubMed

    Martins-Pinheiro, Marinalva; Schons-Fonseca, Luciane; da Silva, Josefa B; Domingos, Renan H; Momo, Leonardo Hiroyuki Santos; Simões, Ana Carolina Quirino; Ho, Paulo Lee; da Costa, Renata M A

    2016-04-01

    Leptospirosis is an emerging zoonosis with important economic and public health consequences and is caused by pathogenic leptospires. The genus Leptospira belongs to the order Spirochaetales and comprises saprophytic (L. biflexa), pathogenic (L. interrogans) and host-dependent (L. borgpetersenii) members. Here, we present an in silico search for DNA repair pathways in Leptospira spp. The relevance of such DNA repair pathways was assessed through the identification of mRNA levels of some genes during infection in animal model and after exposition to spleen cells. The search was performed by comparison of available Leptospira spp. genomes in public databases with known DNA repair-related genes. Leptospires exhibit some distinct and unexpected characteristics, for instance the existence of a redundant mechanism for repairing a chemically diverse spectrum of alkylated nucleobases, a new mutS-like gene and a new shorter version of uvrD. Leptospira spp. shares some characteristics from Gram-positive, as the presence of PcrA, two RecQ paralogs and two SSB proteins; the latter is considered a feature shared by naturally competent bacteria. We did not find a significant reduction in the number of DNA repair-related genes in both pathogenic and host-dependent species. Pathogenic leptospires were enriched for genes dedicated to base excision repair and non-homologous end joining. Their evolutionary history reveals a remarkable importance of lateral gene transfer events for the evolution of the genus. Up-regulation of specific DNA repair genes, including components of SOS regulon, during infection in animal model validates the critical role of DNA repair mechanisms for the complex interplay between host/pathogen. PMID:26527082

  15. Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene

    PubMed Central

    Hollywood, Jennifer A.; Lee, Ciaran M.; Scallan, Martina F.; Harrison, Patrick T.

    2016-01-01

    To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing to characterise repair tracts from a donor plasmid containing seven nucleotide differences across a 216 bp target region in the human CFTR gene. We found that 90% of the template-dependent repair tracts were >100 bp in length with equal numbers of uni-directional and bi-directional repair tracts. The occurrence of long repair tracts suggests that a single gRNA could be used with variants of the same template to create or correct specific mutations within a 200 bp range, the size of ~80% of human exons. The selection-free strategy used here also allowed detection of non-homologous end joining events in many of the homology-directed repair tracts. This indicates a need to modify the donor, possibly by silent changes in the PAM sequence, to prevent creation of a second double-stranded break in an allele that has already been correctly edited by homology-directed repair. PMID:27557525

  16. Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene.

    PubMed

    Hollywood, Jennifer A; Lee, Ciaran M; Scallan, Martina F; Harrison, Patrick T

    2016-01-01

    To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing to characterise repair tracts from a donor plasmid containing seven nucleotide differences across a 216 bp target region in the human CFTR gene. We found that 90% of the template-dependent repair tracts were >100 bp in length with equal numbers of uni-directional and bi-directional repair tracts. The occurrence of long repair tracts suggests that a single gRNA could be used with variants of the same template to create or correct specific mutations within a 200 bp range, the size of ~80% of human exons. The selection-free strategy used here also allowed detection of non-homologous end joining events in many of the homology-directed repair tracts. This indicates a need to modify the donor, possibly by silent changes in the PAM sequence, to prevent creation of a second double-stranded break in an allele that has already been correctly edited by homology-directed repair. PMID:27557525

  17. The effect of acute dose charge particle radiation on expression of DNA repair genes in mice.

    PubMed

    Tariq, Muhammad Akram; Soedipe, Ayodotun; Ramesh, Govindarajan; Wu, Honglu; Zhang, Ye; Shishodia, Shishir; Gridley, Daila S; Pourmand, Nader; Jejelowo, Olufisayo

    2011-03-01

    The space radiation environment consists of trapped particle radiation, solar particle radiation, and galactic cosmic radiation (GCR), in which protons are the most abundant particle type. During missions to the moon or to Mars, the constant exposure to GCR and occasional exposure to particles emitted from solar particle events (SPE) are major health concerns for astronauts. Therefore, in order to determine health risks during space missions, an understanding of cellular responses to proton exposure is of primary importance. The expression of DNA repair genes in response to ionizing radiation (X-rays and gamma rays) has been studied, but data on DNA repair in response to protons is lacking. Using qPCR analysis, we investigated changes in gene expression induced by positively charged particles (protons) in four categories (0, 0.1, 1.0, and 2.0 Gy) in nine different DNA repair genes isolated from the testes of irradiated mice. DNA repair genes were selected on the basis of their known functions. These genes include ERCC1 (5' incision subunit, DNA strand break repair), ERCC2/NER (opening DNA around the damage, Nucleotide Excision Repair), XRCC1 (5' incision subunit, DNA strand break repair), XRCC3 (DNA break and cross-link repair), XPA (binds damaged DNA in preincision complex), XPC (damage recognition), ATA or ATM (activates checkpoint signaling upon double strand breaks), MLH1 (post-replicative DNA mismatch repair), and PARP1 (base excision repair). Our results demonstrate that ERCC1, PARP1, and XPA genes showed no change at 0.1 Gy radiation, up-regulation at 1.0 Gy radiation (1.09 fold, 7.32 fold, 0.75 fold, respectively), and a remarkable increase in gene expression at 2.0 Gy radiation (4.83 fold, 57.58 fold and 87.58 fold, respectively). Expression of other genes, including ATM and XRCC3, was unchanged at 0.1 and 1.0 Gy radiation but showed up-regulation at 2.0 Gy radiation (2.64 fold and 2.86 fold, respectively). We were unable to detect gene expression for the

  18. Genome analysis of DNA repair genes in the alpha proteobacterium Caulobacter crescentus

    PubMed Central

    Martins-Pinheiro, Marinalva; Marques, Regina CP; Menck, Carlos FM

    2007-01-01

    Background The integrity of DNA molecules is fundamental for maintaining life. The DNA repair proteins protect organisms against genetic damage, by removal of DNA lesions or helping to tolerate them. DNA repair genes are best known from the gamma-proteobacterium Escherichia coli, which is the most understood bacterial model. However, genome sequencing raises questions regarding uniformity and ubiquity of these DNA repair genes and pathways, reinforcing the need for identifying genes and proteins, which may respond to DNA damage in other bacteria. Results In this study, we employed a bioinformatic approach, to analyse and describe the open reading frames potentially related to DNA repair from the genome of the alpha-proteobacterium Caulobacter crescentus. This was performed by comparison with known DNA repair related genes found in public databases. As expected, although C. crescentus and E. coli bacteria belong to separate phylogenetic groups, many of their DNA repair genes are very similar. However, some important DNA repair genes are absent in the C. crescentus genome and other interesting functionally related gene duplications are present, which do not occur in E. coli. These include DNA ligases, exonuclease III (xthA), endonuclease III (nth), O6-methylguanine-DNA methyltransferase (ada gene), photolyase-like genes, and uracil-DNA-glycosylases. On the other hand, the genes imuA and imuB, which are involved in DNA damage induced mutagenesis, have recently been described in C. crescentus, but are absent in E. coli. Particularly interesting are the potential atypical phylogeny of one of the photolyase genes in alpha-proteobacteria, indicating an origin by horizontal transfer, and the duplication of the Ada orthologs, which have diverse structural configurations, including one that is still unique for C. crescentus. Conclusion The absence and the presence of certain genes are discussed and predictions are made considering the particular aspects of the C. crescentus

  19. DNA repair genes are selectively mutated in diffuse large B cell lymphomas

    PubMed Central

    de Miranda, Noel FCC; Peng, Roujun; Georgiou, Konstantinos; Wu, Chenglin; Sörqvist, Elin Falk; Berglund, Mattias; Chen, Longyun; Gao, Zhibo; Lagerstedt, Kristina; Lisboa, Susana; Roos, Fredrik; van Wezel, Tom; Teixeira, Manuel R.; Rosenquist, Richard; Sundström, Christer; Enblad, Gunilla; Nilsson, Mats; Zeng, Yixin; Kipling, David

    2013-01-01

    DNA repair mechanisms are fundamental for B cell development, which relies on the somatic diversification of the immunoglobulin genes by V(D)J recombination, somatic hypermutation, and class switch recombination. Their failure is postulated to promote genomic instability and malignant transformation in B cells. By performing targeted sequencing of 73 key DNA repair genes in 29 B cell lymphoma samples, somatic and germline mutations were identified in various DNA repair pathways, mainly in diffuse large B cell lymphomas (DLBCLs). Mutations in mismatch repair genes (EXO1, MSH2, and MSH6) were associated with microsatellite instability, increased number of somatic insertions/deletions, and altered mutation signatures in tumors. Somatic mutations in nonhomologous end-joining (NHEJ) genes (DCLRE1C/ARTEMIS, PRKDC/DNA-PKcs, XRCC5/KU80, and XRCC6/KU70) were identified in four DLBCL tumors and cytogenetic analyses revealed that translocations involving the immunoglobulin-heavy chain locus occurred exclusively in NHEJ-mutated samples. The novel mutation targets, CHEK2 and PARP1, were further screened in expanded DLBCL cohorts, and somatic as well as novel and rare germline mutations were identified in 8 and 5% of analyzed tumors, respectively. By correlating defects in a subset of DNA damage response and repair genes with genomic instability events in tumors, we propose that these genes play a role in DLBCL lymphomagenesis. PMID:23960188

  20. Preferential Repair of DNA Double-strand Break at the Active Gene in Vivo*

    PubMed Central

    Chaurasia, Priyasri; Sen, Rwik; Pandita, Tej K.; Bhaumik, Sukesh R.

    2012-01-01

    Previous studies have demonstrated transcription-coupled nucleotide/base excision repair. We report here for the first time that DNA double-strand break (DSB) repair is also coupled to transcription. We generated a yeast strain by introducing a homing (Ho) endonuclease cut site followed by a nucleotide sequence for multiple Myc epitopes at the 3′ end of the coding sequence of a highly active gene, ADH1. This yeast strain also contains the Ho cut site at the nearly silent or poorly active mating type α (MATα) locus and expresses Ho endonuclease under the galactose-inducible GAL1 promoter. Using this strain, DSBs were generated at the ADH1 and MATα loci in galactose-containing growth medium that induced HO expression. Subsequently, yeast cells were transferred to dextrose-containing growth medium to stop HO expression, and the DSB repair was monitored at the ADH1 and MATα loci by PCR, using the primer pairs flanking the Ho cut sites. Our results revealed a faster DSB repair at the highly active ADH1 than that at the nearly silent MATα locus, hence implicating a transcription-coupled DSB repair at the active gene in vivo. Subsequently, we extended this study to another gene, PHO5 (carrying the Ho cut site at its coding sequence), under transcriptionally active and inactive growth conditions. We found a fast DSB repair at the active PHO5 gene in comparison to its inactive state. Collectively, our results demonstrate a preferential DSB repair at the active gene, thus supporting transcription-coupled DSB repair in living cells. PMID:22910905

  1. Preferential DNA repair of 3-alkyladenine sites in essential and nonessential genes of human astrocytes

    SciTech Connect

    Lapcevich, R.K.; Weiss, R.B.; Gallagher, P.E. )

    1991-03-11

    In recent years, numbers of studies examining excision rates of DNA damaged lesions in defined, subgenomic sequences have shown that DNA repair is not a uniform process throughout the genome. Here, the authors report data on the preferential, in vivo DNA repair of alkylation-induced lesions within specific DNA sequences of essential and nonessential genes. The formation and rate of removal of 3-alkyladenine were studied in these DNA fragments following treatment of human astrocytes with dimethyl sulfate. The distribution and quantitation of this damaged lesion in the isolated DNA from these cells were determined by a polymerase chain reaction assay. The results indicate that alkyladenines are more efficiently repaired in DNA fragments of essential genes than in comparable fragments of nonessential genes. In subsequent experiments, the repair rate of 3-alkyladenine was examined in DNA isolated from alkylation-treated human astrocytes, grown in serum-free medium to inhibit proliferation. The rate of repair of alkylation-induced lesions in essential and nonessential gene fragments also differed in actively growing and quiescent human astrocytes. The results of this study indicate that transcription plays an important role in the efficient removal of 3-alkyladenine by DNA repair systems.

  2. Exercise as Gene Therapy: BDNF and DNA Damage Repair.

    PubMed

    Schmidt, Robin H; Nickerson, John M; Boatright, Jeffrey H

    2016-01-01

    DNA damage is a common feature of neurodegenerative illnesses, and the ability to repair DNA strand breaks and lesions is crucial for neuronal survival, reported by Jeppesen et al (Prog Neurobiol. 2011;94:166-200) and Shiwaku et al (Curr Mol Med. 2015;15:119-128). Interventions aimed at repairing these lesions, therefore, could be useful for preventing or delaying the progression of disease. One potential strategy for promoting DNA damage repair (DDR) is exercise. Although the role of exercise in DDR is not understood, there is increasing evidence that simple physical activity may impact clinical outcomes for neurodegeneration. Here, we discuss what is currently known about the molecular mechanisms of brain-derived neurotrophic factor and how these mechanisms might influence the DDR process. PMID:27488073

  3. Gene Expression in Experimental Aortic Coarctation and Repair: Candidate Genes for Therapeutic Intervention?

    PubMed Central

    LaDisa, John F.; Bozdag, Serdar; Olson, Jessica; Ramchandran, Ramani; Kersten, Judy R.; Eddinger, Thomas J.

    2015-01-01

    Coarctation of the aorta (CoA) is a constriction of the proximal descending thoracic aorta and is one of the most common congenital cardiovascular defects. Treatments for CoA improve life expectancy, but morbidity persists, particularly due to the development of chronic hypertension (HTN). Identifying the mechanisms of morbidity is difficult in humans due to confounding variables such as age at repair, follow-up duration, coarctation severity and concurrent anomalies. We previously developed an experimental model that replicates aortic pathology in humans with CoA without these confounding variables, and mimics correction at various times using dissolvable suture. Here we present the most comprehensive description of differentially expressed genes (DEGs) to date from the pathology of CoA, which were obtained using this model. Aortic samples (n=4/group) from the ascending aorta that experiences elevated blood pressure (BP) from induction of CoA, and restoration of normal BP after its correction, were analyzed by gene expression microarray, and enriched genes were converted to human orthologues. 51 DEGs with >6 fold-change (FC) were used to determine enriched Gene Ontology terms, altered pathways, and association with National Library of Medicine Medical Subject Headers (MeSH) IDs for HTN, cardiovascular disease (CVD) and CoA. The results generated 18 pathways, 4 of which (cell cycle, immune system, hemostasis and metabolism) were shared with MeSH ID’s for HTN and CVD, and individual genes were associated with the CoA MeSH ID. A thorough literature search further uncovered association with contractile, cytoskeletal and regulatory proteins related to excitation-contraction coupling and metabolism that may explain the structural and functional changes observed in our experimental model, and ultimately help to unravel the mechanisms responsible for persistent morbidity after treatment for CoA. PMID:26207811

  4. Lack of association between polymorphisms of the DNA base excision repair genes MUTYH and hOGG1 and keratoconus in a Polish subpopulation

    PubMed Central

    Synowiec, Ewelina; Wójcik, Katarzyna A.; Czubatka, Anna; Polakowski, Piotr; Izdebska, Justyna; Szaflik, Jerzy; Błasiak, Janusz

    2015-01-01

    Introduction Keratoconus (KC) is a non-inflammatory thinning of the cornea and a leading indication for corneal transplantation. Oxidative stress plays a role in the pathogenesis of this disease. The products of the hOGG1 and MUTYH genes play an important role in the repair of oxidatively modified DNA in the base excision repair pathway. We hypothesized that variability in these genes may change susceptibility to oxidative stress and predispose individuals to the development of KC. We investigated the possible association between the c.977C>G polymorphism of the hOGG1 gene (rs1052133) and the c.972G>C polymorphism of the MUTYH gene (rs3219489) and KC occurrence as well as the modulation of this association by some KC risk factors. Material and methods A total of 205 patients with KC and 220 controls were included in this study. The polymorphisms were genotyped with polymerase chain reaction (PCR) restriction fragment length polymorphism and PCR-confronting two-pair primer techniques. Differences in genotype and allele frequency distributions were evaluated using the χ2 test, and KC risk was estimated with an unconditional multiple logistic regression with and without adjustment for co-occurrence of visual impairment, allergies, sex and family history for KC. Results We did not find any association between the genotypes and combined genotypes of the c.977C>G polymorphism of the hOGG1 gene and the c.972G>C polymorphism of the MUTYH gene and the occurrence of KC. Conclusions Our findings suggest that the c.977C>G-hOGG1 polymorphism and the c.972G>C-MUTYH polymorphism may not be linked with KC occurrence in this Polish subpopulation. PMID:26528356

  5. Different patterns of evolution for duplicated DNA repair genes in bacteria of the Xanthomonadales group

    PubMed Central

    Martins-Pinheiro, Marinalva; Galhardo, Rodrigo S; Lage, Claudia; Lima-Bessa, Keronninn M; Aires, Karina A; Menck, Carlos FM

    2004-01-01

    Background DNA repair genes encode proteins that protect organisms against genetic damage generated by environmental agents and by-products of cell metabolism. The importance of these genes in life maintenance is supported by their high conservation, and the presence of duplications of such genes may be easily traced, especially in prokaryotic genomes. Results The genome sequences of two Xanthomonas species were used as the basis for phylogenetic analyses of genes related to DNA repair that were found duplicated. Although 16S rRNA phylogenetic analyses confirm their classification at the basis of the gamma proteobacteria subdivision, differences were found in the origin of the various genes investigated. Except for lexA, detected as a recent duplication, most of the genes in more than one copy are represented by two highly divergent orthologs. Basically, one of such duplications is frequently positioned close to other gamma proteobacteria, but the second is often positioned close to unrelated bacteria. These orthologs may have occurred from old duplication events, followed by extensive gene loss, or were originated from lateral gene transfer (LGT), as is the case of the uvrD homolog. Conclusions Duplications of DNA repair related genes may result in redundancy and also improve the organisms' responses to environmental challenges. Most of such duplications, in Xanthomonas, seem to have arisen from old events and possibly enlarge both functional and evolutionary genome potentiality. PMID:15333143

  6. [Role of DNA repair genes in radiation-induced changes of lifespan of Drosophila melanogaster].

    PubMed

    Shilova, L A; Pliusnina, E N; Zemskaia, N V; Moskalev, A A

    2014-01-01

    One of the main effects of various stress factors, including ionizing radiation, is DNA damage. Accumulation of DNA damage and somatic mutations in the somatic tissues is regarded as one of the basic mechanisms of aging. We have developed an approach to the study of molecular and genetic mechanisms of radioadaptation, which is based on the analysis of changes in the lifespan of Drosophila with a transformed genotype. In this study we investigated the radioadaptive response and hormesis by radiation-induced changed of the lifespan of different strains of Drosophila melanogaster, such as a wild type strain Canton-Sand strains with mutations in DNA damage response gene (homologue of GADD45), excision repair genes (homologues of XPF, XPC, PCNA) and double-strand breaks repair genes (homologues of RAD54, XRCC3, BLM). The exposure to irradiation at the dose rate of 40 cGy was performed chronically through the stages of fly development; an acute exposure at the dose rate of 30 Gy was applied to the adult stages of flies. Also, we investigated the resistance to acute gamma-radiation of Drosophila with conditional ubiquitous overexpression of genes that are involved in DNA damage recognition (homologues of GADD45, HUS1, CHK2), excision repair (homologues of XPF, XPC, AP-endonuclease-1) and double-strand break repair (homologues of BRCA2, XRCC3, KU80, WRNexo). In the wild type strain Canton-S, manifestation of the radioadaptive response and radiation hormesis were observed. In individuals with DNA repair gene mutations, no radioadaptive response was observed, or observed to a lesser extent than in wild type flies. Mifepristone--inducible transgene activation does not lead to an increase in resistance to acute irradiation by the parameters of lifespan of Drosophila. Overexpression of DNA repair genes led to a sharp decline in lifespan also in the absence of irradiation. PMID:25775840

  7. Mechanisms of double-strand-break repair during gene targeting in mammalian cells.

    PubMed Central

    Ng, P; Baker, M D

    1999-01-01

    In the present study, the mechanism of double-strand-break (DSB) repair during gene targeting at the chromosomal immunoglobulin mu-locus in a murine hybridoma was examined. The gene-targeting assay utilized specially designed insertion vectors genetically marked in the region of homology to the chromosomal mu-locus by six diagnostic restriction enzyme site markers. The restriction enzyme markers permitted the contribution of vector-borne and chromosomal mu-sequences in the recombinant product to be determined. The use of the insertion vectors in conjunction with a plating procedure in which individual integrative homologous recombination events were retained for analysis revealed several important features about the mammalian DSB repair process:The presence of the markers within the region of shared homology did not affect the efficiency of gene targeting.In the majority of recombinants, the vector-borne marker proximal to the DSB was absent, being replaced with the corresponding chromosomal restriction enzyme site. This result is consistent with either formation and repair of a vector-borne gap or an "end" bias in mismatch repair of heteroduplex DNA (hDNA) that favored the chromosomal sequence. Formation of hDNA was frequently associated with gene targeting and, in most cases, began approximately 645 bp from the DSB and could encompass a distance of at least 1469 bp.The hDNA was efficiently repaired prior to DNA replication.The repair of adjacent mismatches in hDNA occurred predominantly on the same strand, suggesting the involvement of a long-patch repair mechanism. PMID:10049929

  8. The ancient and evolving roles of cohesin in DNA repair and gene expression

    PubMed Central

    Dorsett, Dale; Ström, Lena

    2012-01-01

    The cohesin complex, named for its key role in sister chromatid cohesion, also plays critical roles in DNA repair and gene regulation. It performs all three functions in single cell eukaryotes such as yeasts, and in higher organisms such as man. Minor disruption of cohesin function has significant consequences for human development, even in the absence of measurable effects on chromatid cohesion or chromosome segregation. Here we survey the roles of cohesin in DNA repair and gene regulation, and how these functions vary from yeast to man. PMID:22497943

  9. Repair of UV damaged DNA, genes and proteins of yeast and human

    SciTech Connect

    Prakash, L.

    1991-04-01

    Our objectives are to determine the molecular mechanism of the incision step of excision repair of ultraviolet (UV) light damaged DNA in eukaryotic organisms, using the yeast Saccharomyces cerevisiae as a model system, as well as studying the human homologs of yeast excision repair and postreplication repair proteins. In addition to its single-stranded DNA-dependent A TPase and DNA helicase activities, we have found that RAD3 protein also possesses DNA-RNA helicase activity, and that like RAD3, the Schizosaccharomyces pombe RAD3 homolog, rhp3{sup +}, is an essential gene. We have overexpressed the human RAD3 homolog, ERCC2, in yeast to facilitate its purification. The RAD10 protein was purified to homogeneity and shown to bind DNA. ERCC3y, the yeast homolog of the human ERCC-3/XP-B gene, has been sequenced and shown to be essential for viability. The Drosophila and human homologs of RAD6, required for postreplication repair and UV induced mutagenesis, were shown to complement the rad6 {Delta} mutation of yeast. Since defective DNA repair and enhanced neoplasia characterize several human genetic diseases, and repair proteins are highly conserved between yeast and man, a thorough understanding of the molecular mechanisms of DNA repir in yeast should provide a better understanding of the causes of carcinogenesis.

  10. The Convergence of Fracture Repair and Stem Cells: Interplay of Genes, Aging, Environmental Factors and Disease

    PubMed Central

    Hadjiargyrou, Michael; O’Keefe, Regis J

    2015-01-01

    The complexity of fracture repair makes it an ideal process for studying the interplay between the molecular, cellular, tissue, and organ level events involved in tissue regeneration. Additionally, as fracture repair recapitulates many of the processes that occur during embryonic development, investigations of fracture repair provide insights regarding skeletal embryogenesis. Specifically, inflammation, signaling, gene expression, cellular proliferation and differentiation, osteogenesis, chondrogenesis, angiogenesis, and remodeling represent the complex array of interdependent biological events that occur during fracture repair. Here we review studies of bone regeneration in genetically modified mouse models, during aging, following environmental exposure, and in the setting of disease that provide insights regarding the role of multipotent cells and their regulation during fracture repair. Complementary animal models and ongoing scientific discoveries define an increasing number of molecular and cellular targets to reduce the morbidity and complications associated with fracture repair. Last, some new and exciting areas of stem cell research such as the contribution of mitochondria function, limb regeneration signaling, and microRNA (miRNA) posttranscriptional regulation are all likely to further contribute to our understanding of fracture repair as an active branch of regenerative medicine. PMID:25264148

  11. The Finland–United States Investigation of Non–Insulin-Dependent Diabetes Mellitus Genetics (FUSION) Study. I. An Autosomal Genome Scan for Genes That Predispose to Type 2 Diabetes

    PubMed Central

    Ghosh, Soumitra; Watanabe, Richard M.; Valle, Timo T.; Hauser, Elizabeth R.; Magnuson, Victoria L.; Langefeld, Carl D.; Ally, Delphine S.; Mohlke, Karen L.; Silander, Kaisa; Kohtamäki, Kimmo; Chines, Peter; Balow, Jr., James; Birznieks, Gunther; Chang, Jennie; Eldridge, William; Erdos, Michael R.; Karanjawala, Zarir E.; Knapp, Julie I.; Kudelko, Kristina; Martin, Colin; Morales-Mena, Anabelle; Musick, Anjene; Musick, Tiffany; Pfahl, Carrie; Porter, Rachel; Rayman, Joseph B.; Rha, David; Segal, Leonid; Shapiro, Shane; Sharaf, Ravi; Shurtleff, Ben; So, Alistair; Tannenbaum, Joyce; Te, Catherine; Tovar, Jason; Unni, Arun; Welch, Christian; Whiten, Ray; Witt, Alyson; Blaschak-Harvan, Jillian; Douglas, Julie A.; Duren, William L.; Epstein, Michael P.; Fingerlin, Tasha E.; Kaleta, Hong Shi; Lange, Ethan M.; Li, Chun; McEachin, Richard C.; Stringham, Heather M.; Trager, Edward; White, Peggy P.; Eriksson, Johan; Toivanen, Liisa; Vidgren, Gabriele; Nylund, Stella J.; Tuomilehto-Wolf, Eva; Ross, Edna H.; Demirchyan, Elza; Hagopian, William A.; Buchanan, Thomas A.; Tuomilehto, Jaakko; Bergman, Richard N.; Collins, Francis S.; Boehnke, Michael

    2000-01-01

    We performed a genome scan at an average resolution of 8 cM in 719 Finnish sib pairs with type 2 diabetes. Our strongest results are for chromosome 20, where we observe a weighted maximum LOD score (MLS) of 2.15 at map position 69.5 cM from pter and secondary weighted LOD-score peaks of 2.04 at 56.5 cM and 1.99 at 17.5 cM. Our next largest MLS is for chromosome 11 (MLS = 1.75 at 84.0 cM), followed by chromosomes 2 (MLS = 0.87 at 5.5 cM), 10 (MLS = 0.77 at 75.0 cM), and 6 (MLS = 0.61 at 112.5 cM), all under an additive model. When we condition on chromosome 2 at 8.5 cM, the MLS for chromosome 20 increases to 5.50 at 69.0 cM (P=.0014). An ordered-subsets analysis based on families with high or low diabetes-related quantitative traits yielded results that support the possible existence of disease-predisposing genes on chromosomes 6 and 10. Genomewide linkage-disequilibrium analysis using microsatellite marker data revealed strong evidence of association for D22S423 (P=.00007). Further analyses are being carried out to confirm and to refine the location of these putative diabetes-predisposing genes. PMID:11032783

  12. DNA repair gene expression in biological tissues exposed to low-intensity infrared laser.

    PubMed

    de Souza da Fonseca, Adenilson; Mencalha, Andre Luiz; Araújo de Campos, Vera Maria; Ferreira Machado, Samara Cristina; de Freitas Peregrino, Antonio Augusto; Geller, Mauro; de Paoli, Flavia

    2013-07-01

    Special properties of laser light have led to its usefulness in many applications in therapy. Excitation of endogenous chromophores in biotissues and generation of free radicals could be involved in its biological effects. DNA lesions induced by free radicals are repaired by base excision repair pathway. In this work, we evaluated the expression of APE1 and OGG1 genes related to repair of DNA lesions induced by free radicals. Skin and muscle tissues of Wistar rats were exposed to low-intensity infrared laser at different fluences and frequencies. After laser exposition of 1 and 24 h, tissue samples were withdrawn for total RNA extraction, cDNA synthesis, and evaluation of APE1 and OGG1 gene expression by quantitative polymerase chain reaction. Data obtained show that laser radiation alters the expression of APE1 and OGG1 mRNA differently in skin and muscle tissues of Wistar rats depending of the fluence, frequency, and time after exposure. Our study suggests that low-intensity infrared laser affects expression of genes involved in repair of DNA lesions by base excision repair pathway. PMID:22941447

  13. Repair of rDNA in Saccharomyces cerevisiae: RAD4-independent strand-specific nucleotide excision repair of RNA polymerase I transcribed genes.

    PubMed Central

    Verhage, R A; Van de Putte, P; Brouwer, J

    1996-01-01

    Removal of UV-induced pyrimidine dimers from the individual strands of the rDNA locus in Saccharomyces cerevisiae was studied. Yeast rDNA, that is transcribed by RNA polymerase I(RNA pol I), is repaired efficiently, slightly strand-specific and independently of RAD26, which has been implicated in transcription-coupled repair of the RNA pol II transcribed RPB2 gene. No repair of rDNA is observed in rad1,2,3 and 14 mutants, demonstrating that dimer removal from this highly repetitive DNA is accomplished by nucleotide excision repair (NER). In rad7 and rad16 mutants, which are specifically deficient in repair of non-transcribed DNA, there is a clear preferential repair of the transcribed strand of rDNA, indicating that strand-specific and therefore probably transcription-coupled repair of RNA pol I transcribed genes does exist in yeast. Unexpectedly, the transcribed but not the non-transcribed strand of rDNA can be repaired in rad4 mutants, which seem otherwise completely NER-deficient. PMID:8604332

  14. Expression of DNA repair and metabolic genes in response to a flavonoid-rich diet.

    PubMed

    Guarrera, Simonetta; Sacerdote, Carlotta; Fiorini, Laura; Marsala, Rosa; Polidoro, Silvia; Gamberini, Sara; Saletta, Federica; Malaveille, Christian; Talaska, Glenn; Vineis, Paolo; Matullo, Giuseppe

    2007-09-01

    A diet rich in fruit and vegetables can be effective in the reduction of oxidative stress, through the antioxidant effects of phytochemicals and other mechanisms. Protection against the carcinogenic effects of chemicals may also be exerted by an enhancement of detoxification and DNA damage repair mechanisms. To investigate a putative effect of flavonoids, a class of polyphenols, on the regulation of the gene expression of DNA repair and metabolic genes, a 1-month flavonoid-rich diet was administered to thirty healthy male smokers, nine of whom underwent gene expression analysis. We postulated that tobacco smoke is a powerful source of reactive oxygen species. The expression level of twelve genes (APEX, ERCC1, ERCC2, ERCC4, MGMT, OGG1, XPA, XPC, XRCC1, XRCC3, AHR, CYP1A1) was investigated. We found a significant increase (P < 0.001) in flavonoid intake. Urinary phenolic content and anti-mutagenicity did not significantly change after diet, nor was a correlation found between flavonoid intake and urinary phenolic levels or anti-mutagenicity. Phenolic levels showed a significant positive correlation with urinary anti-mutagenicity. AHR levels were significantly reduced after the diet (P = 0.038), whereas the other genes showed a generalized up regulation, significant for XRCC3 gene (P = 0.038). Also in the context of a generalized up regulation of DNA repair genes, we found a non-significant negative correlation between flavonoid intake and the expression of all the DNA repair genes. Larger studies are needed to clarify the possible effects of flavonoids in vivo; our preliminary results could help to better plan new studies on gene expression and diet. PMID:17445347

  15. Mutations in recombinational repair and in checkpoint control genes suppress the lethal combination of srs2Delta with other DNA repair genes in Saccharomyces cerevisiae.

    PubMed Central

    Klein, H L

    2001-01-01

    The SRS2 gene of Saccharomyces cerevisiae encodes a DNA helicase that is active in the postreplication repair pathway and homologous recombination. srs2 mutations are lethal in a rad54Delta background and cause poor growth or lethality in rdh54Delta, rad50Delta, mre11Delta, xrs2Delta, rad27Delta, sgs1Delta, and top3Delta backgrounds. Some of these genotypes are known to be defective in double-strand break repair. Many of these lethalities or poor growth can be suppressed by mutations in other genes in the DSB repair pathway, namely rad51, rad52, rad55, and rad57, suggesting that inhibition of recombination at a prior step prevents formation of a lethal intermediate. Lethality of the srs2Delta rad54Delta and srs2Delta rdh54Delta double mutants can also be rescued by mutations in the DNA damage checkpoint functions RAD9, RAD17, RAD24, and MEC3, indicating that the srs2 rad54 and srs2 rdh54 mutant combinations lead to an intermediate that is sensed by these checkpoint functions. When the checkpoints are intact the cells never reverse from the arrest, but loss of the checkpoints releases the arrest. However, cells do not achieve wild-type growth rates, suggesting that unrepaired damage is still present and may lead to chromosome loss. PMID:11156978

  16. An update on targeted gene repair in mammalian cells: methods and mechanisms.

    PubMed

    Jensen, Nanna M; Dalsgaard, Trine; Jakobsen, Maria; Nielsen, Roni R; Sørensen, Charlotte B; Bolund, Lars; Jensen, Thomas G

    2011-01-01

    Transfer of full-length genes including regulatory elements has been the preferred gene therapy strategy for clinical applications. However, with significant drawbacks emerging, targeted gene alteration (TGA) has recently become a promising alternative to this method. By means of TGA, endogenous DNA repair pathways of the cell are activated leading to specific genetic correction of single-base mutations in the genome. This strategy can be implemented using single-stranded oligodeoxyribonucleotides (ssODNs), small DNA fragments (SDFs), triplex-forming oligonucleotides (TFOs), adeno-associated virus vectors (AAVs) and zinc-finger nucleases (ZFNs). Despite difficulties in the use of TGA, including lack of knowledge on the repair mechanisms stimulated by the individual methods, the field holds great promise for the future. The objective of this review is to summarize and evaluate the different methods that exist within this particular area of human gene therapy research. PMID:21284895

  17. An update on targeted gene repair in mammalian cells: methods and mechanisms

    PubMed Central

    2011-01-01

    Transfer of full-length genes including regulatory elements has been the preferred gene therapy strategy for clinical applications. However, with significant drawbacks emerging, targeted gene alteration (TGA) has recently become a promising alternative to this method. By means of TGA, endogenous DNA repair pathways of the cell are activated leading to specific genetic correction of single-base mutations in the genome. This strategy can be implemented using single-stranded oligodeoxyribonucleotides (ssODNs), small DNA fragments (SDFs), triplex-forming oligonucleotides (TFOs), adeno-associated virus vectors (AAVs) and zinc-finger nucleases (ZFNs). Despite difficulties in the use of TGA, including lack of knowledge on the repair mechanisms stimulated by the individual methods, the field holds great promise for the future. The objective of this review is to summarize and evaluate the different methods that exist within this particular area of human gene therapy research. PMID:21284895

  18. Semiconservative replication, genetic repair, and many-gened genomes: Extending the quasispecies paradigm to living systems

    NASA Astrophysics Data System (ADS)

    Tannenbaum, Emmanuel; Shakhnovich, Eugene I.

    2005-12-01

    Quasispecies theory has emerged as an important tool for modeling the evolutionary dynamics of biological systems. We review recent advances in the field, with an emphasis on the quasispecies dynamics of semiconservatively replicating genomes. Applications to cancer and adult stem cell growth are discussed. Additional topics, such as genetic repair and many-gene genomes, are covered as well.

  19. Role of RAD52 Epistasis Group Genes in Homologous Recombination and Double-Strand Break Repair

    PubMed Central

    Symington, Lorraine S.

    2002-01-01

    The process of homologous recombination is a major DNA repair pathway that operates on DNA double-strand breaks, and possibly other kinds of DNA lesions, to promote error-free repair. Central to the process of homologous recombination are the RAD52 group genes (RAD50, RAD51, RAD52, RAD54, RDH54/TID1, RAD55, RAD57, RAD59, MRE11, and XRS2), most of which were identified by their requirement for the repair of ionizing-radiation-induced DNA damage in Saccharomyces cerevisiae. The Rad52 group proteins are highly conserved among eukaryotes, and Rad51, Mre11, and Rad50 are also conserved in prokaryotes and archaea. Recent studies showing defects in homologous recombination and double-strand break repair in several human cancer-prone syndromes have emphasized the importance of this repair pathway in maintaining genome integrity. Although sensitivity to ionizing radiation is a universal feature of rad52 group mutants, the mutants show considerable heterogeneity in different assays for recombinational repair of double-strand breaks and spontaneous mitotic recombination. Herein, I provide an overview of recent biochemical and structural analyses of the Rad52 group proteins and discuss how this information can be incorporated into genetic studies of recombination. PMID:12456786

  20. Cytogenetic Response to Ionizing Radiation Exposure in Human Fibroblasts with Suppressed Expression of Non-DSB Repair Genes

    NASA Technical Reports Server (NTRS)

    Zhang, Ye; Rohde, Larry H.; Emami, Kamal; Hammond, Dianne; Mehta, Satish K.; Jeevarajan, Antony S.; Pierson, Duane L.; Wu, Honglu

    2009-01-01

    Changes of gene expression profile are one of the most important biological responses in living cells after ionizing radiation (IR) exposure. Although some studies have shown that genes up-regulated by IR may play important roles in DNA damage repair, the relationship between the regulation of gene expression by IR, particularly genes not known for their roles in double-strand break (DSB) repair, and its impact on cytogenetic responses has not been well studied. The purpose of this study is to identify new roles of IR inducible genes in radiation-induced chromosome aberrations and micronuclei formation. In the study, the expression of 25 genes selected on the basis of their transcriptional changes in response to IR was individually knocked down by small interfering RNA in human fibroblast cells. Frequencies of micronuclei (MN) formation and chromosome aberrations were measured to determine the efficiency of cytogenetic repair, and the fraction of bi-nucleated cells in the MN analysis was used as a marker for cell cycle progression. In response to gamma radiation, the formation of MN was significantly increased by suppressed expression of five genes: Ku70 (DSB repair pathway), XPA (nucleotide excision repair pathway), RPA1 (mismatch repair pathway), RAD17 and RBBP8 (cell cycle control). Knocked-down expression of four genes (MRE11A, RAD51 in the DSB pathway, SESN1, and SUMO1) significantly inhibited cell cycle progression, possibly because of severe impairment of DNA damage repair. Moreover, decreased XPA, p21, or MLH1 expression resulted in both significantly enhanced cell cycle progression and increased yields of chromosome aberrations, indicating that these gene products modulate both cell cycle control and DNA damage repair. Nine of these eleven genes, whose knock-down expression affected cytogenetic repair, were up-regulated in cells exposed to gamma radiation, suggesting that genes transcriptionally modulated by IR were critical to regulate IR

  1. Loss of transcription factor early growth response gene 1 results in impaired endochondral bone repair

    PubMed Central

    Reumann, Marie K.; Strachna, Olga; Yagerman, Sarah; Torrecilla, Daniel; Kim, Jihye; Doty, Steven B.; Lukashova, Lyudmila; Boskey, Adele L.; Mayer-Kuckuk, Philipp

    2011-01-01

    Transcription factors that play a role in ossification during development are expected to participate in postnatal fracture repair since the endochondral bone formation that occurs in embryos is recapitulated during fracture repair. However, inherent differences exist between bone development and fracture repair, including a sudden disruption of tissue integrity followed by an inflammatory response. This raises the possibility that repair-specific transcription factors participate in bone healing. Here, we assessed the consequence of loss of early growth response gene 1 (EGR-1) on endochondral bone healing because this transcription factor has been shown to modulate repair in vascularized tissues. Model fractures were created in ribs of wild type (wt) and EGR-1−/− mice. Differences in tissue morphology and composition between these two animal groups were followed over 28 post fracture days (PFDs). In wt mice, bone healing occurred in healing phases characteristic of endochondral bone repair. A similar healing sequence was observed in EGR-1−/− mice but was impaired by alterations. A persistent accumulation of fibrin between the disconnected bones was observed on PFD7 and remained pronounced in the callus on PFD14. Additionally, the PFD14 callus was abnormally enlarged and showed increased deposition of mineralized tissue. Cartilage ossification in the callus was associated with hyper-vascularity and -proliferation. Moreover, cell deposits located in proximity to the callus within skeletal muscle were detected on PFD14. Despite these impairments, repair in EGR-1−/− callus advanced on PFD28, suggesting EGR-1 is not essential for healing. Together, this study provides genetic evidence that EGR-1 is a pleiotropic regulator of endochondral fracture repair. PMID:21726677

  2. Biochemical studies of DNA strand break repair and molecular characterization of mei-41, a gene involved in DNA break repair

    SciTech Connect

    Oliveri, D.R.

    1989-01-01

    The ability to repair X-irradiation induced single-strand DNA breaks was examined in mutagen-sensitive mutants of Drosophila melanogaster. This analysis demonstrated that examined stocks possess a normal capacity to repair X-ray induced single-strand breaks. One of the mutants in this study, mei-41, has been shown to be involved in a number of DNA metabolizing functions. A molecular characterization of this mutant is presented. A cDNA hybridizing to genomic DNA both proximal and distal to a P element inducing a mei-41 mutation was isolated from both embryonic and adult female recombinant lambda phage libraries. A 2.2 kilobase embryonic cDNA clone was sequenced; the sequence of an open reading frame was identified which would predict a protein of 384 amino acids with a molecular weight of 43,132 daltons. An examination of homologies to sequences in protein and nucleic acid data bases revealed no sequences with significant homology to mei-41, however, two potential Zinc-finger domains were identified. Analysis of RNA hybridizing to the embryonic cDNA demonstrated the existence of a major 2.2 kilobase transcript expressed primarily in embryos and adult flies. An examination of the transcription of this gene in mei-41 mutants revealed significant variation from wild-type, an indication that the embryonic cDNA does represent a mei-41 transcript. Expression in tissues from adult animals demonstrated that the 2.2 kilobase RNA is expressed primarily in reproductive tissues. A 3.8kb transcript is the major species of RNA in the adult head and thorax. Evidence is presented which implies that expression of the mei-41 gene is strongly induced by exposure of certain cells to mutagens.

  3. DNA repair gene variants associated with benign breast disease in high cancer risk women.

    PubMed

    Jorgensen, Timothy J; Helzlsouer, Kathy J; Clipp, Sandra C; Bolton, Judy Hoffman; Crum, Rosa M; Visvanathan, Kala

    2009-01-01

    Benign breast disease (BBD) is a risk factor for breast cancer and may have a heritable component. Deficient DNA repair has been implicated in breast cancer etiology and may exert its effect before BBD, a known precursor. The association between allelic variants in DNA repair genes and BBD was examined in a cohort of women in Washington County, Maryland. BBD was defined by two criteria: (a) a physician diagnosis of BBD or fibrocystic disease and/or (b) a benign breast biopsy. 3,212 women without BBD at baseline were genotyped for 12 candidate single nucleotide polymorphisms in seven DNA repair genes. Of these women, 482 subsequently reported a diagnosis of BBD. The Cox model was used to calculate hazard ratios (HR). Variant alleles of XRCC1 Arg(194)Trp (rs1799782) and ERCC4 Arg(415)Gln (rs1800067) were significantly associated with BBD [HR, 1.36; 95% confidence interval (95% CI), 1.06-1.74 and HR, 1.39; 95% CI, 1.09-1.76, respectively]. Similar estimates were also observed for each of the BBD criterion used. The BBD association for ERCC4 was even stronger among women with a family history of breast cancer (HR, 2.68; 95% CI, 1.52-4.66; P(interaction) = 0.02). This study suggests that variant alleles in DNA repair genes may modify BBD risk, a potential intermediate marker of breast cancer risk, particularly among high-risk subgroups. PMID:19124519

  4. Low-level infrared laser modulates muscle repair and chromosome stabilization genes in myoblasts.

    PubMed

    da Silva Neto Trajano, Larissa Alexsandra; Stumbo, Ana Carolina; da Silva, Camila Luna; Mencalha, Andre Luiz; Fonseca, Adenilson S

    2016-08-01

    Infrared laser therapy is used for skeletal muscle repair based on its biostimulative effect on satellite cells. However, shortening of telomere length limits regenerative potential in satellite cells, which occurs after each cell division cycle. Also, laser therapy could be more effective on non-physiologic tissues. This study evaluated low-level infrared laser exposure effects on mRNA expression from muscle injury repair and telomere stabilization genes in myoblasts in normal and stressful conditions. Laser fluences were those used in clinical protocols. C2C12 myoblast cultures were exposed to low-level infrared laser (10, 35, and 70 J/cm(2)) in standard or normal (10 %) and reduced (2 %) fetal bovine serum concentrations; total RNA was extracted for mRNA expression evaluation from muscle injury repair (MyoD and Pax7) and chromosome stabilization (TRF1 and TRF2) genes by real time quantitative polymerization chain reaction. Data show that low-level infrared laser increases the expression of MyoD and Pax7 in 10 J/cm(2) fluence, TRF1 expression in all fluences, and TRF2 expression in 70 J/cm(2) fluence in both 10 and 2 % fetal bovine serum. Low-level infrared laser increases mRNA expression from genes related to muscle repair and telomere stabilization in myoblasts in standard or normal and stressful conditions. PMID:27220530

  5. The impact of detoxifying and repair gene polymorphisms on oxidative stress in ischemic stroke.

    PubMed

    Orhan, Gürdal; Elkama, Aylin; Mungan, Semra Öztürk; Eruyar, Esra; Karahalil, Bensu

    2016-06-01

    Stroke is a multifactorial disease caused by the combination of certain risk factors and genetic factors. There are possible risk factors having important role in the pathogenesis of stroke. The most important environmental factors are cigarette smoking and oxidative stress which have different sources. GST (M1, T1, P1) have major roles in detoxification of the products of oxidative stress and they are polymorphic. DNA damages can also be repaired by repair enzymes such as OGG1 and XRCC1 which are highly polymorphic and have pivotal roles in repair systems. In the present study, we investigated that polymorphisms in genes involved in detoxification and DNA-repair pathways might modify the individual's risk for ischemic stroke. Furthermore, the products of oxidative stress and antioxidant capacity were measured and the impact of gene polymorphism on them was evaluated. Our data showed that OGG1 Ser326Cys and XRCC1 Arg399Gln gene polymorphisms had impacts on the development of stroke. PMID:26936466

  6. Transcription-coupled and global genome repair in the Saccharomyces cerevisiae RPB2 gene at nucleotide resolution.

    PubMed Central

    Tijsterman, M; Tasseron-de Jong, J G; van de Putte, P; Brouwer, J

    1996-01-01

    Repair of UV-induced cyclobutane pyrimidine dimers (CPDs) was examined at single nucleotide resolution in the yeast Saccharomyces cerevisiae, using an improved protocol for genomic end-labelling. To obtain the sensitivity required for adduct detection in yeast, an oligonucleotide-directed enrichment step was introduced into the current methodology developed for adduct detection in Escherichia coli. With this method, heterogeneous repair of CPDs within the RPB2 locus is observed. Individual CPDs positioned in the transcribed strand are removed very efficiently with identical kinetics. This fast repair starts within 23 bases downstream of the transcription initiation site. The non-transcribed strand of the active gene exhibits slow repair without detectable repair variations between individual lesions. In contrast, CPDs positioned in the promoter region show profound repair heterogeneity. Here, CPDs at specific sites are removed very quickly, with comparable rates to CPDs positioned in the transcribed strand, while at other positions lesions are not repaired at all during the period studied. Interestingly, the fast repair in the promoter region is dependent on the RAD7 and RAD16 genes, as are the slowly repaired CPDs in this region and in the non-transcribed strand. This indicates that the global genome repair pathway is not intrinsically slow and at specific positions can be as efficient as the transcription-coupled repair pathway. PMID:8836174

  7. Gene Delivery Strategies to Promote Spinal Cord Repair

    PubMed Central

    Walthers, Christopher M; Seidlits, Stephanie K

    2015-01-01

    Gene therapies hold great promise for the treatment of many neurodegenerative disorders and traumatic injuries in the central nervous system. However, development of effective methods to deliver such therapies in a controlled manner to the spinal cord is a necessity for their translation to the clinic. Although essential progress has been made to improve efficiency of transgene delivery and reduce the immunogenicity of genetic vectors, there is still much work to be done to achieve clinical strategies capable of reversing neurodegeneration and mediating tissue regeneration. In particular, strategies to achieve localized, robust expression of therapeutic transgenes by target cell types, at controlled levels over defined time periods, will be necessary to fully regenerate functional spinal cord tissues. This review summarizes the progress over the last decade toward the development of effective gene therapies in the spinal cord, including identification of appropriate target genes, improvements to design of genetic vectors, advances in delivery methods, and strategies for delivery of multiple transgenes with synergistic actions. The potential of biomaterials to mediate gene delivery while simultaneously providing inductive scaffolding to facilitate tissue regeneration is also discussed. PMID:25922572

  8. Transcriptional and Post-Transcriptional Regulation of Nucleotide Excision Repair Genes in Human Cells

    PubMed Central

    Lefkofsky, Hailey B.; Veloso, Artur; Ljungman, Mats

    2014-01-01

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death. PMID:26255935

  9. Transcriptional and post-transcriptional regulation of nucleotide excision repair genes in human cells.

    PubMed

    Lefkofsky, Hailey B; Veloso, Artur; Ljungman, Mats

    2015-06-01

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death. PMID:26255935

  10. 32 CFR 644.391 - Predisposal conference.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 32 National Defense 4 2010-07-01 2010-07-01 true Predisposal conference. 644.391 Section 644.391... ESTATE HANDBOOK Disposal Predisposal Action § 644.391 Predisposal conference. (a) Where a substantial Army installation, or portion thereof, is involved, the DE will convene a predisposal conference...

  11. DNA-repair genes and vitamin E in the prevention of N-nitrosodiethylamine mutagenicity.

    PubMed

    Aiub, Claudia Alessandra Fortes; Pinto, Luis Felipe Ribeiro; Felzenszwalb, Israel

    2009-08-01

    Nitrosamines are stable compounds, biologically and chemically inert unless activated. In biological systems, N-nitrosodiethylamine (NDEA) can be activated by a variety of enzymes, leading to aldehydes and/or intermediates which are themselves alkylating agents. Additionally, it has been shown that NDEA causes reactive oxygen species (ROS) production and induces mutagenicity. The cell defense seeks to neutralize ROS that escape the primary defense mechanisms (antioxidants) by DNA-repair mechanisms. NDEA is present at low concentrations in major dietary sources, like cured meats, salami, millet flour, and dried cuttlefish, where NDEA mutagenicity has been detected. These facts lead us to evaluate vitamin E as a ROS scavenger, in Escherichia coli mutants system, against genotoxicity induced by NDEA at low concentrations under exogenous metabolic activation. Statistical analysis were performed in order to compare the effects of NDEA-induced genotoxicity (a) between the mutants and the wild-type strains, at the same metabolic activation conditions and, (b) between the same strains in the presence or in the absence of vitamin E (150 muM). The indirect evaluation of ROS production by NDEA metabolizing shows that vitamin E protects E. coli cells proficient or deficient in the DNA-repair genes from cytotoxic effects. Our results underscore the role of scavenger molecules such as vitamin E in the diet, avoiding lesions induced by NDEA at low concentrations, via ROS, that could be repaired by nucleotide excision repair and base excision repair proteins. PMID:18581242

  12. Effect of transporter and DNA repair gene polymorphisms to lung cancer chemotherapy toxicity.

    PubMed

    Chen, Juan; Wu, Lin; Wang, Ying; Yin, Jiye; Li, Xiangping; Wang, Zhan; Li, Huihua; Zou, Ting; Qian, Chenyue; Li, Chuntian; Zhang, Wei; Zhou, Honghao; Liu, Zhaoqian

    2016-02-01

    Lung cancer is the first leading cause of cancer deaths. Chemotherapy toxicity is one of factors that limited the efficacy of platinum-based chemotherapy in lung cancer patients. Transporters and DNA repair genes play critical roles in occurrence of platinum-based chemotherapy toxicity. To investigate the relationships between transporter and DNA repair gene polymorphisms and platinum-based chemotherapy toxicity in lung cancer patients, we selected 60 polymorphisms in 14 transporters and DNA repair genes. The polymorphisms were genotyped in 317 lung cancer patients by Sequenom MassARRAY. Logistic regression was performed to estimate the association of toxicity outcome with the polymorphisms by PLINK. Our results showed that polymorphisms of SLC2A1 (rs3738514, rs4658, rs841844) were significantly related to overall toxicity. XRCC5 (rs1051685, rs6941) and AQP2 (10875989, rs3759125) polymorphisms were associated with hematologic toxicity. AQP2 polymorphisms (rs461872, rs7305534) were correlated with gastrointestinal toxicity. In conclusion, genotypes of these genes may be used to predict the platinum-based chemotherapy toxicity in lung cancer patients. PMID:26358256

  13. Recurrent gross mutations of the PTEN tumor suppressor gene in breast cancers with deficient DSB repair

    PubMed Central

    Saal, Lao H; Gruvberger-Saal, Sofia K; Persson, Camilla; Lövgren, Kristina; Jumppanen, Mervi; Staaf, Johan; Jönsson, Göran; Pires, Maira M; Maurer, Matthew; Holm, Karolina; Koujak, Susan; Subramaniyam, Shivakumar; Vallon-Christersson, Johan; Olsson, Haökan; Su, Tao; Memeo, Lorenzo; Ludwig, Thomas; Ethier, Stephen P; Krogh, Morten; Szabolcs, Matthias; Murty, Vundavalli VVS; Isola, Jorma; Hibshoosh, Hanina; Parsons, Ramon; Borg, Åke

    2010-01-01

    Basal-like breast cancer (BBC) is a subtype of breast cancer with poor prognosis1–3. Inherited mutations of BRCA1, a cancer susceptibility gene involved in double-strand DNA break (DSB) repair, lead to breast cancers that are nearly always of the BBC subtype3–5; however, the precise molecular lesions and oncogenic consequences of BRCA1 dysfunction are poorly understood. Here we show that heterozygous inactivation of the tumor suppressor gene Pten leads to the formation of basal-like mammary tumors in mice, and that loss of PTEN expression is significantly associated with the BBC subtype in human sporadic and BRCA1-associated hereditary breast cancers. In addition, we identify frequent gross PTEN mutations, involving intragenic chromosome breaks, inversions, deletions and micro copy number aberrations, specifically in BRCA1-deficient tumors. These data provide an example of a specific and recurrent oncogenic consequence of BRCA1-dependent dysfunction in DNA repair and provide insight into the pathogenesis of BBC with therapeutic implications. These findings also argue that obtaining an accurate census of genes mutated in cancer will require a systematic examination for gross gene rearrangements, particularly in tumors with deficient DSB repair. PMID:18066063

  14. DNA damage and gene therapy of xeroderma pigmentosum, a human DNA repair-deficient disease.

    PubMed

    Dupuy, Aurélie; Sarasin, Alain

    2015-06-01

    Xeroderma pigmentosum (XP) is a genetic disease characterized by hypersensitivity to ultra-violet and a very high risk of skin cancer induction on exposed body sites. This syndrome is caused by germinal mutations on nucleotide excision repair genes. No cure is available for these patients except a complete protection from all types of UV radiations. We reviewed the various techniques to complement or to correct the genetic defect in XP cells. We, particularly, developed the correction of XP-C skin cells using the fidelity of the homologous recombination pathway during repair of double-strand break (DSB) in the presence of XPC wild type sequences. We used engineered nucleases (meganuclease or TALE nuclease) to induce a DSB located at 90 bp of the mutation to be corrected. Expression of specific TALE nuclease in the presence of a repair matrix containing a long stretch of homologous wild type XPC sequences allowed us a successful gene correction of the original TG deletion found in numerous North African XP patients. Some engineered nucleases are sensitive to epigenetic modifications, such as cytosine methylation. In case of methylated sequences to be corrected, modified nucleases or demethylation of the whole genome should be envisaged. Overall, we showed that specifically-designed TALE-nuclease allowed us to correct a 2 bp deletion in the XPC gene leading to patient's cells proficient for DNA repair and showing normal UV-sensitivity. The corrected gene is still in the same position in the human genome and under the regulation of its physiological promoter. This result is a first step toward gene therapy in XP patients. PMID:26255934

  15. Coevolution between Nuclear-Encoded DNA Replication, Recombination, and Repair Genes and Plastid Genome Complexity.

    PubMed

    Zhang, Jin; Ruhlman, Tracey A; Sabir, Jamal S M; Blazier, John Chris; Weng, Mao-Lun; Park, Seongjun; Jansen, Robert K

    2016-01-01

    Disruption of DNA replication, recombination, and repair (DNA-RRR) systems has been hypothesized to cause highly elevated nucleotide substitution rates and genome rearrangements in the plastids of angiosperms, but this theory remains untested. To investigate nuclear-plastid genome (plastome) coevolution in Geraniaceae, four different measures of plastome complexity (rearrangements, repeats, nucleotide insertions/deletions, and substitution rates) were evaluated along with substitution rates of 12 nuclear-encoded, plastid-targeted DNA-RRR genes from 27 Geraniales species. Significant correlations were detected for nonsynonymous (dN) but not synonymous (dS) substitution rates for three DNA-RRR genes (uvrB/C, why1, and gyrA) supporting a role for these genes in accelerated plastid genome evolution in Geraniaceae. Furthermore, correlation between dN of uvrB/C and plastome complexity suggests the presence of nucleotide excision repair system in plastids. Significant correlations were also detected between plastome complexity and 13 of the 90 nuclear-encoded organelle-targeted genes investigated. Comparisons revealed significant acceleration of dN in plastid-targeted genes of Geraniales relative to Brassicales suggesting this correlation may be an artifact of elevated rates in this gene set in Geraniaceae. Correlation between dN of plastid-targeted DNA-RRR genes and plastome complexity supports the hypothesis that the aberrant patterns in angiosperm plastome evolution could be caused by dysfunction in DNA-RRR systems. PMID:26893456

  16. Coevolution between Nuclear-Encoded DNA Replication, Recombination, and Repair Genes and Plastid Genome Complexity

    PubMed Central

    Zhang, Jin; Ruhlman, Tracey A.; Sabir, Jamal S. M.; Blazier, John Chris; Weng, Mao-Lun; Park, Seongjun; Jansen, Robert K.

    2016-01-01

    Disruption of DNA replication, recombination, and repair (DNA-RRR) systems has been hypothesized to cause highly elevated nucleotide substitution rates and genome rearrangements in the plastids of angiosperms, but this theory remains untested. To investigate nuclear–plastid genome (plastome) coevolution in Geraniaceae, four different measures of plastome complexity (rearrangements, repeats, nucleotide insertions/deletions, and substitution rates) were evaluated along with substitution rates of 12 nuclear-encoded, plastid-targeted DNA-RRR genes from 27 Geraniales species. Significant correlations were detected for nonsynonymous (dN) but not synonymous (dS) substitution rates for three DNA-RRR genes (uvrB/C, why1, and gyrA) supporting a role for these genes in accelerated plastid genome evolution in Geraniaceae. Furthermore, correlation between dN of uvrB/C and plastome complexity suggests the presence of nucleotide excision repair system in plastids. Significant correlations were also detected between plastome complexity and 13 of the 90 nuclear-encoded organelle-targeted genes investigated. Comparisons revealed significant acceleration of dN in plastid-targeted genes of Geraniales relative to Brassicales suggesting this correlation may be an artifact of elevated rates in this gene set in Geraniaceae. Correlation between dN of plastid-targeted DNA-RRR genes and plastome complexity supports the hypothesis that the aberrant patterns in angiosperm plastome evolution could be caused by dysfunction in DNA-RRR systems. PMID:26893456

  17. Low intensity infrared laser affects expression of oxidative DNA repair genes in mitochondria and nucleus

    NASA Astrophysics Data System (ADS)

    Fonseca, A. S.; Magalhães, L. A. G.; Mencalha, A. L.; Geller, M.; Paoli, F.

    2014-11-01

    Practical properties and physical characteristics of low intensity lasers have made possible their application to treat soft tissue diseases. Excitation of intracellular chromophores by red and infrared radiation at low energy fluences with increase of mitochondrial metabolism is the basis of the biostimulation effect but free radicals can be produced. DNA lesions induced by free radicals are repaired by the base excision repair pathway. In this work, we evaluate the expression of POLγ and APEX2 genes related to repair of mitochondrial and nuclear DNA, respectively. Skin and muscle tissue of Wistar rats were exposed to low intensity infrared laser at different fluences. One hour and 24 hours after laser exposure, tissue samples were withdrawn for total RNA extraction, cDNA synthesis, and evaluation of POLγ and APEX2 mRNA expression by real time quantitative polymerase chain reaction. Skin and muscle tissue of Wistar rats exposed to laser radiation show different expression of POLγ and APEX2 mRNA depending of the fluence and time after exposure. Our study suggests that a low intensity infrared laser affects expression of genes involved in repair of oxidative lesions in mitochondrial and nuclear DNA.

  18. Capturing the biological impact of CDKN2A and MC1R genes as an early predisposing event in melanoma and non melanoma skin cancer

    PubMed Central

    Puig-Butille, Joan Anton; Escámez, María José; Garcia-Garcia, Francisco; Tell-Marti, Gemma; Fabra, Àngels; Martínez-Santamaría, Lucía; Badenas, Celia; Aguilera, Paula; Pevida, Marta; Dopazo, Joaquín; del Río, Marcela; Puig, Susana

    2014-01-01

    Germline mutations in CDKN2A and/or red hair color variants in MC1R genes are associated with an increased susceptibility to develop cutaneous melanoma or non melanoma skin cancer. We studied the impact of the CDKN2A germinal mutation p.G101W and MC1R variants on gene expression and transcription profiles associated with skin cancer. To this end we set-up primary skin cell co-cultures from siblings of melanoma prone-families that were later analyzed using the expression array approach. As a result, we found that 1535 transcripts were deregulated in CDKN2A mutated cells, with over-expression of immunity-related genes (HLA-DPB1, CLEC2B, IFI44, IFI44L, IFI27, IFIT1, IFIT2, SP110 and IFNK) and down-regulation of genes playing a role in the Notch signaling pathway. 3570 transcripts were deregulated in MC1R variant carriers. In particular, genes related to oxidative stress and DNA damage pathways were up-regulated as well as genes associated with neurodegenerative diseases such as Parkinson’s, Alzheimer and Huntington. Finally, we observed that the expression signatures indentified in phenotypically normal cells carrying CDKN2A mutations or MC1R variants are maintained in skin cancer tumors (melanoma and squamous cell carcinoma). These results indicate that transcriptome deregulation represents an early event critical for skin cancer development. PMID:24742402

  19. Association and Interactions between DNA Repair Gene Polymorphisms and Adult Glioma

    PubMed Central

    Liu, Yanhong; Scheurer, Michael E.; El-Zein, Randa; Cao, Yumei; Do, Kim-Anh; Gilbert, Mark; Aldape, Kenneth D.; Wei, Qingyi; Etzel, Carol; Bondy, Melissa L.

    2010-01-01

    It is generally accepted that glioma develops through accumulation of genetic alterations. We hypothesized that polymorphisms of candidate genes involved in the DNA repair pathways may contribute to susceptibility to glioma. To address this possibility, we conducted a study of 373 Caucasian glioma cases and 365 cancer-free Caucasian controls to assess associations between glioma risk and 18 functional SNPs in DNA repair genes. We evaluated potential gene-gene and gene-environment interactions using a multi-analytic strategy combining logistic regression, multifactor dimensionality reduction (MDR), and classification and regression tree (CART) approaches. In the single-locus analysis, six SNPs (ERCC1 3’ UTR, XRCC1 R399Q, APEX1 E148D, PARP1 A762V, MGMT F84L, and LIG1 5’UTR) showed a significant association with glioma risk. In the analysis of cumulative genetic risk of multiple SNPs, a significant gene-dosage effect was found for increased glioma risk with increasing numbers of adverse genotypes involving the above-mentioned six SNPs (P trend = 0.0004). Further, both the MDR and CART analyses identified MGMT F84L as the predominant risk factor for glioma, and revealed strong interactions among ionizing radiation (IR) exposure, PARP1 A762V, MGMT F84L and APEX1 E148D. Interestingly, the risk for glioma was dramatically increased in IR exposure individuals who had the wild-type genotypes of both MGMT F84L and PARP1 A762V [adjusted odds ratios (OR), 5.95; 95% confidence intervals (CI), 2.21–16.65]. Taken together, these results suggest that polymorphisms in DNA repair genes may act individually or together to contribute to glioma risk. PMID:19124499

  20. Rearrangement of Rag-1 recombinase gene in DNA-repair deficient/immunodeficient ``wasted`` mice

    SciTech Connect

    Woloschak, G.E.; Weaver, P.; Churchill, M.; Chang-Liu, C-M.; Libertin, C.R.

    1992-11-01

    Mice recessive for the autosomal gene ``wasted`` (wst) display a disease pattern which includes increased sensitivity to the killing effects of ionizing radiation, immunodeficiency, and neurologic dysfunction. The recent cloning and characterization of recombinase genes (Rag-l/Rag-2) expressed in lymphoid and possibly central nervous system tissues prompted us to examine expression of these genes in DNA repair-deficient/immunodeficient wasted mice. Our results revealed that in thymus tissue, a small Rag-I transcript (1.0 kb) was detected in wst/wst mice that was not evident in thymus from control mice. In wst/{sm_bullet} mice, a two-fold increase in Rag-1 mRNA was evident in thymus tissue. Rag-2 mRNA could only be detected in thymus tissue from wst/{sm_bullet} and not from wst/wst or parental control BCF, mice. Southern blots revealed a rearrangement or deletion within the Rag-1 gene of affected wasted mice that was not evident in known strain-specific parental or littermate controls. These results support the idea that the Rag-1 gene may map at or near the locus for the wasted mutation. In addition, they suggest the importance of recombinase function in normal immune and central nervous system development as well as the potential contribution of this gene family to the normal repair of radiation-induced DNA damage.

  1. Rearrangement of Rag-1 recombinase gene in DNA-repair deficient/immunodeficient wasted'' mice

    SciTech Connect

    Woloschak, G.E.; Weaver, P.; Churchill, M.; Chang-Liu, C-M. ); Libertin, C.R. )

    1992-01-01

    Mice recessive for the autosomal gene wasted'' (wst) display a disease pattern which includes increased sensitivity to the killing effects of ionizing radiation, immunodeficiency, and neurologic dysfunction. The recent cloning and characterization of recombinase genes (Rag-l/Rag-2) expressed in lymphoid and possibly central nervous system tissues prompted us to examine expression of these genes in DNA repair-deficient/immunodeficient wasted mice. Our results revealed that in thymus tissue, a small Rag-I transcript (1.0 kb) was detected in wst/wst mice that was not evident in thymus from control mice. In wst/[sm bullet] mice, a two-fold increase in Rag-1 mRNA was evident in thymus tissue. Rag-2 mRNA could only be detected in thymus tissue from wst/[sm bullet] and not from wst/wst or parental control BCF, mice. Southern blots revealed a rearrangement or deletion within the Rag-1 gene of affected wasted mice that was not evident in known strain-specific parental or littermate controls. These results support the idea that the Rag-1 gene may map at or near the locus for the wasted mutation. In addition, they suggest the importance of recombinase function in normal immune and central nervous system development as well as the potential contribution of this gene family to the normal repair of radiation-induced DNA damage.

  2. DNA Repair Gene Expression and Risk of Locoregional Relapse in Breast Cancer Patients

    SciTech Connect

    Le Scodan, Romuald; Cizeron-Clairac, Geraldine

    2010-10-01

    Purpose: Radiation therapy appears to kill cells mainly by inducing DNA double-strand breaks. We investigated whether the DNA repair gene expression status might influence the risk of locoregional recurrence (LRR) in breast cancer patients. Methods and Materials: We used a quantitative reverse transcriptase PCR-based approach to measure messenger RNA levels of 20 selected DNA repair genes in tumor samples from 97 breast cancer patients enrolled in a phase III trial (Centre Rene Huguenin cohort). Normalized mRNA levels were tested for an association with LRR-free survival (LRR-FS) and overall survival (OS). The findings were validated in comparison with those of an independent cohort (Netherlands Cancer Institute (NKI) cohort). Multivariate analysis encompassing known prognostic factors was used to assess the association between DNA repair gene expression and patient outcome. Results: RAD51 was the only gene associated with LRR in both cohorts. With a median follow-up of 126 months in the CRH cohort, the 5-year LRR-FS and OS rates were 100% and 95% in the 61 patients with low RAD51 expression, compared with 70% and 69% in the 36 patients with high RAD51 expression, respectively (p < 0.001). RAD51 overexpression was associated with a higher risk of LRR (hazard ratio [HR], 12.83; 95% confidence interval [CI], 3.6-45.6) and death (HR, 4.10; 95% CI, 1.7-9.7). RAD51 overexpression was also significantly associated with shorter LRR-FS and OS in the NKI cohort. Conclusions: Overexpression of RAD51, a key component of the homologous DNA repair pathway, is associated with poor breast cancer outcome. This finding warrants prospective studies of RAD51 as a prognosticator and therapeutic target.

  3. Evaluation of psychosocial effects of pre-symptomatic testing for breast/ovarian and colon cancer pre-disposing genes: a 12-month follow-up.

    PubMed

    Arver, Brita; Haegermark, Aina; Platten, Ulla; Lindblom, Annika; Brandberg, Yvonne

    2004-01-01

    A prospective study of psychosocial consequences following predictive testing for inherited mutations in breast/ovarian and colon cancer susceptibility genes BRCA1, BRCA2, MLH1, and MSH2 was performed. Eighty-seven healthy women were tested for known family mutations and self-assessment scales were used to evaluate anxiety, depression and quality of life. Extensive pre- and post-test information was given. Questionnaires were responded before testing and four times after during the following year. A statistically significant decrease in anxiety mean scores over time was observed among the studied participants. The levels of depression in cancer genes carriers decreased over time while, surprisingly the levels in non-carriers increased. Compared to a normative Swedish sample all women tested showed similar levels of anxiety but women tested for breast cancer genes showed statistically lower levels of depression. Vitality dropped initially after disclosure of the testing of colon cancer genes carriers, followed by increasing levels. No change in vitality or in other quality of life parameters was seen in the other groups and the levels were similar to Swedish norm data. Most tested individuals were satisfied with the testing procedure including genetic counselling and testing and all of them but one would redo the testing. Healthy self-referred women going through predictive breast/ovarian or colon cancer gene testing, including extensive pre- and post-test information and support, in general, will not experience adverse psychological consequences. PMID:15340261

  4. The C677T mutation in the methylenetetrahydrofolate reductase gene predisposes to hyperhomocysteinemia in children with familial hypercholesterolemia treated with cholestyramine.

    PubMed

    Tonstad, S; Refsum, H; Ose, L; Ueland, P M

    1998-02-01

    In children with familial hypercholesterolemia, heterozygosity and homozygosity for the C677T mutation in the methylenetetrahydrofolate reductase gene was associated with low serum folate and increased susceptibility to elevation of plasma total homocysteine during cholestyramine treatment. Because of the independent relationship between elevated plasma total homocysteine and cardiovascular disease, folate supplementation may be prudent in these children. PMID:9506661

  5. Microsatellites in the Eukaryotic DNA Mismatch Repair Genes as Modulators of Evolutionary Mutation Rate

    NASA Technical Reports Server (NTRS)

    Chang, Dong Kyung; Metzgar, David; Wills, Christopher; Boland, C. Richard

    2003-01-01

    All "minor" components of the human DNA mismatch repair (MMR) system-MSH3, MSH6, PMS2, and the recently discovered MLH3-contain mononucleotide microsatellites in their coding sequences. This intriguing finding contrasts with the situation found in the major components of the DNA MMR system-MSH2 and MLH1-and, in fact, most human genes. Although eukaryotic genomes are rich in microsatellites, non-triplet microsatellites are rare in coding regions. The recurring presence of exonal mononucleotide repeat sequences within a single family of human genes would therefore be considered exceptional.

  6. A Genome-Scale DNA Repair RNAi Screen Identifies SPG48 as a Novel Gene Associated with Hereditary Spastic Paraplegia

    PubMed Central

    Słabicki, Mikołaj; Theis, Mirko; Krastev, Dragomir B.; Samsonov, Sergey; Mundwiller, Emeline; Junqueira, Magno; Paszkowski-Rogacz, Maciej; Teyra, Joan; Heninger, Anne-Kristin; Poser, Ina; Prieur, Fabienne; Truchetto, Jérémy; Confavreux, Christian; Marelli, Cécilia; Durr, Alexandra; Camdessanche, Jean Philippe; Brice, Alexis; Shevchenko, Andrej; Pisabarro, M. Teresa; Stevanin, Giovanni; Buchholz, Frank

    2010-01-01

    DNA repair is essential to maintain genome integrity, and genes with roles in DNA repair are frequently mutated in a variety of human diseases. Repair via homologous recombination typically restores the original DNA sequence without introducing mutations, and a number of genes that are required for homologous recombination DNA double-strand break repair (HR-DSBR) have been identified. However, a systematic analysis of this important DNA repair pathway in mammalian cells has not been reported. Here, we describe a genome-scale endoribonuclease-prepared short interfering RNA (esiRNA) screen for genes involved in DNA double strand break repair. We report 61 genes that influenced the frequency of HR-DSBR and characterize in detail one of the genes that decreased the frequency of HR-DSBR. We show that the gene KIAA0415 encodes a putative helicase that interacts with SPG11 and SPG15, two proteins mutated in hereditary spastic paraplegia (HSP). We identify mutations in HSP patients, discovering KIAA0415/SPG48 as a novel HSP-associated gene, and show that a KIAA0415/SPG48 mutant cell line is more sensitive to DNA damaging drugs. We present the first genome-scale survey of HR-DSBR in mammalian cells providing a dataset that should accelerate the discovery of novel genes with roles in DNA repair and associated medical conditions. The discovery that proteins forming a novel protein complex are required for efficient HR-DSBR and are mutated in patients suffering from HSP suggests a link between HSP and DNA repair. PMID:20613862

  7. The rad18 gene of Schizosaccharomyces pombe defines a new subgroup of the SMC superfamily involved in DNA repair

    SciTech Connect

    Lehmann, A.R.; Walicka, M.; Griffiths, D.J.F.; Carr, A.M.

    1995-12-01

    This report describes the cloning and sequencing of the rad18 gene of Schizosaccharomyces pombe and its essential role in cell proliferation. It also describes the isolation and sequencing of its homolog from Saccharomyces cerevisiae, designated RHC18. Genetic radiation effects were explored and results indicate the gene product`s importance in a DNA repair pathway that is distinct from classical nucleotide excision repair. 57 refs., 20 figs., 1 tab.

  8. Enhanced pyrimidine dimer repair in cultured murine epithelial cells transfected with the denV gene of bacteriophage T4.

    PubMed

    Kusewitt, D F; Budge, C L; Ley, R D

    1994-04-01

    The patch size for excision repair of ultraviolet radiation (UV)-induced pyrimidine dimers was determined in cultured murine epithelial cells with normal and enhanced pyrimidine dimer repair capabilities. Cells with enhanced pyrimidine dimer repair were produced by transfecting 308 cells with the denV gene of bacteriophage T4; this gene encodes the enzyme endonuclease V. Pyrimidine dimer repair following exposure to UV from an FS-40 sunlamp was determined by micrococcal dimer-specific nuclease digestion and alkaline sucrose ultracentrifugation. Patch size ws estimated based on the photolytic lability of bromodeoxyuridine-substituted DNA. Excision repair of UV-induced pyrimidine dimers in denV-transfected 308 cells was enhanced two- to threefold. Production of mRNA from the denV gene in cell lines with enhanced repair was confirmed by RNA blotting. In control cells, the patch size for excision repair of DNA photoproducts was estimated to be 34 nucleotides per photoproduct removed; in denV-transfected cells, a smaller average patch size of 10-16 nucleotides per photoproduct removed was calculated. Thus, endonuclease V activity appears to alter not only the extent, but also the nature of excision repair in UV-exposed mammalian epithelial cells. PMID:8151125

  9. Non-homologous DNA increases gene disruption efficiency by altering DNA repair outcomes.

    PubMed

    Richardson, C D; Ray, G J; Bray, N L; Corn, J E

    2016-01-01

    The Cas9 endonuclease can be targeted to genomic sequences by programming the sequence of an associated single guide RNA (sgRNA). For unknown reasons, the activity of these Cas9-sgRNA combinations varies widely at different genomic loci and in different cell types. Thus, disrupting genes in polyploid cell lines or when using poorly performing sgRNAs can require extensive downstream screening to identify homozygous clones. Here we find that non-homologous single-stranded DNA greatly stimulates Cas9-mediated gene disruption in the absence of homology-directed repair. This stimulation increases the frequency of clones with homozygous gene disruptions and rescues otherwise ineffective sgRNAs. The molecular outcome of enhanced gene disruption depends upon cellular context, stimulating deletion of genomic sequence or insertion of non-homologous DNA at the edited locus in a cell line specific manner. Non-homologous DNA appears to divert cells towards error-prone instead of error-free repair pathways, dramatically increasing the frequency of gene disruption. PMID:27530320

  10. Non-homologous DNA increases gene disruption efficiency by altering DNA repair outcomes

    PubMed Central

    Richardson, C. D.; Ray, G. J.; Bray, N. L.; Corn, J. E.

    2016-01-01

    The Cas9 endonuclease can be targeted to genomic sequences by programming the sequence of an associated single guide RNA (sgRNA). For unknown reasons, the activity of these Cas9–sgRNA combinations varies widely at different genomic loci and in different cell types. Thus, disrupting genes in polyploid cell lines or when using poorly performing sgRNAs can require extensive downstream screening to identify homozygous clones. Here we find that non-homologous single-stranded DNA greatly stimulates Cas9-mediated gene disruption in the absence of homology-directed repair. This stimulation increases the frequency of clones with homozygous gene disruptions and rescues otherwise ineffective sgRNAs. The molecular outcome of enhanced gene disruption depends upon cellular context, stimulating deletion of genomic sequence or insertion of non-homologous DNA at the edited locus in a cell line specific manner. Non-homologous DNA appears to divert cells towards error-prone instead of error-free repair pathways, dramatically increasing the frequency of gene disruption. PMID:27530320

  11. Existence and expression of photoreactivation repair genes in various yeast species.

    PubMed

    Yasui, A; Eker, A P; Koken, M

    1989-01-01

    Photoreactivation repair (Phr) activities in cell extracts of 13 different yeast species were measured by the Haemophilus influenzae transformation assay. Five species including Schizosaccharomyces pombe showed no or low enzymatic activity. In contrast to the other species, chromosomal DNAs of these 5 species did not show detectable hybridization using a DNA fragment of the photolyase PHR1 gene of Saccharomyces cervisiae as a probe even at a low stringency condition. When the PHR1 gene was attached to the 5'-flanking sequence of the iso-1-cytochrome c (CYC-1) gene of S. cerevisiae and introduced into S. pombe cells, the transformants acquired a high Phr activity, indicating that the PHR1 gene alone can provide a Phr-negative species with this repair activity and the light-absorbing cofactor(s) must be present in S. pombe. Our results also demonstrated that the 5'-flanking sequence of the S. cerevisiae CYC-1 gene works in S. pombe as a regulatory element. PMID:2911265

  12. The Polycomb Group Protein EZH2 Impairs DNA Damage Repair Gene Expression in Human Uterine Fibroids.

    PubMed

    Yang, Qiwei; Nair, Sangeeta; Laknaur, Archana; Ismail, Nahed; Diamond, Michael P; Al-Hendy, Ayman

    2016-03-01

    Uterine fibroids are benign, smooth muscle tumors that occur in approximately 70%-80% of women by age 50 yr. The cellular and molecular mechanism(s) by which uterine fibroids (UFs) develop are not fully understood. Accumulating evidence demonstrates that several genetic abnormalities, including deletions, rearrangements, translocations, as well as mutations, have been found in UFs. These genetic anomalies suggest that low DNA damage repair capacity may be involved in UF formation. The objective of this study was to determine whether expression levels of DNA damage repair-related genes were altered, and how they were regulated in the pathogenesis of UFs. Expression levels of DNA repair-related genes RAD51 and BRCA1 were deregulated in fibroid tissues as compared to adjacent myometrial tissues. Expression levels of chromatin protein enhancer of zeste homolog 2 (EZH2) were higher in a subset of fibroids as compared to adjacent myometrial tissues by both immunohistochemistry and Western blot analysis. Treatment with an inhibitor of EZH2 markedly increased expression levels of RAD51 and BRCA1 in fibroid cells and inhibited cell proliferation paired with cell cycle arrest. Restoring the expression of RAD51 and BRCA1 by treatment with EZH2 inhibitor was dependent on reducing the enrichment of trimethylation of histone 3 lysine 27 epigenetic mark in their promoter regions. This study reveals the important role of EZH2-regulated DNA damage-repair genes via histone methylation in fibroid biology, and may provide novel therapeutic targets for the medical treatment of women with symptomatic UFs. PMID:26888970

  13. Nucleotide excision repair of the 5 S ribosomal RNA gene assembled into a nucleosome.

    PubMed

    Liu, X; Smerdon, M J

    2000-08-01

    A-175-base pair fragment containing the Xenopus borealis somatic 5 S ribosomal RNA gene was used as a model system to determine the effect of nucleosome assembly on nucleotide excision repair (NER) of the major UV photoproduct (cyclobutane pyrimidine dimer (CPD)) in DNA. Xenopus oocyte nuclear extracts were used to carry out repair in vitro on reconstituted, positioned 5 S rDNA nucleosomes. Nucleosome structure strongly inhibits NER at many CPD sites in the 5 S rDNA fragment while having little effect at a few sites. The time course of CPD removal at 35 different sites indicates that >85% of the CPDs in the naked DNA fragment have t(12) values <2 h, whereas <26% of the t(12) values in nucleosomes are <2 h, and 15% are >8 h. Moreover, removal of histone tails from these mononucleosomes has little effect on the repair rates. Finally, nucleosome inhibition of repair shows no correlation with the rotational setting of a 14-nucleotide-long pyrimidine tract located 30 base pairs from the nucleosome dyad. These results suggest that inhibition of NER by mononucleosomes is not significantly influenced by the rotational orientation of CPDs on the histone surface, and histone tails play little (or no) role in this inhibition. PMID:10821833

  14. Recent developments in brain tumor predisposing syndromes.

    PubMed

    Johansson, Gunnar; Andersson, Ulrika; Melin, Beatrice

    2016-01-01

    The etiologies of brain tumors are in the most cases unknown, but improvements in genetics and DNA screening have helped to identify a wide range of brain tumor predisposition disorders. In this review we are discussing some of the most common predisposition disorders, namely: neurofibromatosis type 1 and 2, schwannomatosis, rhabdoid tumor predisposition disorder, nevoid basal cell carcinoma syndrome (Gorlin), tuberous sclerosis complex, von Hippel-Lindau, Li-Fraumeni and Turcot syndromes. Recent findings from the GLIOGENE collaboration and the newly identified glioma causing gene POT1, will also be discussed. Genetics. We will describe these disorders from a genetic and clinical standpoint, focusing on the difference in clinical symptoms depending on the underlying gene or germline mutation. Central nervous system (CNS) tumors. Most of these disorders predispose the carriers to a wide range of symptoms. Herein, we will focus particularly on tumors affecting the CNS and discuss improvements of targeted therapy for the particular disorders. PMID:26634384

  15. Molecular cloning of the human nucleotide-excision-repair gene ERCC4

    SciTech Connect

    Thompson, L.H.; Brookman, K.W.; Weber, C.A.; Salazar, E.P.; Reardon, J.T.; Sancar, A.; Deng, Z.; Siciliano, M.J.

    1994-07-19

    ERCC4 was previously identified in somatic cell hybrids as a human gene that corrects the nucleotide-excision-repair deficiency in mutant hamster cells. The cloning strategy for ERCC4 involved transfection of the repair-deficient hamster cell line UV41 with a human sCos-1 cosmid library derived from chromosome 16. Enhanced UV resistance was seen with one cosmid-library transformant and two secondary transformants of UV41. Cosmid clones carrying a functional ERCC4 gene were isolated from a library of a second transformant by selecting in Escherichia coli for expression of a linked neomycin-resistance gene that was present in the sCos-1 vector. The cosmids mapped to 16p13.13-p13.2, the location assigned to ERCC4 by using somatic cell hybrids. Upon transfection into UV41, six cosmid clones gave partial correction ranging from 30% to 64%, although all appeared to contain the complete gene. The capacity for in vitro excision of thymine dimers from a plasmid by transformant cell extracts correlated qualitatively with enhanced UV resistance.

  16. Cloning and molecular characterization of the Chinese hamster ERCC2 nucleotide excision repair gene

    SciTech Connect

    Kirchner, J.M.; Salazar, E.P.; Lamerdin, J.E.

    1994-10-01

    The Chinese hamster ERCC2 nucleotide excision repair gene, encoding a presumed ATP-dependent DNA helicase, was cloned from the V79 cell line, and its nucleotide sequence was determined. The {approximately}15-kb gene comprises 23 exons with a 2283-base open reading frame. The predicted 760-amino-acid protein is 98% identical to the human ERCC2/EXP (760 amino acids), 51% identical to the Saccharomyces cerevisiae RAD3 (778 amino acids), and 54% identical to the Schizosaccharomyces pombe rad15 (772 amino acids) proteins. The promoter region of the hamster ERCC2 gene contains a pyrimidine-rich stretch (42 nucleotides, 88% C+T) similar to sequences found in the promoter regions of two other nucleotide excision repair genes, a GC box, a putative {alpha}-Pal transcription factor binding site, and two CAAT boxes. There is no apparent TAATA box. No consensus polyadenylation sequence (AATAAA or its variants) was found with 663 bases 3{prime} of the translation termination codon. 54 refs., 2 figs., 2 tabs.

  17. Polymorphisms in DNA repair genes, traffic-related polycyclic aromatic hydrocarbon exposure and breast cancer incidence.

    PubMed

    Mordukhovich, Irina; Beyea, Jan; Herring, Amy H; Hatch, Maureen; Stellman, Steven D; Teitelbaum, Susan L; Richardson, David B; Millikan, Robert C; Engel, Lawrence S; Shantakumar, Sumitra; Steck, Susan E; Neugut, Alfred I; Rossner, Pavel; Santella, Regina M; Gammon, Marilie D

    2016-07-15

    Vehicular traffic polycyclic aromatic hydrocarbons (PAHs) have been associated with breast cancer incidence in epidemiologic studies, including our own. Because PAHs damage DNA by forming adducts and oxidative lesions, genetic polymorphisms that alter DNA repair capacity may modify associations between PAH-related exposures and breast cancer risk. Our goal was to examine the association between vehicular traffic exposure and breast cancer incidence within strata of a panel of nine biologically plausible nucleotide excision repair (NER) and base excision repair (BER) genotypes. Residential histories of 1,508 cases and 1,556 controls were assessed in the Long Island Breast Cancer Study Project between 1996 and 1997 and used to reconstruct residential traffic exposures to benzo[a]pyrene, as a proxy for traffic-related PAHs. Likelihood ratio tests from adjusted unconditional logistic regression models were used to assess multiplicative interactions. A gene-traffic interaction was evident (p = 0.04) for ERCC2 (Lys751); when comparing the upper and lower tertiles of 1995 traffic exposure estimates, the odds ratio (95% confidence interval) was 2.09 (1.13, 3.90) among women with homozygous variant alleles. Corresponding odds ratios for 1960-1990 traffic were also elevated nearly 2-3-fold for XRCC1(Arg194Trp), XRCC1(Arg399Gln) and OGG1(Ser326Cys), but formal multiplicative interaction was not evident. When DNA repair variants for ERCC2, XRCC1 and OGG1 were combined, among women with 4-6 variants, the odds ratios were 2.32 (1.22, 4.49) for 1995 traffic and 2.96 (1.06, 8.21) for 1960-1990 traffic. Our study is first to report positive associations between traffic-related PAH exposure and breast cancer incidence among women with select biologically plausible DNA repair genotypes. PMID:26946191

  18. Evaluating the effects of genetic variants of DNA repair genes using cytogenetic mutagen sensitivity approaches

    PubMed Central

    Abdel-Rahman, Sherif Z.; El-Zein, Randa A.

    2011-01-01

    Mutagen sensitivity, measured in short-term cultures of peripheral blood lymphocytes by cytogenetic endpoints, is an indirect measure for DNA repair capacity and has been used for many years as a biomarker for intrinsic susceptibility for cancer. In this article, we briefly give an overview of the different cytogenetic mutagen sensitivity approaches that have been used successfully to evaluate the biological effects of polymorphisms in DNA repair genes based on a current review of the literature and based on the need for biomarkers that would allow the characterization of the biological and functional significance of such polymorphisms. We also address some of the future challenges facing this emerging area of research. PMID:21595606

  19. Altered Gene Expressions and Cytogenetic Repair Efficiency in Cells with Suppressed Expression of XPA after Proton Exposure

    NASA Technical Reports Server (NTRS)

    Zhang, Ye; Rohde, Larry H.; Gridley, Daila S.; Mehta, Satish K.; Pierson, Duane L.; Wu, Honglu

    2009-01-01

    Cellular responses to damages from ionizing radiation (IR) exposure are influenced not only by the genes involved in DNA double strand break (DSB) repair, but also by non- DSB repair genes. We demonstrated previously that suppressed expression of several non-DSB repair genes, such as XPA, elevated IR-induced cytogenetic damages. In the present study, we exposed human fibroblasts that were treated with control or XPA targeting siRNA to 250 MeV protons (0 to 4 Gy), and analyzed chromosome aberrations and expressions of genes involved in DNA repair. As expected, after proton irradiation, cells with suppressed expression of XPA showed a significantly elevated frequency of chromosome aberrations compared with control siRNA treated (CS) cells. Protons caused more severe DNA damages in XPA knock-down cells, as 36% cells contained multiple aberrations compared to 25% in CS cells after 4Gy proton irradiation. Comparison of gene expressions using the real-time PCR array technique revealed that expressions of p53 and its regulated genes in irradiated XPA suppressed cells were altered similarly as in CS cells, suggesting that the impairment of IR induced DNA repair in XPA suppressed cells is p53-independent. Except for XPA, which was more than 2 fold down regulated in XPA suppressed cells, several other DNA damage sensing and repair genes (GTSE1, RBBP8, RAD51, UNG and XRCC2) were shown a more than 1.5 fold difference between XPA knock-down cells and CS cells after proton exposure. The possible involvement of these genes in the impairment of DNA repair in XPA suppressed cells will be further investigated.

  20. Reduced Activity of Double-Strand Break Repair Genes in Prostate Cancer Patients With Late Normal Tissue Radiation Toxicity

    SciTech Connect

    Oorschot, Bregje van; Hovingh, Suzanne E.; Moerland, Perry D.; Medema, Jan Paul; Stalpers, Lukas J.A.; Vrieling, Harry; Franken, Nicolaas A.P.

    2014-03-01

    Purpose: To investigate clinical parameters and DNA damage response as possible risk factors for radiation toxicity in the setting of prostate cancer. Methods and Materials: Clinical parameters of 61 prostate cancer patients, 34 with (overresponding, OR) and 27 without (non-responding, NR) severe late radiation toxicity were assembled. In addition, for a matched subset the DNA damage repair kinetics (γ-H2AX assay) and expression profiles of DNA repair genes were determined in ex vivo irradiated lymphocytes. Results: Examination of clinical data indicated none of the considered clinical parameters to be correlated with the susceptibility of patients to develop late radiation toxicity. Although frequencies of γ-H2AX foci induced immediately after irradiation were similar (P=.32), significantly higher numbers of γ-H2AX foci were found 24 hours after irradiation in OR compared with NR patients (P=.03). Patient-specific γ-H2AX foci decay ratios were significantly higher in NR patients than in OR patients (P<.0001). Consequently, NR patients seem to repair DNA double-strand breaks (DSBs) more efficiently than OR patients. Moreover, gene expression analysis indicated several genes of the homologous recombination pathway to be stronger induced in NR compared with OR patients (P<.05). A similar trend was observed in genes of the nonhomologous end-joining repair pathway (P=.09). This is congruent with more proficient repair of DNA DSBs in patients without late radiation toxicity. Conclusions: Both gene expression profiling and DNA DSB repair kinetics data imply that less-efficient repair of radiation-induced DSBs may contribute to the development of late normal tissue damage. Induction levels of DSB repair genes (eg, RAD51) may potentially be used to assess the risk for late radiation toxicity.

  1. DNA repair gene polymorphisms and tobacco smoking in the risk for colorectal adenomas.

    PubMed

    Gao, Ying; Hayes, Richard B; Huang, Wen-Yi; Caporaso, Neil E; Burdette, Laurie; Yeager, Meredith; Chanock, Stephen J; Berndt, Sonja I

    2011-06-01

    DNA damage is thought to play a critical role in the development of colorectal adenoma. Variation in DNA repair genes may alter their capacity to correct endogenous and exogenous DNA damage. We explored the association between common single-nucleotide polymorphisms (SNPs) in DNA repair genes and adenoma risk with a case-control study nested in the Prostate, Lung, Colorectal and Ovarian Cancer Screening Trial. A total of 1338 left sided, advanced colorectal adenoma cases and 1503 matched controls free of left-sided polyps were included in the study. Using DNA extracted from blood, 3144 tag SNPs in 149 DNA repair genes were successfully genotyped. Among Caucasians, 30 SNPs were associated with adenoma risk at P < 0.01, with four SNPs remaining significant after gene-based adjustment for multiple testing. The most significant finding was for a non-synonymous SNP (rs9350) in Exonuclease-1 (EXO1) [odds ratio (OR) = 1.30, 95% confidence interval (CI) = 1.11-1.51, P = 0.001)], which was predicted to be damaging using bioinformatics methods. However, the association was limited to smokers with a strong risk for current smokers (OR = 2.15, 95% CI = 1.27-3.65) and an intermediate risk for former smokers (OR = 1.45, 95% CI = 1.14-1.82) and no association for never smokers (OR = 0.98, 95% CI = 0.76-1.25) (P(interaction) = 0.002). Among the top findings, an SNP (rs17503908) in ataxia telangiectasia mutated (ATM) was inversely related to adenoma risk (OR = 0.75, 95% CI = 0.63-0.91). The association was restricted to never smokers (OR = 0.55, 95% CI = 0.40-0.76) with no increased risk observed among smokers (OR = 0.89, 95% CI = 0.70-1.13) (P(interaction) = 0.006). This large comprehensive study, which evaluated all presently known DNA repair genes, suggests that polymorphisms in EXO1 and ATM may be associated with risk for advanced colorectal adenoma with the associations modified by tobacco-smoking status. PMID:21504893

  2. Divergent genes in potential inoculant Sinorhizobium strains are related to DNA replication, recombination, and repair.

    PubMed

    Penttinen, Petri; Greco, Dario; Muntyan, Victoria; Terefework, Zewdu; De Lajudie, Philippe; Roumiantseva, Marina; Becker, Anke; Auvinen, Petri; Lindström, Kristina

    2016-06-01

    To serve as inoculants of legumes, nitrogen-fixing rhizobium strains should be competitive and tolerant of diverse environments. We hybridized the genomes of symbiotically efficient and salt tolerant Sinorhizobium inoculant strains onto the Sinorhizobium meliloti Rm1021 microarray. The number of variable genes, that is, divergent or putatively multiplied genes, ranged from 503 to 1556 for S. meliloti AK23, S. meliloti STM 1064 and S. arboris HAMBI 1552. The numbers of divergent genes affiliated with the symbiosis plasmid pSymA and related to DNA replication, recombination and repair were significantly higher than expected. The variation was mainly in the accessory genome, implying that it was important in shaping the adaptability of the strains. PMID:26879331

  3. Use of the HPRT gene to study nuclease-induced DNA double-strand break repair

    PubMed Central

    Gravells, Polly; Ahrabi, Sara; Vangala, Rajani K.; Tomita, Kazunori; Brash, James T.; Brustle, Lena A.; Chung, Christopher; Hong, Julia M.; Kaloudi, Aikaterini; Humphrey, Timothy C.; Porter, Andrew C.G.

    2015-01-01

    Understanding the mechanisms of chromosomal double-strand break repair (DSBR) provides insight into genome instability, oncogenesis and genome engineering, including disease gene correction. Research into DSBR exploits rare-cutting endonucleases to cleave exogenous reporter constructs integrated into the genome. Multiple reporter constructs have been developed to detect various DSBR pathways. Here, using a single endogenous reporter gene, the X-chromosomal disease gene encoding hypoxanthine phosphoribosyltransferase (HPRT), we monitor the relative utilization of three DSBR pathways following cleavage by I-SceI or CRISPR/Cas9 nucleases. For I-SceI, our estimated frequencies of accurate or mutagenic non-homologous end-joining and gene correction by homologous recombination are 4.1, 1.5 and 0.16%, respectively. Unexpectedly, I-SceI and Cas9 induced markedly different DSBR profiles. Also, using an I-SceI-sensitive HPRT minigene, we show that gene correction is more efficient when using long double-stranded DNA than single- or double-stranded oligonucleotides. Finally, using both endogenous HPRT and exogenous reporters, we validate novel cell cycle phase-specific I-SceI derivatives for investigating cell cycle variations in DSBR. The results obtained using these novel approaches provide new insights into template design for gene correction and the relationships between multiple DSBR pathways at a single endogenous disease gene. PMID:26423459

  4. Rearrangement of RAG-1 recombinase gene in DNA-repair deficient ``wasted`` mice

    SciTech Connect

    Woloschak, G.E.; Libertin, C.R.; Weaver, P.; Churchill, M.; Chang-Liu, C.M.

    1993-11-01

    Mice recessive for the autosomal gene ``wasted`` wst display a disease pattern which includes increased sensitivity to the killing effects of ionizing radiation, immunodeficiency, and neurologic dysfunction. The recent cloning and characterization of recombinase genes (RAG-l/RAG-2) expressed in lymphoid and possibly central nervous system tissues prompted us to examine expression of these genes in DNA repair-deficient/immunodeficient wasted mice. Our results revealed expression of RAG-1 mRNA in spinal cord (but not brain) of control mice; no expression of RAG-1 mRNA was detected in spinal cord or brain from wst/wst mice or their normal littermates (wst/{center_dot}mice). In thymus tissue, a small RAG-1 transcript (1.0 kb) was detected in wst/wst mice that was not evident in thymus from control mice. In wst/{center_dot}mice, a two-fold increase in RAG-1 mRNA was evident in thymus tissue. RAG-2 mRNA could only be detected in thymus tissue from wst/{center_dot} and not from wst/wst or parental control BCF{sub 1} mice. Southern blots revealed a rearrangement/deletion within the RAG-1 gene of affected wasted mice, not evident in known strain-specific parental or littermate controls. These results support the idea that the RAG-1 gene may map at or near the locus for the wasted mutation. In addition, they suggest the importance of recombinase function in normal immune and central nervous system development as well as the potential contribution of this gene family to the normal repair of radiation-induced DNA damage.

  5. The democratization of gene editing: Insights from site-specific cleavage and double-strand break repair.

    PubMed

    Jasin, Maria; Haber, James E

    2016-08-01

    DNA double-strand breaks (DSBs) are dangerous lesions that if not properly repaired can lead to genomic change or cell death. Organisms have developed several pathways and have many factors devoted to repairing DSBs, which broadly occurs by homologous recombination, which relies on an identical or homologous sequence to template repair, or nonhomologous end-joining. Much of our understanding of these repair mechanisms has come from the study of induced DNA cleavage by site-specific endonucleases. In addition to their biological role, these cellular pathways can be co-opted for gene editing to study gene function or for gene therapy or other applications. While the first gene editing experiments were done more than 20 years ago, the recent discovery of RNA-guided endonucleases has simplified approaches developed over the years to make gene editing an approach that is available to the entire biomedical research community. Here, we review DSB repair mechanisms and site-specific cleavage systems that have provided insight into these mechanisms and led to the current gene editing revolution. PMID:27261202

  6. MMP3 and TIMP2 gene variants as predisposing factors for Achilles tendon pathologies: Attempted replication study in a British case-control cohort.

    PubMed

    El Khoury, Louis; Ribbans, William J; Raleigh, Stuart M

    2016-09-01

    Variants within the MMP3 (rs679620) and TIMP2 (rs4789932) genes have been associated with the risk of Achilles tendon pathology (ATP) in populations from South Africa and Australia. This study aimed to determine whether these variants were associated with the risk of ATP in British Caucasians. We recruited 118 cases with ATP, including a subset of 25 individuals with Achilles tendon rupture (RUP) and 131 controls. DNA samples were isolated from saliva and genotyped using qPCR. For the TIMP2 rs4789932 variant we found a significant (p = 0.038) difference in the genotype distribution frequency between males with ATP (CC, 39.4%; CT, 43.7%; TT, 16.9%) compared to male controls (CC, 20.7%; CT, 59.8%; TT, 19.5%). We also observed a difference in the TIMP2 rs4789932 genotype distribution between males with rupture compared to male controls (p = 0.038). The MMP3 rs679620 GG genotype was found to be overrepresented in the Achilles tendon rupture (RUP) group (AA, 24.0%; AG, 32.0%; GG, 44.0%) compared to controls (AA, 26.7%; AG, 54.2%; GG, 19.1%). In conclusion, the CT genotype of the TIMP2 rs4789932 variant was associated with lower risk of ATP in males. Furthermore, while we revealed differences for both variants in genotype distribution between the RUP and control groups, the sample size of the RUP group was small and confirmation would be required in additional cohorts. Finally, although both the TIMP2 rs4789932 and MMP3 rs679620 variants tentatively associated with ATP, there were differences in the direction of association compared to earlier work. PMID:27222816

  7. MMP3 and TIMP2 gene variants as predisposing factors for Achilles tendon pathologies: Attempted replication study in a British case–control cohort

    PubMed Central

    El Khoury, Louis; Ribbans, William J.; Raleigh, Stuart M.

    2016-01-01

    Variants within the MMP3 (rs679620) and TIMP2 (rs4789932) genes have been associated with the risk of Achilles tendon pathology (ATP) in populations from South Africa and Australia. This study aimed to determine whether these variants were associated with the risk of ATP in British Caucasians. We recruited 118 cases with ATP, including a subset of 25 individuals with Achilles tendon rupture (RUP) and 131 controls. DNA samples were isolated from saliva and genotyped using qPCR. For the TIMP2 rs4789932 variant we found a significant (p = 0.038) difference in the genotype distribution frequency between males with ATP (CC, 39.4%; CT, 43.7%; TT, 16.9%) compared to male controls (CC, 20.7%; CT, 59.8%; TT, 19.5%). We also observed a difference in the TIMP2 rs4789932 genotype distribution between males with rupture compared to male controls (p = 0.038). The MMP3 rs679620 GG genotype was found to be overrepresented in the Achilles tendon rupture (RUP) group (AA, 24.0%; AG, 32.0%; GG, 44.0%) compared to controls (AA, 26.7%; AG, 54.2%; GG, 19.1%). In conclusion, the CT genotype of the TIMP2 rs4789932 variant was associated with lower risk of ATP in males. Furthermore, while we revealed differences for both variants in genotype distribution between the RUP and control groups, the sample size of the RUP group was small and confirmation would be required in additional cohorts. Finally, although both the TIMP2 rs4789932 and MMP3 rs679620 variants tentatively associated with ATP, there were differences in the direction of association compared to earlier work. PMID:27222816

  8. A recessive syndrome of intellectual disability, moderate overgrowth, and renal dysplasia predisposing to Wilms tumor is caused by a mutation in FIBP gene.

    PubMed

    Akawi, Nadia; Ben-Salem, Salma; Lahti, Laura; Partanen, Juha; Ali, Bassam R; Al-Gazali, Lihadh

    2016-08-01

    Clinical classification of overgrowth syndromes represents a challenge since a wide spectrum of disorders result in marked overgrowth. Therefore, there is a continuous effort to identify the genetic basis of these disorders that will eventually facilitate their molecular classification. Here, we have identified the genetic etiology and the pathogenetic mechanism underlying a rare autosomal recessive overgrowth syndrome in three affected siblings. The overgrowth phenotype in the patients was accompanied by developmental delay, learning disabilities, and variable congenital abnormalities. To elucidate the genetic etiology of the disorder, whole-genome genotyping and whole-exome sequencing were used. The disease was mapped to 3p21.1-p14.2 and 11q13.1-q13.4, where an in-frame insertion (c.175_176insTAA) in FIBP gene was revealed. The resulting indel (p.H59LN) was predicted to change the protein conformation with likely deleterious effect on its function as one of the fibroblast growth factor signaling mediators. In vitro cellular proliferation assay and in situ hypridization in vivo were then performed to understand the pathophysiology of the disease. The patients' skin fibroblasts showed an increased proliferation capacity compared to the controls' explaining the observed overgrowth phenotype. In addition, we detected Fibp expression most notably in the brains of mice embryos suggesting a possible effect on cognitive functions early in development. To date, only one patient has been reported with a homozygous nonsense mutation in FIBP exhibiting an overgrowth syndrome with multiple congenital abnormalities. Taken all together, these findings provide convincing evidence implicating FIBP aberrations in the newly recognized overgrowth syndrome and expand the associated phenotypes to include possible Wilms tumor predisposition. © 2016 Wiley Periodicals, Inc. PMID:27183861

  9. DNA mismatch repair gene MSH6 implicated in determining age at natural menopause

    PubMed Central

    Perry, John R.B.; Hsu, Yi-Hsiang; Chasman, Daniel I.; Johnson, Andrew D.; Elks, Cathy; Albrecht, Eva; Andrulis, Irene L.; Beesley, Jonathan; Berenson, Gerald S.; Bergmann, Sven; Bojesen, Stig E.; Bolla, Manjeet K.; Brown, Judith; Buring, Julie E.; Campbell, Harry; Chang-Claude, Jenny; Chenevix-Trench, Georgia; Corre, Tanguy; Couch, Fergus J.; Cox, Angela; Czene, Kamila; D'adamo, Adamo Pio; Davies, Gail; Deary, Ian J.; Dennis, Joe; Easton, Douglas F.; Engelhardt, Ellen G.; Eriksson, Johan G.; Esko, Tõnu; Fasching, Peter A.; Figueroa, Jonine D.; Flyger, Henrik; Fraser, Abigail; Garcia-Closas, Montse; Gasparini, Paolo; Gieger, Christian; Giles, Graham; Guenel, Pascal; Hägg, Sara; Hall, Per; Hayward, Caroline; Hopper, John; Ingelsson, Erik; Kardia, Sharon L.R.; Kasiman, Katherine; Knight, Julia A.; Lahti, Jari; Lawlor, Debbie A.; Magnusson, Patrik K.E.; Margolin, Sara; Marsh, Julie A.; Metspalu, Andres; Olson, Janet E.; Pennell, Craig E.; Polasek, Ozren; Rahman, Iffat; Ridker, Paul M.; Robino, Antonietta; Rudan, Igor; Rudolph, Anja; Salumets, Andres; Schmidt, Marjanka K.; Schoemaker, Minouk J.; Smith, Erin N.; Smith, Jennifer A.; Southey, Melissa; Stöckl, Doris; Swerdlow, Anthony J.; Thompson, Deborah J.; Truong, Therese; Ulivi, Sheila; Waldenberger, Melanie; Wang, Qin; Wild, Sarah; Wilson, James F; Wright, Alan F.; Zgaga, Lina; Ong, Ken K.; Murabito, Joanne M.; Karasik, David; Murray, Anna

    2014-01-01

    The length of female reproductive lifespan is associated with multiple adverse outcomes, including breast cancer, cardiovascular disease and infertility. The biological processes that govern the timing of the beginning and end of reproductive life are not well understood. Genetic variants are known to contribute to ∼50% of the variation in both age at menarche and menopause, but to date the known genes explain <15% of the genetic component. We have used genome-wide association in a bivariate meta-analysis of both traits to identify genes involved in determining reproductive lifespan. We observed significant genetic correlation between the two traits using genome-wide complex trait analysis. However, we found no robust statistical evidence for individual variants with an effect on both traits. A novel association with age at menopause was detected for a variant rs1800932 in the mismatch repair gene MSH6 (P = 1.9 × 10−9), which was also associated with altered expression levels of MSH6 mRNA in multiple tissues. This study contributes to the growing evidence that DNA repair processes play a key role in ovarian ageing and could be an important therapeutic target for infertility. PMID:24357391

  10. DNA mismatch repair gene MSH6 implicated in determining age at natural menopause.

    PubMed

    Perry, John R B; Hsu, Yi-Hsiang; Chasman, Daniel I; Johnson, Andrew D; Elks, Cathy; Albrecht, Eva; Andrulis, Irene L; Beesley, Jonathan; Berenson, Gerald S; Bergmann, Sven; Bojesen, Stig E; Bolla, Manjeet K; Brown, Judith; Buring, Julie E; Campbell, Harry; Chang-Claude, Jenny; Chenevix-Trench, Georgia; Corre, Tanguy; Couch, Fergus J; Cox, Angela; Czene, Kamila; D'adamo, Adamo Pio; Davies, Gail; Deary, Ian J; Dennis, Joe; Easton, Douglas F; Engelhardt, Ellen G; Eriksson, Johan G; Esko, Tõnu; Fasching, Peter A; Figueroa, Jonine D; Flyger, Henrik; Fraser, Abigail; Garcia-Closas, Montse; Gasparini, Paolo; Gieger, Christian; Giles, Graham; Guenel, Pascal; Hägg, Sara; Hall, Per; Hayward, Caroline; Hopper, John; Ingelsson, Erik; Kardia, Sharon L R; Kasiman, Katherine; Knight, Julia A; Lahti, Jari; Lawlor, Debbie A; Magnusson, Patrik K E; Margolin, Sara; Marsh, Julie A; Metspalu, Andres; Olson, Janet E; Pennell, Craig E; Polasek, Ozren; Rahman, Iffat; Ridker, Paul M; Robino, Antonietta; Rudan, Igor; Rudolph, Anja; Salumets, Andres; Schmidt, Marjanka K; Schoemaker, Minouk J; Smith, Erin N; Smith, Jennifer A; Southey, Melissa; Stöckl, Doris; Swerdlow, Anthony J; Thompson, Deborah J; Truong, Therese; Ulivi, Sheila; Waldenberger, Melanie; Wang, Qin; Wild, Sarah; Wilson, James F; Wright, Alan F; Zgaga, Lina; Ong, Ken K; Murabito, Joanne M; Karasik, David; Murray, Anna

    2014-05-01

    The length of female reproductive lifespan is associated with multiple adverse outcomes, including breast cancer, cardiovascular disease and infertility. The biological processes that govern the timing of the beginning and end of reproductive life are not well understood. Genetic variants are known to contribute to ∼50% of the variation in both age at menarche and menopause, but to date the known genes explain <15% of the genetic component. We have used genome-wide association in a bivariate meta-analysis of both traits to identify genes involved in determining reproductive lifespan. We observed significant genetic correlation between the two traits using genome-wide complex trait analysis. However, we found no robust statistical evidence for individual variants with an effect on both traits. A novel association with age at menopause was detected for a variant rs1800932 in the mismatch repair gene MSH6 (P = 1.9 × 10(-9)), which was also associated with altered expression levels of MSH6 mRNA in multiple tissues. This study contributes to the growing evidence that DNA repair processes play a key role in ovarian ageing and could be an important therapeutic target for infertility. PMID:24357391

  11. Widespread transcriptional gene inactivation initiated by a repair intermediate of 8-oxoguanine

    PubMed Central

    Allgayer, Julia; Kitsera, Nataliya; Bartelt, Solveig; Epe, Bernd; Khobta, Andriy

    2016-01-01

    DNA damage can significantly modulate expression of the affected genes either by direct structural interference with transcription components or as a collateral outcome of cellular repair attempts. Thus, DNA glycosylases of the base excision repair (BER) pathway have been implicated in negative transcriptional response to several spontaneously generated DNA base modifications, including a common oxidative DNA base modification 8-oxoguanine (8-oxoG). Here, we report that single 8-oxoG situated in the non-transcribed DNA strand of a reporter gene has a pronounced negative effect on transcription, driven by promoters of various strength and with different structural properties, including viral, human, and artificial promoters. We further show that the magnitude of the negative effect on the gene expression correlates with excision of the modified base by OGG1 in all promoter constructs tested. Moreover, by using expression vectors with nuclease resistant backbone modifications, we demonstrate that OGG1 does not catalyse DNA strand cleavage in vivo. Rather, cleavage of the phosphate bond 5′ to 8-oxodG (catalysed by APE1) is essential and universally required for the onset of transcriptional silencing, regardless of the promoter structure. Hence, induction of transcriptional silencing emerges as a ubiquitous mode of biological response to 8-oxoG in DNA. PMID:27220469

  12. Expression of DNA repair genes in burned skin exposed to low-level red laser.

    PubMed

    Trajano, Eduardo Tavares Lima; Mencalha, Andre Luiz; Monte-Alto-Costa, Andréa; Pôrto, Luís Cristóvão; de Souza da Fonseca, Adenilson

    2014-11-01

    Although red laser lights lie in the region of non-ionizing radiations in the electromagnetic spectrum, there are doubts whether absorption of these radiations causes lesions in the DNA molecule. Our aim was to investigate the expression of the genes involved with base excision and nucleotide excision repair pathways in skin tissue submitted to burn injury and exposed to low-level red laser. Wistar rats were divided as follows: control group-rats burned and not irradiated, laser group-rats burned and irradiated 1 day after injury for five consecutive days, and later laser group-rats injured and treated 4 days after injury for five consecutive days. Irradiation was performed according to a clinical protocol (20 J/cm(2), 100 mW, continuous wave emission mode). The animals were sacrificed on day 10, and scarred tissue samples were withdrawn for total RNA extraction, complementary DNA (cDNA) synthesis, and evaluation of gene expression by quantitative polymerase chain reaction. Low-level red laser exposure (1) reduces the expression of APE1 messenger (mRNA), (2) increases the expression of OGG1 mRNA, (3) reduces the expression of XPC mRNA, and (4) increases the expression of XPA mRNA both in laser and later laser groups. Red laser exposure at therapeutic fluences alters the expression of genes related to base excision and nucleotide excision pathways of DNA repair during wound healing of burned skin. PMID:24930134

  13. Yeast RAD14 and human xeroderma pigmentosum group A DNA-repair genes encode homologous proteins.

    PubMed

    Bankmann, M; Prakash, L; Prakash, S

    1992-02-01

    Xeroderma pigmentosum (XP), a human autosomal recessive disorder, is characterized by extreme sensitivity to sunlight and high incidence of skin cancers. XP cells are defective in the incision step of excision repair of DNA damaged by ultraviolet light. Cell fusion studies have defined seven XP complementation groups, XP-A to XP-G. Similar genetic complexity of excision repair is observed in the yeast Saccharomyces cerevisiae. Mutations in any one of five yeast genes, RAD1, RAD2, RAD3, RAD4, and RAD10, cause a total defect in incision and an extreme sensitivity to ultraviolet light. Here we report the characterization of the yeast RAD14 gene. The available rad14 point mutant is only moderately ultraviolet-sensitive, and it performs a substantial amount of incision of damaged DNA. Our studies with the rad14 deletion (delta) mutation indicate an absolute requirement of RAD14 in incision. RAD14 encodes a highly hydrophilic protein of 247 amino acids containing zinc-finger motifs, and it is similar to the protein encoded by the human XPAC gene that complements XP group A cell lines. PMID:1741034

  14. Nucleotide Excision Repair Gene ERCC2 and ERCC5 Variants Increase Risk of Uterine Cervical Cancer

    PubMed Central

    Joo, Jungnam; Yoon, Kyong-Ah; Hayashi, Tomonori; Kong, Sun-Young; Shin, Hye-Jin; Park, Boram; Kim, Young Min; Hwang, Sang-Hyun; Kim, Jeongseon; Shin, Aesun; Kim, Joo-Young

    2016-01-01

    Purpose Defects in the DNA damage repair process can cause genomic instability and play an important role in cervical carcinogenesis. The purpose of this study was to analyze the association of 29 candidate single nucleotide polymorphisms (SNPs) in genes in the DNA repair pathway, TP53, and TP53BP1 with the risk of cervical cancer. Materials and Methods Twenty-nine SNPs in four genes in the DNA repair pathway (ERCC2, ERCC5, NBS1, and XRCC1), TP53, and TP53BP1 were genotyped for 478 cervical cancer patients and 922 healthy control subjects, and their effects on cervical carcinogenesis were analyzed. Results The most significant association was found for rs17655 in ERCC5, with an age-adjusted p-value < 0.0001, for which a strong additive effect of the risk allele C was observed (odds ratio, 2.01 for CC to GG). On the other hand, another significant polymorphism rs454421 in ERCC2 showed a dominant effect (odds ratio, 1.68 for GA+AA to GG) with an age-adjusted p-value of 0.0009. The association of these polymorphisms remained significant regardless of the age of onset. The significant result for rs17655 was also consistent for subgroups of patients defined by histology and human papillomavirus (HPV) types. However, for rs454421, the association was observed only in patients with squamous cell carcinoma and non-HPV 18 type. Conclusion The results of this study show a novel association of cervical cancer and the genes involved in the nucleotide excision pathway in the Korean population. PMID:26130668

  15. Single nucleotide polymorphisms in the DNA repair genes in HPV-positive cervical cancer.

    PubMed

    Bajpai, Deepti; Banerjee, Ayan; Pathak, Sujata; Thakur, Bhaskar; Jain, Sunesh K; Singh, Neeta

    2016-05-01

    Genetic variation in DNA repair genes can modulate DNA repair capacity and may be related to the risk of cancer. The human papillomavirus is considered to be a necessary but not sufficient cause for cervical cancer and, therefore, other factors contribute to the carcinogenesis. A hereditary component for this neoplasia has been reported. Evaluation of the association of six polymorphisms was carried out in the following DNA repair genes: XRCC1 (Arg194Trp, Arg280His, and Arg399Gln), ERCC1 (Asp118Asp), ERCC2 (Lys751Gln), and ERCC4 (Arg415Gln). The cases (n=110) included 65 squamous cell carcinomas (SCCs) and 45 squamous intraepithelial lesions (SIL). Controls (n=68) were recruited from among women without cervical abnormalities. Genotypes were determined by PCR-restriction fragment length polymorphism and DNA sequencing. A positive association was observed between the polymorphisms of XRCC1 genes, that is, in codons 194 [P=0.001, odds ratio (OR)=20.1, 95% confidence interval (CI)=5.9-68.8], 280 (P=0.001, OR=5.4, 95% CI=2.3-12.6), and 399 (P=0.008, OR=4.2, 95% CI=1.5-12.1) and cervical cancer. SIL patients also showed a significant association with codon 194 (P=0.012, OR=3.8, 95% CI=1.3-10.6), but not with 280 (P=0.35) and 399 (P=0.81). A positive correlation was also found in ERCC4 Gln415Gln in both SCCs and SILs (P=0.001, OR=21.3, 95% CI=7.1-64.0 and P=0.001, OR=7.8, 95% CI=2.9-20.9, respectively). For ERCC2 Gln751Gln, the association was significant for both SCCs (P=0.001, OR=10.1, 95% CI=2.6-37.9) and SILs (P=0.001, OR=8.9, 95% CI=2.8-28.3). However, the risk of SCC did not appear to differ significantly among individuals with the ERCC1 Asp118Asp genotype (P=0.404). For SILs, it appeared to be a protective genotype (95% CI=0.1-0.7). This study indicates that variant types of DNA repair genes play an important role in modifying individual susceptibility to SCC. PMID:25812040

  16. Polymorphisms in DNA repair genes, hair dye use, and the risk of non-Hodgkin lymphoma

    PubMed Central

    Guo, Huan; Bassig, Bryan A.; Lan, Qing; Zhu, Yong; Zhang, Yawei; Holford, Theodore R.; Leaderer, Brian; Boyle, Peter; Qin, Qin; Zhu, Cairong; Li, Ni; Rothman, Nathaniel

    2016-01-01

    Purpose Genetic polymorphisms in DNA repair genes and hair dye use may both have a role in the development of non-Hodgkin lymphoma (NHL). We aimed to examine the interaction between variants in DNA repair genes and hair dye use with risk of NHL in a population-based case– control study of Connecticut women. Methods We examined 24 single nucleotide polymorphisms in 16 DNA repair genes among 518 NHL cases and 597 controls and evaluated the associations between hair dye use and risk of overall NHL and common NHL subtypes, stratified by genotype, using unconditional logistic regression. Results Women who used hair dye before 1980 had a significantly increased risk of NHL, particularly for the follicular lymphoma (FL) subtype, but not for diffuse large B-cell lymphoma. The following genotypes in combination with hair dye use before 1980 were associated with FL risk: BRCA2 rs144848 AC+CC [odds ratio (OR) (95 % confidence interval (CI)) 3.28(1.27–8.50)], WRN rs1346044 TT [OR(95 % CI) 2.70(1.30–5.65)], XRCC3 rs861539 CT+TT [OR(95 % CI) 2.76(1.32–5.77)], XRCC4 rs1805377 GG [OR(95 % CI) 2.07(1.10–3.90)] and rs1056503 TT [OR(95 % CI) 2.17(1.16–4.07)], ERCC1 rs3212961 CC [OR(95 % CI) 1.93(1.00–3.72)], RAD23B rs1805329 CC [OR(95 % CI) 2.28(1.12–4.64)], and MGMT rs12917 CC, rs2308321 AA, and rs2308327 AA genotypes [OR(95 % CI) 1.96(1.06–3.63), 2.02(1.09–3.75), and 2.23(1.16–4.29), respectively]. In addition, a significant interaction with risk of overall NHL was observed between WRN rs1346044 and hair dye use before 1980 (pinteraction = 0.032). Conclusions Our results indicated that genetic variation in DNA repair genes modifies susceptibility to NHL in relation to hair dye use, particularly for the FL subtype and in women who began using hair dye before 1980. Further studies are needed to confirm these observations. PMID:25178586

  17. DNA repair gene polymorphisms and risk of adult meningioma, glioma, and acoustic neuroma.

    PubMed

    Rajaraman, Preetha; Hutchinson, Amy; Wichner, Sara; Black, Peter M; Fine, Howard A; Loeffler, Jay S; Selker, Robert G; Shapiro, William R; Rothman, Nathaniel; Linet, Martha S; Inskip, Peter D

    2010-01-01

    Although the etiology of primary brain tumors is largely unknown, prior studies suggest that DNA repair polymorphisms may influence risk of glioma. Altered DNA repair is also likely to affect the risk of meningioma and acoustic neuroma, but these tumors have not been well studied. We estimated the risk of glioma (n = 362), meningioma (n = 134), and acoustic neuroma (n = 69) in non-Hispanic whites with respect to 36 single nucleotide polymorphisms from 26 genes involved in DNA repair in a hospital-based, case-control study conducted by the National Cancer Institute. We observed significantly increased risk of meningioma with the T variant of GLTSCR1 rs1035938 (OR(CT/TT) = 3.5; 95% confidence interval: 1.8-6.9; P(trend) .0006), which persisted after controlling for multiple comparisons (P = .019). Significantly increased meningioma risk was also observed for the minor allele variants of ERCC4 rs1800067 (P(trend) .01); MUTYH rs3219466 (P(trend) .02), and PCNA rs25406 (P(trend) .03). The NBN rs1805794 minor allele variant was associated with decreased meningioma risk (P(trend) .006). Risk of acoustic neuroma was increased for the ERCC2 rs1799793 (P(trend) .03) and ERCC5 rs17655 (P(trend) .05) variants and decreased for the PARP1 rs1136410 (P(trend) .03). Decreased glioma risk was observed with the XRCC1 rs1799782 variant (P(trend) .04). Our results suggest that common DNA repair variants may affect the risk of adult brain tumors, especially meningioma. PMID:20150366

  18. Differential expression of thymic DNA repair genes in low-dose-rate irradiated AKR/J mice.

    PubMed

    Bong, Jin Jong; Kang, Yu Mi; Shin, Suk Chul; Choi, Seung Jin; Lee, Kyung Mi; Kim, Hee Sun

    2013-01-01

    We previously determined that AKR/J mice housed in a low-dose-rate (LDR) ((137)Cs, 0.7 mGy/h, 2.1 Gy) γ-irradiation facility developed less spontaneous thymic lymphoma and survived longer than those receiving sham or high-dose-rate (HDR) ((137)Cs, 0.8 Gy/min, 4.5 Gy) radiation. Interestingly, histopathological analysis showed a mild lymphomagenesis in the thymus of LDR-irradiated mice. Therefore, in this study, we investigated whether LDR irradiation could trigger the expression of thymic genes involved in the DNA repair process of AKR/J mice. The enrichment analysis of Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes pathways showed immune response, nucleosome organization, and the peroxisome proliferator-activated receptors signaling pathway in LDR-irradiated mice. Our microarray analysis and quantitative polymerase chain reaction data demonstrated that mRNA levels of Lig4 and RRM2 were specifically elevated in AKR/J mice at 130 days after the start of LDR irradiation. Furthermore, transcriptional levels of H2AX and ATM, proteins known to recruit DNA repair factors, were also shown to be upregulated. These data suggest that LDR irradiation could trigger specific induction of DNA repair-associated genes in an attempt to repair damaged DNA during tumor progression, which in turn contributed to the decreased incidence of lymphoma and increased survival. Overall, we identified specific DNA repair genes in LDR-irradiated AKR/J mice. PMID:23820165

  19. Platelet-Derived Growth Factor Gene Delivery Stimulates ex Vivo Gingival Repair

    PubMed Central

    ANUSAKSATHIEN, ORASA; WEBB, SARAH A.; JIN, QI-MING; GIANNOBILE, WILLIAM V.

    2008-01-01

    Destruction of tooth support due to the chronic inflammatory disease periodontitis is a major cause of tooth loss. There are limitations with available treatment options to tissue engineer soft tissue periodontal defects. The exogenous application of growth factors (GFs) such as platelet-derived growth factor (PDGF) has shown promise to enhance oral and periodontal tissue regeneration. However, the topical administration of GFs has not led to clinically significant improvements in tissue regeneration because of problems in maintaining therapeutic protein levels at the defect site. The utilization of PDGF gene transfer may circumvent many of the limitations with protein delivery to soft tissue wounds. The objective of this study was to test the effect of PDGF-A and PDGF-B gene transfer to human gingival fibroblasts (HGFs) on ex vivo repair in three-dimensional collagen lattices. HGFs were transduced with adenovirus encoding PDGF-A and PDGF-B genes. Defect fill of bilayer collagen gels was measured by image analysis of cell repopulation into the gingival defects. The modulation of gene expression at the defect site and periphery was measured by RT-PCR during a 10-day time course after gene delivery. The results demonstrated that PDGF-B gene transfer stimulated potent (>4-fold) increases in cell repopulation and defect fill above that of PDGF-A and corresponding controls. PDGF-A and PDGF-B gene expression was maintained for at least 10 days. PDGF gene transfer upregulated the expression of phosphatidylinosital 3-kinase and integrin α5 subunit at 5 days after adenovirus transduction. These results suggest that PDGF gene transfer has potential for periodontal soft tissue-engineering applications. PMID:13678451

  20. p63 and p73 Transcriptionally Regulate Genes Involved in DNA Repair

    PubMed Central

    Gurdziel, Katherine; Bell, George W.; Jacks, Tyler; Flores, Elsa R.

    2009-01-01

    The p53 family activates many of the same genes in response to DNA damage. Because p63 and p73 have structural differences from p53 and play distinct biological functions in development and metastasis, it is likely that they activate a unique transcriptional network. Therefore, we performed a genome-wide analysis using cells lacking the p53 family members after treatment with DNA damage. We identified over 100 genes involved in multiple pathways that were uniquely regulated by p63 or p73, and not p53. Further validation indicated that BRCA2, Rad51, and mre11 are direct transcriptional targets of p63 and p73. Additionally, cells deficient for p63 and p73 are impaired in DNA repair and p63+/−;p73+/− mice develop mammary tumors suggesting a novel mechanism whereby p63 and p73 suppress tumorigenesis. PMID:19816568

  1. Polymorphisms in DNA Repair Pathway Genes, Body Mass Index, and Risk of Non-Hodgkin Lymphoma

    PubMed Central

    Chen, Yingtai; Zheng, Tongzhang; Lan, Qing; Kim, Christopher; Qin, Qin; Foss, Francine; Chen, Xuezhong; Holford, Theodore; Leaderer, Brian; Boyle, Peter; Wang, Chengfeng; Dai, Min; Liu, Zhenjiang; Ma, Shuangge; Chanock, Stephen J.; Rothman, Nathaniel; Zhang, Yawei

    2013-01-01

    We conducted a population-based case-control study in Connecticut women to test the hypothesis that genetic variations in DNA repair pathway genes may modify the relationship between body mass index (BMI) and risk of non-Hodgkin lymphoma (NHL). Compared to those with BMI < 25, women with BMI ≥ 25 had significantly increased risk of NHL among women who carried BRCA1 (rs799917) CT/TT, ERCC2 (rs13181) AA, XRCC1 (rs1799782) CC, and WRN (rs1801195) GG genotypes, but no increase in NHL risk among women who carried BRCA1 CC, ERCC2 AC/CC, XRCC1 CT/TT, and WRN GT/TT genotypes. A significant interaction with BMI was only observed for WRN (rs1801195, P=0.004) for T-cell lymphoma and ERCC2 (rs13181, P=0.002) for diffuse large B-cell lymphoma. The results suggest that common genetic variation in DNA repair pathway genes may modify the association between BMI and NHL risk. PMID:23619945

  2. Tissue repair genes: the TiRe database and its implication for skin wound healing

    PubMed Central

    Yanai, Hagai; Budovsky, Arie; Tacutu, Robi; Barzilay, Thomer; Abramovich, Amir; Ziesche, Rolf; Fraifeld, Vadim E.

    2016-01-01

    Wound healing is an inherent feature of any multicellular organism and recent years have brought about a huge amount of data regarding regular and abnormal tissue repair. Despite the accumulated knowledge, modulation of wound healing is still a major biomedical challenge, especially in advanced ages. In order to collect and systematically organize what we know about the key players in wound healing, we created the TiRe (Tissue Repair) database, an online collection of genes and proteins that were shown to directly affect skin wound healing. To date, TiRe contains 397 entries for four organisms: Mus musculus, Rattus norvegicus, Sus domesticus, and Homo sapiens. Analysis of the TiRe dataset of skin wound healing-associated genes showed that skin wound healing genes are (i) over-conserved among vertebrates, but are under-conserved in invertebrates; (ii) enriched in extracellular and immuno-inflammatory genes; and display (iii) high interconnectivity and connectivity to other proteins. The latter may provide potential therapeutic targets. In addition, a slower or faster skin wound healing is indicative of an aging or longevity phenotype only when assessed in advanced ages, but not in the young. In the long run, we aim for TiRe to be a one-station resource that provides researchers and clinicians with the essential data needed for a better understanding of the mechanisms of wound healing, designing new experiments, and the development of new therapeutic strategies. TiRe is freely available online at http://www.tiredb.org. PMID:27049721

  3. Tissue repair genes: the TiRe database and its implication for skin wound healing.

    PubMed

    Yanai, Hagai; Budovsky, Arie; Tacutu, Robi; Barzilay, Thomer; Abramovich, Amir; Ziesche, Rolf; Fraifeld, Vadim E

    2016-04-19

    Wound healing is an inherent feature of any multicellular organism and recent years have brought about a huge amount of data regarding regular and abnormal tissue repair. Despite the accumulated knowledge, modulation of wound healing is still a major biomedical challenge, especially in advanced ages. In order to collect and systematically organize what we know about the key players in wound healing, we created the TiRe (Tissue Repair) database, an online collection of genes and proteins that were shown to directly affect skin wound healing. To date, TiRe contains 397 entries for four organisms: Mus musculus, Rattus norvegicus, Sus domesticus, and Homo sapiens. Analysis of the TiRe dataset of skin wound healing-associated genes showed that skin wound healing genes are (i) over-conserved among vertebrates, but are under-conserved in invertebrates; (ii) enriched in extracellular and immuno-inflammatory genes; and display (iii) high interconnectivity and connectivity to other proteins. The latter may provide potential therapeutic targets. In addition, a slower or faster skin wound healing is indicative of an aging or longevity phenotype only when assessed in advanced ages, but not in the young. In the long run, we aim for TiRe to be a one-station resource that provides researchers and clinicians with the essential data needed for a better understanding of the mechanisms of wound healing, designing new experiments, and the development of new therapeutic strategies. TiRe is freely available online at http://www.tiredb.org. PMID:27049721

  4. beta-Glucosidase as a reporter for the gene expression studies in Thermus thermophilus and constitutive expression of DNA repair genes.

    PubMed

    Ohta, Toshihiro; Tokishita, Shin-Ichi; Imazuka, Reiko; Mori, Ichiro; Okamura, Jin; Yamagata, Hideo

    2006-07-01

    Thermus thermophilus is an extremely thermophilic eubacterium that grows optimally at 70-75 degrees C. Because the frequency of DNA damage, such as deamination, depurination and single-strand breaks, increases as the temperature rises, the regulation of expression as well as the specificities and activities of T.thermophilus DNA repair systems are of particular interest. To study those systems, we developed a gene expression vector using the T.thermophilus beta-glucosidase gene (bgl) with host strain JOS9 (Deltabgl) derived from the T.thermophilus wild-type strain HB27. Since HB27 has two putative beta-galactosidase genes, the use of a single bgl gene as a reporter in combination with a Deltabgl host strain permits the study of gene expression against a low background level. We assayed Bgl activity with 2-nitrophenyl-beta-d-glucopyranoside as the substrate at 80 degrees C. We measured the expression of seven genes involved in DNA repair--three nucleotide excision repair genes (uvrA, uvrB and uvrC) and four recombinational repair genes (recA, ruvA, ruvB and ruvC). Expression levels of uvrA and uvrB were about three times those of uvrC, while those of ruvA, ruvB and ruvC were almost equal. Both ruvA and ruvC formed an operon with their adjacent 5'-upstream gene paaG and ftsQAZ, respectively. recA was transcribed as an operon of four genes, amt-cinA-ligT-recA. All seven DNA repair genes were expressed constitutively, and the DNA damaging agent mitomycin C did not increase their expression. PMID:16777922

  5. The study of the relation of DNA repair pathway genes SNPs and the sensitivity to radiotherapy and chemotherapy of NSCLC.

    PubMed

    Wang, Chunbo; Nie, Huan; Li, Yiqun; Liu, Guiyou; Wang, Xu; Xing, Shijie; Zhang, Liping; Chen, Xin; Chen, Yue; Li, Yu

    2016-01-01

    To analyze the relation between SNPs in DNA repair pathway-related genes and sensitivity of tumor radio-chemotherapy, 26 SNPs in 20 DNA repair genes were genotyped on 176 patients of NSCLC undertaking radio-chemotherapy treatment. In squamous cell carcinoma (SCC), as the rs2228000, rs2228001 (XPC), rs2273953 (TP73), rs2279744 (MDM2), rs2299939 (PTEN) and rs8178085, rs12334811 (DNA-PKcs) affected the sensitivity to chemotherapy, so did the rs8178085, rs12334811 to radiotherapy. Moreover rs344781, rs2273953 and rs12334811 were related with the survival time of SCC. In general, the "good" genotype GG (rs12334811) showed greater efficacy of radio-chemotherapy and MSF (24 months) on SCC. In adenocarcinoma, as the rs2699887 (PIK3), rs12334811 (DNA-PKcs) influenced the sensitivity to chemotherapy, so did the rs2299939, rs2735343 (PTEN) to radiotherapy. And rs402710, rs80270, rs2279744 and rs2909430 impacted the survival time of the adenocarcinoma patients. Both GG (rs2279744) and AG (rs2909430) showed a shorter survival time (MFS = 6). Additionally, some SNPs such as rs2228000, rs2228001 and rs344781 were found to regulate the expression of DNA repair pathway genes through eQTLs dataset analysis. These results indicate that SNPs in DNA repair pathway genes might regulate the expression and affect the DNA damage repair, and thereby impact the efficacy of radio-chemotherapy and the survival time of NSCLC. PMID:27246533

  6. The study of the relation of DNA repair pathway genes SNPs and the sensitivity to radiotherapy and chemotherapy of NSCLC

    PubMed Central

    Wang, Chunbo; Nie, Huan; Li, Yiqun; Liu, Guiyou; Wang, Xu; Xing, Shijie; Zhang, Liping; Chen, Xin; Chen, Yue; Li, Yu

    2016-01-01

    To analyze the relation between SNPs in DNA repair pathway-related genes and sensitivity of tumor radio-chemotherapy, 26 SNPs in 20 DNA repair genes were genotyped on 176 patients of NSCLC undertaking radio-chemotherapy treatment. In squamous cell carcinoma (SCC), as the rs2228000, rs2228001 (XPC), rs2273953 (TP73), rs2279744 (MDM2), rs2299939 (PTEN) and rs8178085, rs12334811 (DNA-PKcs) affected the sensitivity to chemotherapy, so did the rs8178085, rs12334811 to radiotherapy. Moreover rs344781, rs2273953 and rs12334811 were related with the survival time of SCC. In general, the “good” genotype GG (rs12334811) showed greater efficacy of radio-chemotherapy and MSF (24 months) on SCC. In adenocarcinoma, as the rs2699887 (PIK3), rs12334811 (DNA-PKcs) influenced the sensitivity to chemotherapy, so did the rs2299939, rs2735343 (PTEN) to radiotherapy. And rs402710, rs80270, rs2279744 and rs2909430 impacted the survival time of the adenocarcinoma patients. Both GG (rs2279744) and AG (rs2909430) showed a shorter survival time (MFS = 6). Additionally, some SNPs such as rs2228000, rs2228001 and rs344781 were found to regulate the expression of DNA repair pathway genes through eQTLs dataset analysis. These results indicate that SNPs in DNA repair pathway genes might regulate the expression and affect the DNA damage repair, and thereby impact the efficacy of radio-chemotherapy and the survival time of NSCLC. PMID:27246533

  7. Horizontal Gene Transfer Regulation in Bacteria as a “Spandrel” of DNA Repair Mechanisms

    PubMed Central

    Fall, Saliou; Mercier, Anne; Bertolla, Franck; Calteau, Alexandra; Gueguen, Laurent; Perrière, Guy; Vogel, Timothy M.; Simonet, Pascal

    2007-01-01

    Horizontal gene transfer (HGT) is recognized as the major force for bacterial genome evolution. Yet, numerous questions remain about the transferred genes, their function, quantity and frequency. The extent to which genetic transformation by exogenous DNA has occurred over evolutionary time was initially addressed by an in silico approach using the complete genome sequence of the Ralstonia solanacearum GMI1000 strain. Methods based on phylogenetic reconstruction of prokaryote homologous genes families detected 151 genes (13.3%) of foreign origin in the R. solanacearum genome and tentatively identified their bacterial origin. These putative transfers were analyzed in comparison to experimental transformation tests involving 18 different genomic DNA positions in the genome as sites for homologous or homeologous recombination. Significant transformation frequency differences were observed among these positions tested regardless of the overall genomic divergence of the R. solanacearum strains tested as recipients. The genomic positions containing the putative exogenous DNA were not systematically transformed at the highest frequencies. The two genomic “hot spots”, which contain recA and mutS genes, exhibited transformation frequencies from 2 to more than 4 orders of magnitude higher than positions associated with other genes depending on the recipient strain. These results support the notion that the bacterial cell is equipped with active mechanisms to modulate acquisition of new DNA in different genomic positions. Bio-informatics study correlated recombination “hot-spots” to the presence of Chi-like signature sequences with which recombination might be preferentially initiated. The fundamental role of HGT is certainly not limited to the critical impact that the very rare foreign genes acquired mainly by chance can have on the bacterial adaptation potential. The frequency to which HGT with homologous and homeologous DNA happens in the environment might have led

  8. Single-stranded oligonucleotide-mediated in vivo gene repair in the rd1 retina

    PubMed Central

    Andrieu-Soler, Charlotte; Halhal, Mounia; Boatright, Jeffrey H.; Padove, Staci A.; Nickerson, John M.; Stodulkova, Eva; Stewart, Rachael E.; Ciavatta, Vincent T.; Doat, Marc; Jeanny, Jean-Claude; de Bizemont, Therèse; Sennlaub, Florian; Courtois, Yves

    2007-01-01

    Purpose The aim of this study was to test whether oligonucleotide-targeted gene repair can correct the point mutation in genomic DNA of PDE6brd1 (rd1) mouse retinas in vivo. Methods Oligonucleotides (ODNs) of 25 nucleotide length and complementary to genomic sequence subsuming the rd1 point mutation in the gene encoding the β-subunit of rod photoreceptor cGMP-phosphodiesterase (β-PDE), were synthesized with a wild type nucleotide base at the rd1 point mutation position. Control ODNs contained the same nucleotide bases as the wild type ODNs but with varying degrees of sequence mismatch. We previously developed a repeatable and relatively non-invasive technique to enhance ODN delivery to photoreceptor nuclei using transpalpebral iontophoresis prior to intravitreal ODN injection. Three such treatments were performed on C3H/henJ (rd1) mouse pups before postnatal day (PN) 9. Treatment outcomes were evaluated at PN28 or PN33, when retinal degeneration was nearly complete in the untreated rd1 mice. The effect of treatment on photoreceptor survival was evaluated by counting the number of nuclei of photoreceptor cells and by assessing rhodopsin immunohistochemistry on flat-mount retinas and sections. Gene repair in the retina was quantified by allele-specific real time PCR and by detection of β-PDE-immunoreactive photoreceptors. Confirmatory experiments were conducted using independent rd1 colonies in separate laboratories. These experiments had an additional negative control ODN that contained the rd1 mutant nucleotide base at the rd1 point mutation site such that the sole difference between treatment with wild type and control ODN was the single base at the rd1 point mutation site. Results Iontophoresis enhanced the penetration of intravitreally injected ODNs in all retinal layers. Using this delivery technique, significant survival of photoreceptors was observed in retinas from eyes treated with wild type ODNs but not control ODNs as demonstrated by cell counting and

  9. AAV-Mediated Gene Editing via Double-Strand Break Repair

    PubMed Central

    Hirsch, Matthew L.; Samulski, R. Jude

    2016-01-01

    Traditionally, the ability to edit the mammalian genome was inhibited by the inherent low efficiency of homologous recombination (HR; approximately <1 in a million events) and the inability to deliver DNA efficiently to dividing and non-dividing cells/tissue. Despite these limitations, creative selections designed over 20 years ago, clearly demonstrated the powerful implications of gene knock-in and knockout technology for the genetic engineering of mice (Doetschman et al. Nat 330(6148): 576–578, 1987; Thomas and Capecchi. Cell 51(3): 503–512, 1987). The development and application of recombinant vectors based on adeno-associated virus (rAAV) have helped to overcome both of the initial limitations regarding DNA delivery and the frequency of HR. Considering DNA delivery, rAAV infects non-dividing and dividing cultured cells as well as most tissues in mouse and larger animal models (including humans). At the DNA editing level, rAAV genomes have been reported to increase the frequency of HR several orders of magnitude by serving as the repair substrate (Russell and Hirata. Nat Genet 18(4): 325–330, 1998). However, reports on the ability of rAAV genomes to stimulate HR, compared to plasmid DNA and oligonucleotides, are variable, and many labs have found it necessary to augment the frequency of rAAV-induced HR using site-specific endonucleases (Ellis et al. Gene Ther, 2012; Hirsch et al. Gene Ther 17(9): 1175–1180, 2010; Porteus et al. Mol Cell Biol 23(10): 3558–3565, 2003; Radecke et al. Mol Ther 14(6): 798–808, 2006). In this protocol, we describe a method to perform rAAV-mediated double-strand break (DSB) repair for precise genetic engineering in human cells. PMID:24557911

  10. Isolation of a human DNA repair gene by selection in Chinese hamster ovary cells

    SciTech Connect

    Ding, R.C.; Eastman, A.; Bresnick, E.

    1987-05-01

    Alkylation of DNA at the O/sup 6/-position of guanine represents a potent mutagenic and carcinogenic lesion. O/sup 6/-Methylguanine DNA methyltransferase is the repair system responsible for catalyzing the transfer of the methyl group to a cysteine of the protein in a suicide reaction. The gene controlling its expression in mammalian systems is designated mex. Resistance to chloroethylnitrosourea (CNU) is also mediated by this protein; this was used to select cells into which the max gene has been introduced. DNA purified from human liver has been transfected into mex/sup -/ CHO cells by the CaPO/sub 4/ method. pSV2gpt, containing a marker gene, gpt, was cotransfected. The transformed cells were initially selected for the expression of gpt (mycophenolic acid resistance) and reselected in CNU for mex/sup +/. Several clones were resistant to both demonstrating the linkage of these genes. A cosmid library was made from a mex/sup +/gpt/sup +/ clone and grown in a gpt/sup -/ strain of E. coli. gpt/sup +/ colonies were selected and the cosmid DNA rescued. One of the tested cosmid DNA's produced CNU resistance upon introduction into CHO cells. This cosmid was subcloned, restriction endonuclease-treated and a 5.3 kb fragment showed mex activity. This fragment is being further characterized and the DNA sequenced.

  11. Resveratrol improves bone repair by modulation of bone morphogenetic proteins and osteopontin gene expression in rats.

    PubMed

    Casarin, R C; Casati, M Z; Pimentel, S P; Cirano, F R; Algayer, M; Pires, P R; Ghiraldini, B; Duarte, P M; Ribeiro, F V

    2014-07-01

    This study investigated the effect of resveratrol on bone healing and its influence on the gene expression of osteogenic markers. Two calvarial defects were created and one screw-shaped titanium implant was inserted in the tibia of rats that were assigned to daily administration of placebo (control group, n=15) or 10mg/kg of resveratrol (RESV group, n=15) for 30 days. The animals were then sacrificed. One of the calvarial defects was processed for histomorphometric analysis and the tissue relative to the other was collected for mRNA quantification of bone morphogenetic protein (BMP)-2, BMP-7, osteopontin (OPN), bone sialoprotein (BSP), osteoprotegrin (OPG), and receptor activator of NF-κB ligand (RANKL). Implants were removed by applying a counter-torque force. Histomorphometric analysis revealed higher remaining defect in the calvarial defects of the control group than the RESV group (P=0.026). Resveratrol increased the counter-torque values of implant removal when compared to control therapy (P=0.031). Gene expression analysis showed a higher expression of BMP-2 (P=0.011), BMP-7 (P=0.049), and OPN (P=0.002) genes in the RESV group than in the control group. In conclusion, resveratrol improved the repair of critical-sized bone defects and the biomechanical retention of implants. Indeed, this natural agent may up-regulate the gene expression of important osteogenic markers. PMID:24530035

  12. DNA Repair Gene Polymorphism and the Risk of Mitral Chordae Tendineae Rupture

    PubMed Central

    Kalayci Yigin, Aysel; Bulent Vatan, Mehmet; Akdemir, Ramazan; Necati Murat Aksoy, Muhammed; Cakar, Mehmet Akif; Kilic, Harun; Erkorkmaz, Unal; Karacan, Keziban; Kaleli, Suleyman

    2015-01-01

    Polymorphisms in Lys939Gln XPC gene may diminish DNA repair capacity, eventually increasing the risk of carcinogenesis. The aim of the present study was to evaluate the significance of polymorphism Lys939Gln in XPC gene in patients with mitral chordae tendinea rupture (MCTR). Twenty-one patients with MCTR and thirty-seven age and sex matched controls were enrolled in the study. Genotyping of XPC gene Lys939Gln polymorphism was carried out using polymerase chain reaction- (PCR-) restriction fragment length polymorphism (RFLP). The frequencies of the heterozygote genotype (Lys/Gln-AC) and homozygote genotype (Gln/Gln-CC) were significantly different in MCTR as compared to control group, respectively (52.4% versus 43.2%, p = 0.049; 38.15% versus 16.2%, p = 0.018). Homozygote variant (Gln/Gln) genotype was significantly associated with increased risk of MCTR (OR = 2.059; 95% CI: 1.097–3.863; p = 0.018). Heterozygote variant (Lys/Gln) genotype was also highly significantly associated with increased risk of MCTR (OR = 1.489; 95% CI: 1.041–2.129; p = 0.049). The variant allele C was found to be significantly associated with MCTR (OR = 1.481; 95% CI: 1.101–1.992; p = 0.011). This study has demonstrated the association of XPC gene Lys939Gln polymorphism with MCTR, which is significantly associated with increased risk of MCTR. PMID:26604426

  13. Mismatch repair genes expression defects & association with clinicopathological characteristics in colorectal carcinoma

    PubMed Central

    Kaur, Gurjeet; Masoud, Abdelhafid; Raihan, N.; Radzi, M.; Khamizar, W.; Kam, Lee Suk

    2011-01-01

    Background & objectives: DNA mismatch repair gene (MMR) abnormalities are seen in 95 per cent of hereditary nonpolyposis colorectal cancer (HNPCC) and 10-15 per cent of sporadic colorectal cancers. There are no data on MMR abnormalities in Malaysian colorectal cancer patients. This study was aimed to determine the frequency of abnormal MMR gene protein expression in colorectal carcinoma in Northern Peninsular Malaysia using immunohistochemistry. Methods: Clinicopathological information was obtained from 148 patients’ records who underwent bowel resection for colorectal cancer (CRC) at the three hospitals in Malaysia. Immunohistochemistry for MLH1, MSH2, MSH6 and PMS2 proteins were performed on paraffin embedded tissue containing carcinoma. Results: A total of 148 subjects and 150 colorectal carcinomas of sporadic and hereditary types were assessed. Three patients had synchronous tumours. Twenty eight cancers (18.6%) from 26 subjects (17.6%) had absent immunohistochemical expression of any one of the MMR gene proteins. This comprised absent MLH1 only – 3 cancers, absent MSH2 only – 3, absent MSH6 only – 2, absent PMS2 only – 3, absent MLH1 and PMS2 – 14, absent MSH2 and MSH6 – 2 and absent MLH1, MSH6 and PMS2 – 1. There was significant association between abnormal MMR gene protein expression and proximal colon cancers, mucinous, signet ring and poorly differentiated morphology. Interpretation & conclusions: Cancers with abnormal MMR gene expression were associated with microsatellite instability-high (MSI-H) phenotype. About 15 per cent demonstrated absent MSH2, MSH6 and PMS2 protein expression in isolation or in combination with other MMR genes, which often predicts a germline mutation, synonymous with a diagnosis of HNPCC. This appears to be high frequency compared to reported data. PMID:21911971

  14. Variation in DNA Base Excision Repair Genes in Fuchs Endothelial Corneal Dystrophy

    PubMed Central

    Wójcik, Katarzyna A.; Synowiec, Ewelina; Polakowski, Piotr; Błasiak, Janusz; Szaflik, Jerzy; Szaflik, Jacek P.

    2015-01-01

    Background Fuchs endothelial corneal dystrophy (FECD) is a corneal disease characterized by abnormalities in the Descemet membrane and the corneal endothelium. The etiology of this disease is poorly understood. An increased level of oxidative DNA damage reported in FECD corneas suggests a role of DNA base excision repair (BER) genes in its pathogenesis. In this work, we searched for the association between variation of the PARP-1, NEIL1, POLG, and XRCC1 genes and FECD occurrence. Material/Methods This study was conducted on 250 FECD patients and 353 controls using polymerase chain reaction-restriction fragment length polymorphism, high-resolution melting analysis, and the TaqMan® SNP Genotyping Assay. Results We observed that the A/A genotype and the A allele of the c.1196A>G polymorphism of the XRCC1 gene were positively correlated with an increased FECD occurrence, whereas the G allele had the opposite effect. A weak association between the C/G genotype of the g.46438521G>C polymorphism of the NEIL1 gene and an increased incidence of FECD was also detected. Haplotypes of both polymorphisms of the XRCC1 were associated with FECD occurrence. No association of the c.2285T>C, c.–1370T>A and c.580C>T polymorphisms of the PARP-1, POLG and XRCC1 genes, respectively, with FECD occurrence was observed. Conclusions Our results suggest that the c.1196A>G polymorphism in the XRCC1 gene may be an independent genetic risk factor for FECD. PMID:26388025

  15. Dual requirement for the yeast MMS19 gene in DNA repair and RNA polymerase II transcription.

    PubMed Central

    Lauder, S; Bankmann, M; Guzder, S N; Sung, P; Prakash, L; Prakash, S

    1996-01-01

    Genetic and biochemical studies of Saccharomyces cerevisiae have indicated the involvement of a large number of protein factors in nucleotide excision repair (NER) of UV-damaged DNA. However, how MMS19 affects this process has remained unclear. Here, we report on the isolation of the MMS19 gene and the determination of its role in NER and other cellular processes. Genetic and biochemical evidence indicates that besides its function in NER, MMS19 also affects RNA polymerase II (Pol II) transcription. mms19delta cells do not grow at 37 degrees C, and mutant extract exhibits a thermolabile defect in Pol II transcription. Thus, Mms19 protein resembles TFIIH in that it is required for both transcription and DNA repair. However, addition of purified Mms19 protein does not alleviate the transcriptional defect of the mms19delta extract, nor does it stimulate the incision of UV-damaged DNA reconstituted from purified proteins. Interestingly, addition of purified TFIIH corrects the transcriptional defect of the mms19delta extract. Mms19 is, however, not a component of TFIIH or of Pol II holoenzyme. These and other results suggest that Mms19 affects NER and transcription by influencing the activity of TFIIH as an upstream regulatory element. It is proposed that mutations in the human MMS19 counterpart could result in syndromes in which both NER and transcription are affected. PMID:8943333

  16. Increasing frequencies of site-specific mutagenesis and gene targeting in Arabidopsis by manipulating DNA repair pathways.

    PubMed

    Qi, Yiping; Zhang, Yong; Zhang, Feng; Baller, Joshua A; Cleland, Spencer C; Ryu, Yungil; Starker, Colby G; Voytas, Daniel F

    2013-03-01

    Improved methods for engineering sequence-specific nucleases, including zinc finger nucleases (ZFNs) and TAL effector nucleases (TALENs), have made it possible to precisely modify plant genomes. However, the success of genome modification is largely dependent on the intrinsic activity of the engineered nucleases. In this study, we sought to enhance ZFN-mediated targeted mutagenesis and gene targeting (GT) in Arabidopsis by manipulating DNA repair pathways. Using a ZFN that creates a double-strand break (DSB) at the endogenous ADH1 locus, we analyzed repair outcomes in the absence of DNA repair proteins such as KU70 and LIG4 (both involved in classic nonhomologous end-joining, NHEJ) and SMC6B (involved in sister-chromatid-based homologous recombination, HR). We achieved a fivefold to 16-fold enhancement in HR-based GT in a ku70 mutant and a threefold to fourfold enhancement in GT in the lig4 mutant. Although the NHEJ mutagenesis frequency was not significantly changed in ku70 or lig4, DNA repair was shifted to microhomology-dependent alternative NHEJ. As a result, mutations in both ku70 and lig4 were predominantly large deletions, which facilitates easy screening for mutations by PCR. Interestingly, NHEJ mutagenesis and GT at the ADH1 locus were enhanced by sixfold to eightfold and threefold to fourfold, respectively, in a smc6b mutant. The increase in NHEJ-mediated mutagenesis by loss of SMC6B was further confirmed using ZFNs that target two other Arabidopsis genes, namely, TT4 and MPK8. Considering that components of DNA repair pathways are highly conserved across species, mutations in DNA repair genes likely provide a universal strategy for harnessing repair pathways to achieve desired targeted genome modifications. PMID:23282329

  17. A critical re-assessment of DNA repair gene promoter methylation in non-small cell lung carcinoma

    PubMed Central

    Do, Hongdo; Wong, Nicholas C.; Murone, Carmel; John, Thomas; Solomon, Benjamin; Mitchell, Paul L.; Dobrovic, Alexander

    2014-01-01

    DNA repair genes that have been inactivated by promoter methylation offer potential therapeutic targets either by targeting the specific repair deficiency, or by synthetic lethal approaches. This study evaluated promoter methylation status for eight selected DNA repair genes (ATM, BRCA1, ERCC1, MGMT, MLH1, NEIL1, RAD23B and XPC) in 56 non-small cell lung cancer (NSCLC) tumours and 11 lung cell lines using the methylation-sensitive high resolution melting (MS-HRM) methodology. Frequent methylation in NEIL1 (42%) and infrequent methylation in ERCC1 (2%) and RAD23B (2%) are reported for the first time in NSCLC. MGMT methylation was detected in 13% of the NSCLCs. Contrary to previous studies, methylation was not detected in ATM, BRCA1, MLH1 and XPC. Data from The Cancer Genome Atlas (TCGA) was consistent with these findings. The study emphasises the importance of using appropriate methodology for accurate assessment of promoter methylation. PMID:24569633

  18. Statin Therapy and the Expression of Genes that Regulate Calcium Homeostasis and Membrane Repair in Skeletal Muscle

    PubMed Central

    Draeger, Annette; Sanchez-Freire, Verónica; Monastyrskaya, Katia; Hoppeler, Hans; Mueller, Matthias; Breil, Fabio; Mohaupt, Markus G.; Babiychuk, Eduard B.

    2010-01-01

    In skeletal muscle of patients with clinically diagnosed statin-associated myopathy, discrete signs of structural damage predominantly localize to the T-tubular region and are suggestive of a calcium leak. The impact of statins on skeletal muscle of non-myopathic patients is not known. We analyzed the expression of selected genes implicated in the molecular regulation of calcium and membrane repair, in lipid homeostasis, myocyte remodeling and mitochondrial function. Microscopic and gene expression analyses were performed using validated TaqMan custom arrays on skeletal muscle biopsies of 72 age-matched subjects who were receiving statin therapy (n = 38), who had discontinued therapy due to statin-associated myopathy (n = 14), and who had never undergone statin treatment (n = 20). In skeletal muscle, obtained from statin-treated, non-myopathic patients, statins caused extensive changes in the expression of genes of the calcium regulatory and the membrane repair machinery, whereas the expression of genes responsible for mitochondrial function or myocyte remodeling was unaffected. Discontinuation of treatment due to myopathic symptoms led to a normalization of gene expression levels, the genes encoding the ryanodine receptor 3, calpain 3, and dystrophin being the most notable exceptions. Hence, even in clinically asymptomatic (non-myopathic) patients, statin therapy leads to an upregulation in the expression of genes that are concerned with skeletal muscle regulation and membrane repair. PMID:20489141

  19. Characterization of Chondrocyte Scaffold Carriers for Cell-based Gene Therapy in Articular Cartilage Repair

    PubMed Central

    Shui, Wei; Yin, Liangjun; Luo, Jeffrey; Li, Ruidong; Zhang, Wenwen; Zhang, Jiye; Huang, Wei; Hu, Ning; Liang, Xi; Deng, Zhong-Liang; Hu, Zhenming; Shi, Lewis; Luu, Hue H.; Haydon, Rex C.; He, Tong-Chuan; Ho, Sherwin

    2014-01-01

    Articular cartilage lesions in the knee are common injuries. Chondrocyte transplant represents a promising therapeutic modality for articular cartilage injuries. Here, we characterize the viability and transgene expression of articular chondrocytes cultured in 3-D scaffolds provided by four types of carriers. Articular chondrocytes are isolated from rabbit knees and cultured in four types of scaffolds: type I collagen sponge, fibrin glue, hyaluronan, and Open-cell PolyLactic Acid (OPLA). The cultured cells are transduced with adenovirus expressing green fluorescence protein (AdGFP) and luciferase (AdGL3-Luc). The viability and gene expression in the chondrocytes are determined with fluorescence microscopy and luciferase assay. Cartilage matrix production is assessed by Alcian blue staining. Rabbit articular chondrocytes are effectively infected by AdGFP and exhibited sustained GFP expression. All tested scaffolds support the survival and gene expression of the infected chondrocytes. However, the highest transgene expression is observed in the OPLA carrier. At four weeks, Alcian blue-positive matrix materials are readily detected in OPLA cultures. Thus, our results indicate that, while all tested carriers can support the survival of chondrocytes, OPLA supports the highest transgene expression and is the most conductive scaffold for matrix production, suggesting that OPLA may be a suitable scaffold for cell-based gene therapy of articular cartilage repairs. PMID:23629940

  20. ABCB5 is a limbal stem cell gene required for corneal development and repair

    PubMed Central

    Ksander, Bruce R.; Kolovou, Paraskevi E.; Wilson, Brian J.; Saab, Karim R.; Guo, Qin; Ma, Jie; McGuire, Sean P.; Gregory, Meredith S.; Vincent, William J. B.; Perez, Victor L.; Cruz-Guilloty, Fernando; Kao, Winston W. Y.; Call, Mindy K.; Tucker, Budd A.; Zhan, Qian; Murphy, George F.; Lathrop, Kira L.; Alt, Clemens; Mortensen, Luke J.; Lin, Charles P.; Zieske, James D.; Frank, Markus H.; Frank, Natasha Y.

    2014-01-01

    Corneal epithelial homeostasis and regeneration are sustained by limbal stem cells (LSCs)1–3, and LSC deficiency is a major cause of blindness worldwide4. Transplantation is often the only therapeutic option available to patients with LSC deficiency. However, while transplant success depends foremost on LSC frequency within grafts5, a gene allowing for prospective LSC enrichment has not been identified so far5. Here we show that ATP-binding cassette, sub-family B, member 5 (ABCB5)6,7 marks LSCs and is required for LSC maintenance, corneal development and repair. Furthermore, we demonstrate that prospectively isolated human or murine ABCB5-positive LSCs possess the exclusive capacity to fully restore the cornea upon grafting to LSC-deficient mice in xenogeneic or syngeneic transplantation models. ABCB5 is preferentially expressed on label-retaining LSCs2 in mice and p63α-positive LSCs8 in humans. Consistent with these findings, ABCB5-positive LSC frequency is reduced in LSC-deficient patients. Abcb5 loss of function in Abcb5 knockout mice causes depletion of quiescent LSCs due to enhanced proliferation and apoptosis, and results in defective corneal differentiation and wound healing. Our results from gene knockout studies, LSC tracing and transplantation models, as well as phenotypic and functional analyses of human biopsy specimens, provide converging lines of evidence that ABCB5 identifies mammalian LSCs. Identification and prospective isolation of molecularly defined LSCs with essential functions in corneal development and repair has important implications for the treatment of corneal disease, particularly corneal blindness due to LSC deficiency. PMID:25030174

  1. Repair of uv damaged DNA: Genes and proteins of yeast and human

    SciTech Connect

    Prakash, L.

    1992-04-01

    Our objectives are to determine the molecular mechanism of the incision step of excision repair of ultraviolet (UV) light damaged DNA in eukaryotic organisms, using the yeast Saccharomyces cerevisiae as a model system, and to study the human homologs of yeast excision repair and postreplication repair proteins progress is described.

  2. Characterisation of antimicrobial resistance-associated integrons and mismatch repair gene mutations in Salmonella serotypes.

    PubMed

    Yang, Baowei; Zheng, Jie; Brown, Eric W; Zhao, Shaohua; Meng, Jianghong

    2009-02-01

    In this study, we examined the presence of integrons and Salmonella genomic island 1 (SGI1) and assessed their contribution to antimicrobial resistance as well as determining the extent of the mutator phenotype in Salmonella isolates. A total of 81 Salmonella enterica serotype Typhimurium isolates were examined for the presence of integrons and SGI1 and for hypermutators using polymerase chain reaction (PCR) and the mutator assay, respectively. An additional 336 Salmonella isolates were also used to screen for hypermutators. Fourteen S. Typhimurium isolates carried class 1 integrons, of which six were shown to possess SGI1. Five putative mutators, S. Typhimurium ST20751, S. enterica serotype Heidelberg 22396 and S. enterica serotype Enteritidis 17929, 17929N and 17929R, were identified among the 417 Salmonella isolates. Complementation analysis with the wild-type mutH, mutL, mutS and uvrD genes indicated that none of the five mutators contained defective mismatch repair (MMR) system alleles. DNA sequence analysis revealed that single point mutations resulting in aspartic acid (codon 87) substitution in the gyrA gene conferred resistance to nalidixic acid and/or other fluoroquinolone drugs (ciprofloxacin and enrofloxacin) among four isolates. Our findings indicated that integrons and SGI1 play an important role in multidrug resistance in Salmonella. The incidence of hypermutators owing to defective MMR in Salmonella appears to be rare. PMID:19013057

  3. Targeted next-generation sequencing of 22 mismatch repair genes identifies Lynch syndrome families.

    PubMed

    Talseth-Palmer, Bente A; Bauer, Denis C; Sjursen, Wenche; Evans, Tiffany J; McPhillips, Mary; Proietto, Anthony; Otton, Geoffrey; Spigelman, Allan D; Scott, Rodney J

    2016-05-01

    Causative germline mutations in mismatch repair (MMR) genes can only be identified in ~50% of families with a clinical diagnosis of the inherited colorectal cancer (CRC) syndrome hereditary nonpolyposis colorectal cancer (HNPCC)/Lynch syndrome (LS). Identification of these patients are critical as they are at substantially increased risk of developing multiple primary tumors, mainly colorectal and endometrial cancer (EC), occurring at a young age. This demonstrates the need to develop new and/or more thorough mutation detection approaches. Next-generation sequencing (NGS) was used to screen 22 genes involved in the DNA MMR pathway in constitutional DNA from 14 HNPCC and 12 sporadic EC patients, plus 2 positive controls. Several softwares were used for analysis and functional annotation. We identified 5 exonic indel variants, 42 exonic nonsynonymous single-nucleotide variants (SNVs) and 1 intronic variant of significance. Three of these variants were class 5 (pathogenic) or class 4 (likely pathogenic), 5 were class 3 (uncertain clinical relevance) and 40 were classified as variants of unknown clinical significance. In conclusion, we have identified two LS families from the sporadic EC patients, one without a family history of cancer, supporting the notion for universal MMR screening of EC patients. In addition, we have detected three novel class 3 variants in EC cases. We have, in addition discovered a polygenic interaction which is the most likely cause of cancer development in a HNPCC patient that could explain previous inconsistent results reported on an intronic EXO1 variant. PMID:26811195

  4. Evaluation of DNA damage in COPD patients and its correlation with polymorphisms in repair genes

    PubMed Central

    2013-01-01

    Background We investigated a potential link between genetic polymorphisms in genes XRCC1 (Arg399Gln), OGG1 (Ser326Cys), XRCC3 (Thr241Met), and XRCC4 (Ile401Thr) with the level of DNA damage and repair, accessed by comet and micronucleus test, in 51 COPD patients and 51 controls. Methods Peripheral blood was used to perform the alkaline and neutral comet assay; and genetic polymorphisms by PCR/RFLP. To assess the susceptibility to exogenous DNA damage, the cells were treated with methyl methanesulphonate for 1-h or 3-h. After 3-h treatment the % residual damage was calculated assuming the value of 1-h treatment as 100%. The cytogenetic damage was evaluated by buccal micronucleus cytome assay (BMCyt). Results COPD patients with the risk allele XRCC1 (Arg399Gln) and XRCC3 (Thr241Met) showed higher DNA damage by comet assay. The residual damage was higher for COPD with risk allele in the four genes. In COPD patients was showed negative correlation between BMCyt (binucleated, nuclear bud, condensed chromatin and karyorrhexic cells) with pulmonary function and some variant genotypes. Conclusion Our results suggest a possible association between variant genotypes in XRCC1 (Arg399Gln), OGG1 (Ser326Cys), XRCC3 (Thr241Met), and XRCC4 (Ile401Thr), DNA damage and progression of COPD. PMID:24053728

  5. Cell and gene therapy for arrhythmias: Repair of cardiac conduction damage

    PubMed Central

    Xiao, Yong-Fu

    2011-01-01

    Action potentials generated in the sinoatrial node (SAN) dominate the rhythm and rate of a healthy human heart. Subsequently, these action potentials propagate to the whole heart via its conduction system. Abnormalities of impulse generation and/or propagation in a heart can cause arrhythmias. For example, SAN dysfunction or conduction block of the atrioventricular node can lead to serious bradycardia which is currently treated with an implanted electronic pacemaker. On the other hand, conduction damage may cause reentrant tachyarrhythmias which are primarily treated pharmacologically or by medical device-based therapies, including defibrillation and tissue ablation. However, drug therapies sometimes may not be effective or are associated with serious side effects. Device-based therapies for cardiac arrhythmias, even with well developed technology, still face inadequacies, limitations, hardware complications, and other challenges. Therefore, scientists are actively seeking other alternatives for antiarrhythmic therapy. In particular, cells and genes used for repairing cardiac conduction damage/defect have been investigated in various studies both in vitro and in vivo. Despite the complexities of the excitation and conduction systems of the heart, cell and gene-based strategies provide novel alternatives for treatment or cure of cardiac arrhythmias. This review summarizes some highlights of recent research progress in this field. PMID:22783301

  6. Influence of functional polymorphisms in DNA repair genes of myelodysplastic syndrome.

    PubMed

    Ribeiro, Howard Lopes; Soares Maia, Allan Rodrigo; Costa, Marília Braga; Farias, Izabelle Rocha; de Paula Borges, Daniela; de Oliveira, Roberta Taiane Germano; de Sousa, Juliana Cordeiro; Magalhães, Silvia Maria Meira; Pinheiro, Ronald Feitosa

    2016-09-01

    Myelodysplastic syndromes (MDS) are a heterogeneous group of hematopoietic stem cell (HSC) malignances characterized by peripheral cytopenias and predisposition to acute myeloid leukemia transformation. Several studies show that the MDS pathogenesis is a complex and heterogeneous process that involves multiple steps through a sequence of genetic lesions in the DNA which lead to functional changes in the cell and the emergence and subsequent evolution of pre-malignant clone. Double strand breaks (DSB) lesions are the most severe type of DNA damage in HSCs, which, if not properly repaired, might contribute to the development of chromosomal abnormalities, which in turn may lead to leukemia development. We assessed the mRNA expression levels of ATM, BRCA1, BRCA2, RAD51, XRCC5, XRCC6 and LIG4 genes in bone marrow samples of 47 MDS patients in order to evaluate the association with functional polymorphisms rs228593, rs4793191, rs9567623, rs1801320, rs3835, rs2267437 and rs1805388, respectively, and try to detect clinical associations. We found that the rs228593, rs2267437 and rs1805388 functional polymorphisms probably alter the level of expression of the ATM, XRCC6 and LIG4 genes, respectively, being important in the maintenance of genomic instability in MDS. PMID:27497341

  7. Necrotic enteritis predisposing factors in broiler chickens.

    PubMed

    Moore, Robert J

    2016-06-01

    Necrotic enteritis in chickens develops as a result of infection with pathogenic strains of Clostridium perfringens and the presence of predisposing factors. Predisposing factors include elements that directly change the physical properties of the gut, either damaging the epithelial surface, inducing mucus production, or changing gut transit times; factors that disrupt the gut microbiota; and factors that alter the immune status of birds. In the past research into necrotic enteritis predisposing factors was directed by the simple hypothesis that low-level colonization of C. perfringens commonly occurred within the gut of healthy chickens and the predisposing factors lead to a proliferation of those bacteria to produce disease. More recently, with an increasing understanding of the major virulence factors of C. perfringens and the application of molecular techniques to define different clades of C. perfringens strains, it has become clear that the C. perfringens isolates commonly found in healthy chickens are generally not strains that have the potential to cause disease. Therefore, we need to re-evaluate hypotheses regarding the development of disease, the origin of disease causing isolates of C. perfringens, and the importance of interactions with other C. perfringens strains and with predisposing factors. Many predisposing factors that affect the physical and immunological characteristics of the gastrointestinal tract may also change the resident microbiota. Research directed towards defining the relative importance of each of these different actions of predisposing factors will improve the understanding of disease pathogenesis and may allow refinement of experiment disease models. PMID:26926926

  8. Mutational analysis of the Drosophila DNA repair and recombination gene mei-9.

    PubMed Central

    Yildiz, Ozlem; Kearney, Hutton; Kramer, Benjamin C; Sekelsky, Jeff J

    2004-01-01

    Drosophila mei-9 is essential for several DNA repair and recombination pathways, including nucleotide excision repair (NER), interstrand crosslink repair, and meiotic recombination. To better understand the role of MEI-9 in these processes, we characterized 10 unique mutant alleles of mei-9. These include a P-element insertion that disrupts repair functions but not the meiotic function; three nonsense mutations, one of which has nearly wild-type levels of protein; three missense mutations, one of which disrupts the meiotic function but not repair functions; two small in-frame deletions; and one frameshift. PMID:15166153

  9. DNA repair gene ERCC1 polymorphisms may contribute to the risk of glioma.

    PubMed

    Yuan, Guanqian; Gao, Dandan; Ding, Shaofeng; Tan, Jun

    2014-05-01

    Polymorphisms in excision repair cross-complementing rodent repair deficiency complementation group 1 (ERCC1) gene have been shown to affect individual susceptibility to glioma, though studies have yielded conflicting results. This meta-analysis aims to derive a more precise estimation of the association between ERCC1 C8092A and C118T polymorphisms and glioma risk. A literature search of PubMed, Embase, Web of Science, Cochrane Library, and CBM databases was conducted to identify all eligible studies published before August 5, 2013. Crude odds ratios (ORs) with their corresponding confidence intervals (95% CIs) were used to assess the strength of this association. A meta-analysis was performed by reviewing seven studies on the C8092A polymorphism (2,978 cases and 4,051 controls) and four studies on the C118T polymorphism (1,390 Asian cases and 1,546 Asian controls). Pooled analysis yielded a significant association between the C8092A variant genotype and increased risk of glioma. As for ethnicity, the A allele was associated with increased risk of glioma in Asians, while no similar finding was observed in Caucasians. Stratified analyses by histological subtype indicated that the C8092A polymorphism showed a significant association with the risk of non-glioblastoma multiforme. For the C118T polymorphism, increased glioma susceptibility was also observed among Asians. Taken together, results from our meta-analysis support the view that common variants in ERCC1 may contribute to susceptibility to glioma, especially in Asians. However, further studies investigating the significance of these two polymorphisms as markers of susceptibility to and disease progression of glioma are still needed. PMID:24453030

  10. HLA-DQ2.5 genes associated with celiac disease risk are preferentially expressed with respect to non-predisposing HLA genes: Implication for anti-gluten T cell response.

    PubMed

    Pisapia, Laura; Camarca, Alessandra; Picascia, Stefania; Bassi, Virginia; Barba, Pasquale; Del Pozzo, Giovanna; Gianfrani, Carmen

    2016-06-01

    HLA genes represent the main risk factor in autoimmune disorders. In celiac disease (CD), the great majority of patients carry the HLA DQA1*05 and DQB1*02 alleles, both of which encode the DQ2.5 molecule. The formation of complexes between DQ2.5 and gluten peptides on antigen-presenting cells (APCs) is necessary to activate pathogenic CD4(+) T lymphocytes. It is widely accepted that the DQ2.5 genes establish the different intensities of anti-gluten immunity, depending whether they are in a homozygous or a heterozygous configuration. Here, we demonstrated that HLA DQA1*05 and DQB1*02 gene expression is much higher than expression of non-CD-associated genes. This influences the protein levels and causes a comparable cell surface exposure of DQ2.5 heterodimers between DQ2.5 homozygous and heterozygous celiac patients. As a consequence, the magnitude of the anti-gluten CD4(+) T cell response is strictly dependent on the antigen dose and not on the DQ2.5 gene configuration of APCs. Furthermore, our findings support the concept that the expression of DQ2.5 genes is an important risk factor in celiac disease. The preferential expression of DQ2.5 alleles provides a new functional explanation of why these genes are so frequently associated with celiac disease and with other autoimmune disorders. PMID:27083396

  11. NKX3.1 Suppresses TMPRSS2-ERG Gene Rearrangement and Mediates Repair of Androgen Receptor-Induced DNA Damage

    PubMed Central

    Bowen, Cai; Zheng, Tian; Gelmann, Edward P.

    2015-01-01

    TMPRSS2 gene rearrangements occur at DNA breaks formed during androgen receptor-mediated transcription and activate expression of ETS transcription factors at the early stages of more than half of prostate cancers. NKX3.1, a prostate tumor suppressor that accelerates the DNA repair response, binds to androgen receptor at the ERG gene breakpoint and inhibits both the juxtaposition of the TMPRSS2 and ERG gene loci and also their recombination. NKX3.1 acts by accelerating DNA repair after androgen-induced transcriptional activation. NKX3.1 influences the recruitment of proteins that promote homology-directed DNA repair. Loss of NKX3.1 favors recruitment to the ERG gene breakpoint of proteins that promote error-prone nonhomologous end-joining. Analysis of prostate cancer tissues showed that the presence of a TMPRSS2-ERG rearrangement was highly correlated with lower levels of NKX3.1 expression consistent with the role of NKX3.1 as a suppressor of the pathogenic gene rearrangement. PMID:25977336

  12. Genome-wide Transcriptome Profiling of Homologous Recombination DNA Repair

    PubMed Central

    Peng, Guang; Lin, Curtis Chun-Jen; Mo, Wei; Dai, Hui; Park, Yun-Yong; Kim, Soo-Mi; Peng, Yang; Mo, Qianxing; Siwko, Stefan; Hu, Ruozhen; Lee, Ju-Seog; Hennessy, Bryan; Hanash, Samir; Mills, Gordon B.; Lin, Shiaw-Yih

    2014-01-01

    Homologous recombination (HR) repair deficiency predisposes to cancer development, but also sensitizes cancer cells to DNA-damage-inducing therapeutics. Here we identify an HR-defect (HRD) gene signature, which can be used to functionally assess HR repair status without interrogating individual genetic alterations in cells. By using this HRD gene signature as a functional network analysis tool, we discover that simultaneous loss of two major tumor suppressors BRCA1 and PTEN extensively rewire the HR repair-deficient phenotype, which is found in cells with defects in either BRCA1 or PTEN alone. Moreover, the HRD gene signature serves as an effective drug discovery platform to identify agents targeting HR repair as potential chemo/radio-sensitizers. More importantly, this HRD gene signature is able to predict clinical outcomes across multiple cancer lineages. Our findings, therefore, provide a molecular profile of HR repair to assess its status at a functional network level, which can provide both biological insights and have clinical implications in cancer. PMID:24553445

  13. Downregulation of Homologous Recombination DNA Repair Genes by HDAC Inhibition in Prostate Cancer Is Mediated through the E2F1 Transcription Factor

    PubMed Central

    Kachhap, Sushant K.; Rosmus, Nadine; Collis, Spencer J.; Kortenhorst, Madeleine S. Q.; Wissing, Michel D.; Hedayati, Mohammad; Shabbeer, Shabana; Mendonca, Janet; Deangelis, Justin; Marchionni, Luigi; Lin, Jianqing; Höti, Naseruddin; Nortier, Johan W. R.; DeWeese, Theodore L.; Hammers, Hans; Carducci, Michael A.

    2010-01-01

    Background Histone deacetylase inhibitors (HDACis) re-express silenced tumor suppressor genes and are currently undergoing clinical trials. Although HDACis have been known to induce gene expression, an equal number of genes are downregulated upon HDAC inhibition. The mechanism behind this downregulation remains unclear. Here we provide evidence that several DNA repair genes are downregulated by HDAC inhibition and provide a mechanism involving the E2F1 transcription factor in the process. Methodology/Principal Findings Applying Analysis of Functional Annotation (AFA) on microarray data of prostate cancer cells treated with HDACis, we found a number of genes of the DNA damage response and repair pathways are downregulated by HDACis. AFA revealed enrichment of homologous recombination (HR) DNA repair genes of the BRCA1 pathway, as well as genes regulated by the E2F1 transcription factor. Prostate cancer cells demonstrated a decreased DNA repair capacity and an increased sensitization to chemical- and radio-DNA damaging agents upon HDAC inhibition. Recruitment of key HR repair proteins to the site of DNA damage, as well as HR repair capacity was compromised upon HDACi treatment. Based on our AFA data, we hypothesized that the E2F transcription factors may play a role in the downregulation of key repair genes upon HDAC inhibition in prostate cancer cells. ChIP analysis and luciferase assays reveal that the downregulation of key repair genes is mediated through decreased recruitment of the E2F1 transcription factor and not through active repression by repressive E2Fs. Conclusions/Significance Our study indicates that several genes in the DNA repair pathway are affected upon HDAC inhibition. Downregulation of the repair genes is on account of a decrease in amount and promoter recruitment of the E2F1 transcription factor. Since HDAC inhibition affects several pathways that could potentially have an impact on DNA repair, compromised DNA repair upon HDAC inhibition could

  14. Changes in Macrophage Phenotype and Induction of Epithelial-to-Mesenchymal Transition Genes Following Acute Achilles Tenotomy and Repair

    PubMed Central

    Sugg, Kristoffer B; Lubardic, Jovan; Gumucio, Jonathan P; Mendias, Christopher L

    2014-01-01

    Tendon injuries occur frequently in physically active individuals, but the clinical outcomes for these injuries can be poor. In many injured tissues the repair process is orchestrated by two types of cells, macrophages and fibroblasts. Macrophages, which have both proinflammatory (M1) and antiinflammatory (M2) phenotypes, can directly participate in tissue remodeling and direct the response of other cells through the secretion of cytokines and growth factors. In many organ systems, epithelial cells can transdifferentiate into fibroblasts, which can then regenerate damaged ECM. This process is triggered via activation of epithelial-to-mesenchymal transition (EMT) signaling programs. Most tendons are surrounded by sheets of epithelial cells, and these tissue layers could provide a source of fibroblasts to repair injured tendons. To gain greater insight into the biology of tendon repair, we performed a tenotomy and repair in Achilles tendons of adult rats and determined changes in macrophage phenotype, and ECM- and EMT-related genes over a four week time course. The results from this study suggest that changes in macrophage phenotype and activation of EMT-related programs likely contribute to the degradation and subsequent repair of injured tendon tissue. PMID:24700411

  15. Changes in macrophage phenotype and induction of epithelial-to-mesenchymal transition genes following acute Achilles tenotomy and repair.

    PubMed

    Sugg, Kristoffer B; Lubardic, Jovan; Gumucio, Jonathan P; Mendias, Christopher L

    2014-07-01

    Tendon injuries occur frequently in physically active individuals, but the clinical outcomes for these injuries can be poor. In many injured tissues the repair process is orchestrated by two types of cells, macrophages and fibroblasts. Macrophages, which have both pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes, can directly participate in tissue remodeling and direct the response of other cells through the secretion of cytokines and growth factors. In many organ systems, epithelial cells can trans-differentiate into fibroblasts, which can then regenerate damaged ECM. This process is triggered via activation of epithelial-to-mesenchymal transition (EMT) signaling programs. Most tendons are surrounded by sheets of epithelial cells, and these tissue layers could provide a source of fibroblasts to repair injured tendons. To gain greater insight into the biology of tendon repair, we performed a tenotomy and repair in Achilles tendons of adult rats and determined changes in macrophage phenotype, and ECM- and EMT-related genes over a 4-week time course. The results from this study suggest that changes in macrophage phenotype and activation of EMT-related programs likely contribute to the degradation and subsequent repair of injured tendon tissue. PMID:24700411

  16. Combination strategies for repair, plasticity, and regeneration using regulation of gene expression during the chronic phase after spinal cord injury.

    PubMed

    Gerin, Christine G; Madueke, Ikenna C; Perkins, Tina; Hill, Seritta; Smith, Kristin; Haley, Benjamin; Allen, Shannon A; Garcia, Richard P; Paunesku, Tanjana; Woloschak, Gayle

    2011-12-01

    Although recovery after spinal cord injury (SCI) is rare in humans, recent literature indicates that some patients do recover sensorimotor function years after the trauma. This study seeks to elucidate the genetic underpinnings of SCI repair through the investigation of neurodegenerative and regenerative associated genes involved in the response to SCI during the chronic phase in adult rats. Intervention on the level of gene regulation focused on enhancing naturally attempting SCI regenerative genes has the potential to promote SCI repair. Our aim was to analyze gene expression characteristics of candidate genes involved in the neuro-degenerative and -regenerative processes following various animal models of SCI. We compiled data showing gene expression changes after SCI in adult rats and created a chronological time-line of candidate genes differentially expressed during the chronic phase of SCI. Compiled data showed that SCI induced a transient upregulation of endogenous neuro-regenerative genes not only within a few hours but also within a few days, weeks, and months after SCI. For example, gene controlling growth-associated protein-43 (GAP-43), brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and others, showed significant changes in mRNA accumulation in SCI animals, from 48 hours to 12 weeks after SCI. Similarly, inhibitory genes, such as RhoA, LINGO-1, and others, were upregulated as late as 4 to 14 days after injury. This indicates that gene specific regulation changes, corresponding to repair and regenerative attempts, are naturally orchestrated over time after injury. These delayed changes after SCI give ample time for therapeutic gene modulation through upregulation or silencing of specific genes responsible for the synthesis of the corresponding biogenic proteins. By following the examination of differential gene regulation during the chronic phase, we have determined times, successions, co

  17. Mechanical properties and structure-function relationships in articular cartilage repaired using IGF-I gene-enhanced chondrocytes.

    PubMed

    Griffin, Darvin J; Ortved, Kyla F; Nixon, Alan J; Bonassar, Lawrence J

    2016-01-01

    Several studies have demonstrated the benefits of IGF-I gene therapy in enhancing the histologic and biochemical content of cartilage repaired by chondrocyte transplantation. However, there is little to no data on the mechanical performance of IGF-I augmented cartilage grafts. This study evaluated the compressive properties of full-thickness chondral defects in the equine femur repaired with and without IGF-I gene therapy. Animals were randomly assigned to one of three study cohorts based on chondrocyte treatment provided in each defect: (i) IGF-I gene delivered by recombinant adeno-associated virus (rAAV)-5; (ii) AAV-5 delivering GFP as a reporter; (iii) naïve cells without virus. In each case, the opposite limb was implanted with a fibrin carrier without cells. Samples were prepared for confined compression testing to measure the aggregate modulus and hydraulic permeability. All treatment groups, regardless of cell content or transduction, had mechanical properties inferior to native cartilage. Overexpression of IGF-I increased modulus and lowered permeability relative to other treatments. Investigation of structure-property relationships revealed that Ha and k were linearly correlated with GAG content but logarithmically correlated with collagen content. This provides evidence that IGF-I gene therapy can improve healing of articular cartilage and can greatly increase the mechanical properties of repaired grafts. PMID:26308948

  18. Frequency distribution of DNA repair genes ERCC1 and ERCC2 polymorphisms in South Indian healthy population.

    PubMed

    Rao, Katiboina Srinivasa; SureshKumar, Srinivasamurthy; Umamaheswaran, Gurusamy; Paul, Abialbon; Dubashi, Biswajit; Gunaseelan, Karunanithi; Dkhar, Steven Aibor

    2014-09-01

    DNA repair genes are crucial in maintaining the integrity of the whole genome. Single nucleotide polymorphisms (SNPs) in DNA repair genes have been attributed to the development of various cancers. SNPs of DNA repair genes (ERCC1 and ERCC2) have been implicated in the causation of various cancers as well as inter-individual variability in the therapeutic outcomes of platinum based therapy. Thus establishing the frequency of these functional SNPs in the healthy population is of significance. The present study was aimed to establish the allele and genotype frequencies of ERCC1 (19007C>T, rs11615; 8092C>A, rs3212986) and ERCC2 (Asp312Asn, rs1799793) genes in South Indian healthy population and to compare the data from HapMap populations. The study population consisted of 128 healthy South Indian unrelated individuals of either sex aged between 18 and 60 years. Standard phenol-chloroform method was used to extract DNA from peripheral leukocytes. The genotype of DNA repair gene polymorphisms was determined by quantitative real-time polymerase chain reaction using TaqMan genotyping assay. The observed frequency of the studied polymorphisms followed Hardy-Weinberg equilibrium (p>0.05). The frequencies of the minor alleles of the SNPs rs11615 (T), rs3212986 (A) and rs1799793 (A) were 43.8%, 29.3% and 35.6%, respectively. Gender-based analysis showed no significant difference in the frequency pattern. The observed allele and genotype frequencies showed significant ethnic difference between South Indians and other HapMap populations. This is the first study to provide the normative frequency data of allele and genotype distribution of three SNPs of ERCC1 and ERCC2 in South Indian healthy population. It might be useful in future genotype-phenotype association studies, especially for predicting the efficacy and adverse events of platinum based drugs. PMID:25155628

  19. Genetic polymorphisms in homologous recombination repair genes in healthy Slovenian population and their influence on DNA damage

    PubMed Central

    Goricar, Katja; Erculj, Nina; Zadel, Maja; Dolzan, Vita

    2012-01-01

    Background Homologous recombination (HR) repair is an important mechanism involved in repairing double-strand breaks in DNA and for maintaining genomic stability. Polymorphisms in genes coding for enzymes involved in this pathway may influence the capacity for DNA repair. The aim of this study was to select tag single nucleotide polymorphisms (SNPs) in specific genes involved in HR repair, to determine their allele frequencies in a healthy Slovenian population and their influence on DNA damage detected with comet assay. Materials and methods In total 373 individuals were genotyped for nine tag SNPs in three genes: XRCC3 722C>T, XRCC3 -316A>G, RAD51 -98G>C, RAD51 -61G>T, RAD51 1522T>G, NBS1 553G>C, NBS1 1197A>G, NBS1 37117C>T and NBS1 3474A>C using competitive allele-specific amplification (KASPar assay). Comet assay was performed in a subgroup of 26 individuals to determine the influence of selected SNPs on DNA damage. Results We observed that age significantly affected genotype frequencies distribution of XRCC3 -316A>G (P = 0.039) in healthy male blood donors. XRCC3 722C>T (P = 0.005), RAD51 -61G>T (P = 0.023) and NBS1 553G>C (P = 0.008) had a statistically significant influence on DNA damage. Conclusions XRCC3 722C>T, RAD51 -61G>T and NBS1 553G>C polymorphisms significantly affect the repair of damaged DNA and may be of clinical importance as they are common in Slovenian population. PMID:22933979

  20. Smoking and polymorphisms in xenobiotic metabolism and DNA repair genes are additive risk factors affecting bladder cancer in Northern Tunisia.

    PubMed

    Rouissi, Kamel; Ouerhani, Slah; Hamrita, Bechr; Bougatef, Karim; Marrakchi, Raja; Cherif, Mohamed; Ben Slama, Mohamed Riadh; Bouzouita, Mohamed; Chebil, Mohamed; Ben Ammar Elgaaied, Amel

    2011-12-01

    Cancer epidemiology has undergone marked development since the nineteen-fifties. One of the most spectacular and specific contributions was the demonstration of the massive effect of smoking and genetic polymorphisms on the occurrence of bladder cancer. The tobacco carcinogens are metabolized by various xenobiotic metabolizing enzymes, such as the super-families of N-acetyltransferases (NAT) and glutathione S-transferases (GST). DNA repair is essential to an individual's ability to respond to damage caused by tobacco carcinogens. Alterations in DNA repair genes may affect cancer risk by influencing individual susceptibility to this environmental exposure. Polymorphisms in NAT2, GST and DNA repair genes alter the ability of these enzymes to metabolize carcinogens or to repair alterations caused by this process. We have conducted a case-control study to assess the role of smoking, slow NAT2 variants, GSTM1 and GSTT1 null, and XPC, XPD, XPG nucleotide excision-repair (NER) genotypes in bladder cancer development in North Tunisia. Taken alone, each gene unless NAT2 did not appear to be a factor affecting bladder cancer susceptibility. For the NAT2 slow acetylator genotypes, the NAT2*5/*7 diplotype was found to have a 7-fold increased risk to develop bladder cancer (OR = 7.14; 95% CI: 1.30-51.41). However, in tobacco consumers, we have shown that Null GSTM1, Wild GSTT1, Slow NAT2, XPC (CC) and XPG (CC) are genetic risk factors for the disease. When combined together in susceptible individuals compared to protected individuals these risk factors give an elevated OR (OR = 61). So, we have shown a strong cumulative effect of tobacco and different combinations of studied genetic risk factors which lead to a great susceptibility to bladder cancer. PMID:21647780

  1. Association of DNA Repair Gene APE1 Asp148Glu Polymorphism with Breast Cancer Risk

    PubMed Central

    AlMutairi, Fatima; Ali Khan Pathan, Akbar; Alanazi, Mohammed; Shalaby, Manal; Alabdulkarim, Huda A.; Alamri, Abdullah; Al Naeem, Abdulrahman; Elrobh, Moammad; Shaik, Jilani P.; Khan, Wajahatullah; Khan, Zahid; Reddy Parine, Narasimha

    2015-01-01

    Objective. The aim of this study was to investigate the role of APE1 Asp148Glu polymorphism in breast cancer progression in Saudi population. Methods. We examined the genetic variations (rs1130409) in the DNA base excision repair gene APE1 at codon 148 (Asp148Glu) and its association with breast cancer risk using genotypic assays and in silico structural as well as functional predictions. In silico structural analysis was performed with Asp148Glu allele and compared with the predicted native protein structure. The wild and mutant 3D structures of APE1 were compared and analyzed using solvent accessibility models for protein stability confirmation. Results. Genotypic analysis of APE1 (rs1130409) showed statistically significant association of Asp148Glu with elevated susceptibility to breast cancer. The in silico analysis results indicated that the nsSNP Asp148Glu may cause changes in the protein structure and is associated with breast cancer risk. Conclusion. Taken together, this is the first report that established that Asp148Glu variant has structural and functional effect on the APE1 and may play an important role in breast cancer progression in Saudi population. PMID:26257461

  2. Pattern of antioxidant and DNA repair gene expression in normal airway epithelium associated with lung cancer diagnosis.

    PubMed

    Blomquist, Thomas; Crawford, Erin L; Mullins, D'Anna; Yoon, Youngsook; Hernandez, Dawn-Alita; Khuder, Sadik; Ruppel, Patricia L; Peters, Elizabeth; Oldfield, David J; Austermiller, Brad; Anders, John C; Willey, James C

    2009-11-15

    In previous studies, we reported that key antioxidant and DNA repair genes are regulated differently in normal bronchial epithelial cells of lung cancer cases compared with non-lung cancer controls. In an effort to develop a biomarker for lung cancer risk, we evaluated the transcript expressions of 14 antioxidant, DNA repair, and transcription factor genes in normal bronchial epithelial cells (HUGO names CAT, CEBPG, E2F1, ERCC4, ERCC5, GPX1, GPX3, GSTM3, GSTP1, GSTT1, GSTZ1, MGST1, SOD1, and XRCC1). A test comprising these 14 genes accurately identified the lung cancer cases in two case-control studies. The receiver operating characteristic-area under the curve was 0.82 (95% confidence intervals, 0.68-0.91) for the first case-control set (25 lung cancer cases and 24 controls), and 0.87 (95% confidence intervals, 0.73-0.96) for the second set (18 cases and 22 controls). For each gene included in the test, the key difference between cases and controls was altered distribution of transcript expression among cancer cases compared with controls, with more lung cancer cases expressing at both extremes among all genes (Kolmorogov-Smirnov test, D = 0.0795; P = 0.041). A novel statistical approach was used to identify the lower and upper boundaries of transcript expression that optimally classifies cases and controls for each gene. Based on the data presented here, there is an increased prevalence of lung cancer diagnosis among individuals that express a threshold number of key antioxidant, DNA repair, and transcription factor genes at either very high or very low levels in the normal airway epithelium. PMID:19887610

  3. Explorative study to identify novel candidate genes related to oxaliplatin efficacy and toxicity using a DNA repair array

    PubMed Central

    Kweekel, D M; Antonini, N F; Nortier, J W R; Punt, C J A; Gelderblom, H; Guchelaar, H-J

    2009-01-01

    Purpose: To identify new polymorphisms (single nucleotide polymorphisms, SNPs) in DNA repair pathways that are associated with efficacy and toxicity in patients receiving oxaliplatin and capecitabine for advanced colorectal cancer (ACC). Methods: We studied progression-free survival (PFS) in 91 ACC patients, of whom germ-line DNA was isolated and genotyped using an Asper Biotech array. Overall survival (OS) and toxicity were studied as secondary end points. A step-wise selection of SNPs was performed, involving univariate and multivariate log-rank tests and Cox regression analysis, with age and performance status as covariates. Results: A total of 81 SNPs in 46 genes on the array were selected for further analysis, based on genotyping success rates and minor allele frequencies. After step-wise selection, we found that homozygosity for the ataxia telangiectasia mutated gene (ATM) rs1801516 or excision repair cross-complementing gene (ERCC5) rs1047768 SNPs was associated with shorter PFS; however there were no significant associations (P>0.01) with OS or toxicity. Discussion: This is the first study describing the pathway gene approach for the selection of new candidate genes involved in oxaliplatin efficacy and toxicity. The results suggest that the ATM and ERCC5 genes may be associated with oxaliplatin efficacy in ACC. PMID:19536092

  4. Clinical heterogeneity within xeroderma pigmentosum associated with mutations in the DNA repair and transcription gene ERCC3

    SciTech Connect

    Vermeulen, W.; Kleijer, W.J.; Bootsma, D.; Hoeijmakers, J.H.J.; Weeda, G. ); Scott, R.J.; Rodgers, S.; Mueller, H.J. ); Cole, J.; Arlett, C.F. )

    1994-02-01

    The human DNA excision repair gene ERCC3 specifically corrects the nucleotide excision repair (NER) defect of xeroderma pigmentosum (XP) complementation group B. In addition to its function in NER, the ERCC3 DNA helicase was recently identified as one of the components of the human BTF2/TFIIH transcription factor complex, which is required for initiation of transcription of class II genes. To date, a single patient (XP11BE) has been assigned to this XP group B (XP-B), with the remarkable conjunction of two autosomal recessive DNA repair deficiency disorders: XP and Cockayne syndrome (CS). The intriguing involvement of the ERCC3 protein in the vital process of transcription may provide an explanation for the rarity, severity, and wide spectrum of clinical features in this complementation group. Here the authors report the identification of two new XP-B patients: XPCS1BA and XPCS2BA (siblings), by microneedle injection of the cloned ERCC3 repair gene as well as by cell hybridization. Molecular analysis of the ERCC3 gene in both patients revealed a single base substitution causing a missense mutation in a region that is completely conserved in yeast, Drosophila, mouse, and human ERCC3. As in patient XP11BE, the expression of only one allele (paternal) is detected. The mutation causes a virtually complete inactivation of the NER function of the protein. Despite this severe NER defect, both patients display a late onset of neurologic impairment, mild cutaneous symptoms, and a striking absence of skin tumors even at an age of >40 years. Analysis of the frequency of hprt[sup [minus

  5. A unique horizontal gene transfer event has provided the octocoral mitochondrial genome with an active mismatch repair gene that has potential for an unusual self-contained function

    PubMed Central

    2011-01-01

    Background The mitochondrial genome of the Octocorallia has several characteristics atypical for metazoans, including a novel gene suggested to function in DNA repair. This mtMutS gene is favored for octocoral molecular systematics, due to its high information content. Several hypotheses concerning the origins of mtMutS have been proposed, and remain equivocal, although current weight of support is for a horizontal gene transfer from either an epsilonproteobacterium or a large DNA virus. Here we present new and compelling evidence on the evolutionary origin of mtMutS, and provide the very first data on its activity, functional capacity and stability within the octocoral mitochondrial genome. Results The mtMutS gene has the expected conserved amino acids, protein domains and predicted tertiary protein structure. Phylogenetic analysis indicates that mtMutS is not a member of the MSH family and therefore not of eukaryotic origin. MtMutS clusters closely with representatives of the MutS7 lineage; further support for this relationship derives from the sharing of a C-terminal endonuclease domain that confers a self-contained mismatch repair function. Gene expression analyses confirm that mtMutS is actively transcribed in octocorals. Rates of mitochondrial gene evolution in mtMutS-containing octocorals are lower than in their hexacoral sister-group, which lacks the gene, although paradoxically the mtMutS gene itself has higher rates of mutation than other octocoral mitochondrial genes. Conclusions The octocoral mtMutS gene is active and codes for a protein with all the necessary components for DNA mismatch repair. A lower rate of mitochondrial evolution, and the presence of a nicking endonuclease domain, both indirectly support a theory of self-sufficient DNA mismatch repair within the octocoral mitochondrion. The ancestral affinity of mtMutS to non-eukaryotic MutS7 provides compelling support for an origin by horizontal gene transfer. The immediate vector of transmission

  6. DNA Methylation Leads to DNA Repair Gene Down-Regulation and Trinucleotide Repeat Expansion in Patient-Derived Huntington Disease Cells.

    PubMed

    Mollica, Peter A; Reid, John A; Ogle, Roy C; Sachs, Patrick C; Bruno, Robert D

    2016-07-01

    Huntington disease (HD) is an autosomal dominantly inherited disease that exhibits genetic anticipation of affected progeny due to expansions of a trinucleotide repeat (TNR) region within the HTT gene. DNA repair machinery is a known effector of TNR instability; however, the specific defects in HD cells that lead to TNR expansion are unknown. We hypothesized that HD cells would be deficient in DNA repair gene expression. To test this hypothesis, we analyzed expression of select DNA repair genes involved in mismatch/loop-out repair (APEX1, BRCA1, RPA1, and RPA3) in patient-derived HD cells and found each was consistently down-regulated relative to wild-type samples taken from unaffected individuals in the same family. Rescue of DNA repair gene expression by 5-azacytidine treatment identified DNA methylation as a mediator of DNA repair gene expression deficiency. Bisulfite sequencing confirmed hypermethylation of the APEX1 promoter region in HD cells relative to control, as well as 5-azacytidine-induced hypomethylation. 5-Azacytidine treatments also resulted in stabilization of TNR expansion within the mutant HTT allele during long-term culture of HD cells. Our findings indicate that DNA methylation leads to DNA repair down-regulation and TNR instability in mitotically active HD cells and offer a proof of principle that epigenetic interventions can curb TNR expansions. PMID:27182645

  7. Criteria and prediction models for mismatch repair gene mutations: a review.

    PubMed

    Win, Aung Ko; Macinnis, Robert J; Dowty, James G; Jenkins, Mark A

    2013-12-01

    One of the strongest predictors of colorectal cancer risk is carrying a germline mutation in a DNA mismatch repair (MMR) gene. Once identified, mutation carriers can be recommended for intensive screening that will substantially reduce their high colorectal cancer risk. Conversely, the relatives of carriers identified as non-carriers can be relieved of the burden of intensive screening. Criteria and prediction models that identify likely mutation carriers are needed for cost-effective, targeted, germline testing for MMR gene mutation. We reviewed 12 criteria/guidelines and 8 prediction models (Leiden, Amsterdam-plus, Amsterdam-alternative, MMRpro, PREMM1,2,6, MMRpredict, Associazione Italiana per lo studio della Familiarità ed Ereditarietà dei tumori Gastrointestinali (AIFEG) and the Myriad Genetics Prevalence table) for identifying mutation carriers. While criteria are only used to identify individuals with colorectal cancer (yes/no for screening followed by germline testing), all prediction models except MMRpredict and Myriad tables can predict the probability of carrying mutations for individuals with or without colorectal cancer. We conducted a meta-analysis of the discrimination performance of 17 studies that validated the prediction models. The pooled estimate for the area under curve was 0.80 (95% CI 0.72 to 0.88) for MMRpro, 0.81 (95% CI 0.73 to 0.88) for MMRpredict, 0.84 (95% CI 0.81 to 0.88) for PREMM, and 0.85 (95% CI 0.78 to 0.91) for Leiden model. Given the high degree of overlap in the CIs, we cannot state that one model has a higher discrimination than any of the others. Overall, the existing statistical models have been shown to be sensitive and specific (at a 5% cut-off) in predicting MMR gene mutation carriers. Future models may need to: provide prediction of PMS2 mutations, take into account a wider range of Lynch syndrome-associated cancers when assessing family history, and be applicable to all people irrespective of any cancer diagnosis

  8. Two DNA repair and recombination genes in Saccharomyces cerevisiae, RAD52 and RAD54, are induced during meiosis

    SciTech Connect

    Cole, G.M.; Mortimer, R.K. ); Schild, D. )

    1989-07-01

    The DNA repair and recombination genes of Saccharomyces cerevisiae, RAD52 and RAD54, were transcriptionally induced approximately 10- to 15-fold in sporulating MATa/{alpha} cells. Congenic MATa/a cells, which did not sporulate, did not show similar increases. Assays of {beta}-galactosidase activity in strains harboring either a RAD52- or RAD54-lacZ gene fusion indicated that this induction occurred at a time concomitant with a commitment to meiotic recombination, as measured by prototroph formation from his1 heteroalleles.

  9. Single-nucleotide polymorphisms in DNA repair genes and association with breast cancer risk in the web study.

    PubMed

    Roberts, Michelle R; Shields, Peter G; Ambrosone, Christine B; Nie, Jing; Marian, Catalin; Krishnan, Shiva S; Goerlitz, David S; Modali, Ramakrishna; Seddon, Michael; Lehman, Teresa; Amend, Kandace L; Trevisan, Maurizio; Edge, Stephen B; Freudenheim, Jo L

    2011-08-01

    Base excision repair (BER) and nucleotide excision repair (NER) pathways repair damaged DNA, and polymorphisms in these genes might affect breast cancer susceptibility. We evaluated associations between seven single-nucleotide polymorphisms in four DNA repair genes (ERCC4 rs1799801, XPC rs2227998, rs2228001, rs2228000, OGG1 rs1052133 and XRCC1 rs25487 and rs25486) and breast cancer risk, examining modification by smoking and alcohol consumption, using data from the Western New York Exposures and Breast Cancer Study. Women aged 35-79 years with incident breast cancer (n = 1170) and age- and race-matched controls (n = 2115) were enrolled. Genotyping was performed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Unconditional logistic regression was used to estimate odds ratios (OR) and 95% confidence intervals (CIs). No significant associations were observed in premenopausal women. Among postmenopausal women, rs25487 and rs25486 (OR = 1.24; 95% CI 1.01-1.51 and OR = 1.23; 95% CI 1.01-1.49, respectively, for combined heterozygous and homozygous variant compared with reference) were associated with increased risk of breast cancer. Postmenopausal women carrying the variant allele of the synonymous XPC polymorphism (rs2227998) were also at borderline significantly increased risk (OR = 1.24; 95% CI 1.01-1.52, heterozygous variant compared with reference; OR = 1.22; 95% CI 1.01-1.48, for combined heterozygous and homozygous variant compared with reference). There was no evidence of genotype-smoking and genotype-alcohol consumption interactions for pre- and postmenopausal women. These results indicate that some of the variants in BER and NER genes may influence risk of postmenopausal breast cancer. PMID:21622940

  10. Ovarian steroids regulate gene expression related to DNA repair and neurodegenerative diseases in serotonin neurons of macaques

    PubMed Central

    Bethea, Cynthia L.; Reddy, Arubala P.

    2015-01-01

    Depression often accompanies the peri-menopausal transition and it often precedes overt symptomology in common neurodegenerative diseases (NDD; Alzheimer's, Parkinson's, Huntington, ALS). Serotonin dysfunction is frequently found in the different etiologies of depression. We have shown that ovariectomized (Ovx) monkeys treated with estradiol (E) for 28 days supplement with placebo or progesterone (P) on days 14-28 had reduced DNA fragmentation in serotonin neurons of the dorsal raphe nucleus; and long-term Ovx monkeys had fewer serotonin neurons than intact controls. We questioned the effect of E alone or E+P on gene expression related to DNA repair, protein folding (chaperones), the ubiquitin proteosome, axon transport, and NDD specific genes in serotonin neurons. Ovx macaques were treated with placebo, E or E+P (n=3/group) for 1 month. Serotonin neurons were laser captured and subjected to microarray analysis and qRT-PCR. Increases were confirmed with qRT-PCR in 5 genes that code for proteins involved in repair of strand breaks and nucleotide excision. NBN1, PCNA, GADD45A, RAD23A and GTF2H5 significantly increased with E or E+P treatment (all ANOVA p< 0.01). Chaperone genes HSP70, HSP60 and HSP27 significantly increased with E or E+P treatment (all ANOVA p<0.05). HSP90 showed a similar trend. Ubiquinase coding genes UBEA5, UBE2D3 and UBE3A (Parkin) increased with E or E+P (all ANOVA p<0.003). Transport related genes coding kinesin, dynein, and dynactin increased with E or E+P (all ANOVA p<0.03). SCNA (α synuclein) and ADAM10 (α secretase) increased (both ANOVA p<0.02), but PSEN1 (presenilin1) decreased (ANOVA p<0.02) with treatment. APP decreased 10-fold with E or E+P administration. Newman-Keuls posthoc comparisons indicated variation in the response to E alone versus E+P across the different genes. In summary, E or E+P increased gene expression for DNA repair mechanisms in serotonin neurons, thereby rendering them less vulnerable to stress-induced DNA

  11. Evaluation of cell proliferation, apoptosis, and dna-repair genes as potential biomarkers for ethanol-induced cns alterations

    PubMed Central

    2012-01-01

    Background Alcohol use disorders (AUDs) lead to alterations in central nervous system (CNS) architecture along with impaired learning and memory. Previous work from our group and that of others suggests that one mechanism underlying these changes is alteration of cell proliferation, apoptosis, and DNA-repair in neural stem cells (NSCs) produced as a consequence of ethanol-induced effects on the expression of genes related to p53-signaling. This study tests the hypothesis that changes in the expression of p53-signaling genes represent biomarkers of ethanol abuse which can be identified in the peripheral blood of rat drinking models and human AUD subjects and posits that specific changes may be correlated with differences in neuropsychological measures and CNS structure. Results Remarkably, microarray analysis of 350 genes related to p53-signaling in peripheral blood leukocytes (PBLs) of binge-drinking rats revealed 190 genes that were significantly altered after correcting for multiple testing. Moreover, 40 of these genes overlapped with those that we had previously observed to be changed in ethanol-exposed mouse NSCs. Expression changes in nine of these genes were tested for independent confirmation by a custom QuantiGene Plex (QGP) assay for a subset of p53-signaling genes, where a consistent trend for decreased expression of mitosis-related genes was observed. One mitosis-related gene (Pttg1) was also changed in human lymphoblasts cultured with ethanol. In PBLs of human AUD subjects seven p53-signaling genes were changed compared with non-drinking controls. Correlation and principal components analysis were then used to identify significant relationships between the expression of these seven genes and a set of medical, demographic, neuropsychological and neuroimaging measures that distinguished AUD and control subjects. Two genes (Ercc1 and Mcm5) showed a highly significant correlation with AUD-induced decreases in the volume of the left parietal supramarginal

  12. [Comparison between gene therapy and gradual release carrier for bone morphogenetic protein-2 in repairing bone defects].

    PubMed

    Li, Jianjun; Bai, Lunhao; Cui, Shaoqian; Wang, Huan; Xu, Xinxiang

    2007-06-01

    To compare the effects between gene therapy and gradual release carrier for bone morphogenetic protein-2 (BMP-2) in repairing bone defects, bone defects for 15 mm were created.on the bilateral radius in rabbits and treated with four kinds of implantations, ie, composite of transgeneic MSCs and PLA/PCL (Group A), composite of MSCs and gradual release carrier for BMP-2 (Group B), composite of MSCs and PLA/PCL (Group C), and PLA/PCL alone (Group D). After 4, 8, and 12 weeks of the operations, X-ray, histological examination, biomechanics analysis, and bone density measurement were conducted. Results showed that both osteoblasts and mesenchymal cells displayed strongly positive expression of BMP-2 in Group A after 4 weeks of the operation, the speed and quality of bone formation in Group A were much better than those in Group B. After 12 weeks of the operations, bone defects were completely repaired in Group A. BMP-2 gene therapy is really a good method to repair segmental bone defects. PMID:17713285

  13. [BMP-2 gene carried by biodegradable scaffold and fibrinous gel for repairing segmental radial defect in rabbit].

    PubMed

    Li, Jianjun; Wang, Enbo; Sun, Hongbin; Han, Dong; Wang, Huan; Bai, Lunhao; Li, Lei; Liu, Xueyong; Xu, Xinxiang

    2007-04-01

    Adenovirus carrying BMP-2 gene, after being mixed with fibrinous gel, was siphoned off on biodegradable scaffolds (PLA/PCL). The composite was used to repair 1.5 cm long radius defect in rabbits. Four methods were in use in the experiments: Ad-BMP-2 plus fibrinous gel and PLA/PCL (Group A), reconstructed hBMP-2 plus fibrinous gel and PLA/PCL (Group B), Ad-Lacz plus fibrinous gel and PLA/PCL (Group C), and fibrinous gel and PLA/PCL (Group D). Results showed that the defects treated in Group A were repaired with much more new bone regenerated, bridged earlier and stronger than those in Group B 12 weeks after operation. The defects treated in the other two groups could not attain osseous tissue healing. BMP-2 gene carried by biodegradable scaffold and fibrinous gel is easy to conduct and has very strong osteoinduction ability. It is really a good method to repair segmental bone defects. PMID:17591257

  14. Yeast DNA-repair gene RAD14 encodes a zinc metalloprotein with affinity for ultraviolet-damaged DNA

    SciTech Connect

    Guzder, S.N.; Sung, P.; Prakash, S. ); Prakash, L. )

    1993-06-15

    Xeroderma pigmentosum (XP) patients suffer from a high incidence of skin cancers due to a defect in excision repair of UV light-damaged DNA. Of the seven XP complementation groups, A--G, group A represents a severe and frequent form of the disease. The Saccharomyces cerevisiae RAD14 gene is a homolog of the XP-A correcting (XPAC) gene. Like XP-A cells, rad14-null mutants are defective in the incision step of excision repair of UV-damaged DNA. The authors have purified RAD14 protein to homogeneity from extract of a yeast strain genetically tailored to overexpress RAD14. As determined by atomic emission spectroscopy, RAD14 contains one zinc atom. They also show in vitro that RAD14 binds zinc but does not bind other divalent metal ions. In DNA mobility-shift assays, RAD14 binds specifically to UV-damaged DNA. Removal of cyclobutane pyrimidine dimers from damaged DNA by enzymatic photoreactivation has no effect on binding, strongly suggesting that RAD14 recognizes pyrimidine(6-4)pyrimidone photoproduct sites. These findings indicate that RAD14 functions in damage recognition during excision repair. 37 refs., 4 figs.

  15. Preferential repair of cyclobutane pyrimidine dimers in the transcribed strand of a gene in yeast chromosomes and plasmids is dependent on transcription.

    PubMed Central

    Sweder, K S; Hanawalt, P C

    1992-01-01

    While preferential repair of the transcribed strands within active genes has been demonstrated in organisms as diverse as humans and Escherichia coli, it has not previously been shown to occur in chromosomal genes in the yeast Saccharomyces cerevisiae. We found that repair of cyclobutane pyrimidine dimers in the transcribed strand of the expressed RPB2 gene in the chromosome of a repair-proficient strain is much more rapid than that in the nontranscribed strand. Furthermore, a copy of the RPB2 gene borne on a centromeric ARS1 plasmid showed the same strand bias in repair. To investigate the relation of this strand bias to transcription, we studied repair in a yeast strain with the temperature-sensitive mutation, rpb1-1, in the largest subunit of RNA polymerase II. When exponentially growing rpb1-1 cells are shifted to the nonpermissive temperature, they rapidly cease mRNA synthesis. At the permissive temperature, both rpb1-1 and the wild-type, parental cells exhibited rapid, proficient repair in the transcribed strand of chromosomal and plasmid-borne copies of the RPB2 gene. At the nonpermissive temperature, the rate of repair in the transcribed strand in rpb1-1 cells was reduced to that in the nontranscribed strand. These findings establish the dependence of strand bias in repair on transcription by RNA polymerase II in the chromosomes and in plasmids, and they validate the use of plasmids for analysis of the relation of repair to transcription in yeast. Images PMID:1438266

  16. Deficiency in nucleotide excision repair family gene activity, especially ERCC3, is associated with non-pigmented hair fiber growth.

    PubMed

    Yu, Mei; Bell, Robert H; Ho, Maggie M; Leung, Gigi; Haegert, Anne; Carr, Nicholas; Shapiro, Jerry; McElwee, Kevin J

    2012-01-01

    We conducted a microarray study to discover gene expression patterns associated with a lack of melanogenesis in non-pigmented hair follicles (HF) by microarray. Pigmented and non-pigmented HFs were collected and micro-dissected into the hair bulb (HB) and the upper hair sheaths (HS) including the bulge region. In comparison to pigmented HS and HBs, nucleotide excision repair (NER) family genes ERCC1, ERCC2, ERCC3, ERCC4, ERCC5, ERCC6, XPA, NTPBP, HCNP, DDB2 and POLH exhibited statistically significantly lower expression in non- pigmented HS and HBs. Quantitative PCR verified microarray data and identified ERCC3 as highly differentially expressed. Immunohistochemistry confirmed ERCC3 expression in HF melanocytes. A reduction in ERCC3 by siRNA interference in human melanocytes in vitro reduced their tyrosinase production ability. Our results suggest that loss of NER gene function is associated with a loss of melanin production capacity. This may be due to reduced gene transcription and/or reduced DNA repair in melanocytes which may eventually lead to cell death. These results provide novel information with regard to melanogenesis and its regulation. PMID:22615732

  17. Redox regulation of genome stability by effects on gene expression, epigenetic pathways and DNA damage/repair

    PubMed Central

    Mikhed, Yuliya; Görlach, Agnes; Knaus, Ulla G.; Daiber, Andreas

    2015-01-01

    Reactive oxygen and nitrogen species (e.g. H2O2, nitric oxide) confer redox regulation of essential cellular signaling pathways such as cell differentiation, proliferation, migration and apoptosis. In addition, classical regulation of gene expression or activity, including gene transcription to RNA followed by translation to the protein level, by transcription factors (e.g. NF-κB, HIF-1α) and mRNA binding proteins (e.g. GAPDH, HuR) is subject to redox regulation. This review will give an update of recent discoveries in this field, and specifically highlight the impact of reactive oxygen and nitrogen species on DNA repair systems that contribute to genomic stability. Emphasis will be placed on the emerging role of redox mechanisms regulating epigenetic pathways (e.g. miRNA, DNA methylation and histone modifications). By providing clinical correlations we discuss how oxidative stress can impact on gene regulation/activity and vise versa, how epigenetic processes, other gene regulatory mechanisms and DNA repair can influence the cellular redox state and contribute or prevent development or progression of disease. PMID:26079210

  18. Analysis of gene expression dynamics revealed delayed and abnormal epidermal repair process in aged compared to young skin.

    PubMed

    Sextius, Peggy; Marionnet, Claire; Tacheau, Charlotte; Bon, François-Xavier; Bastien, Philippe; Mauviel, Alain; Bernard, Bruno A; Bernerd, Françoise; Dubertret, Louis

    2015-05-01

    With aging, epidermal homeostasis and barrier function are disrupted. In a previous study, we analyzed the transcriptomic response of young skin epidermis after stratum corneum removal, and obtained a global kinetic view of the molecular processes involved in barrier function recovery. In the present study, the same analysis was performed in aged skin in order to better understand the defects which occur with aging. Thirty healthy male volunteers (67 ± 4 years old) were involved. Tape-strippings were carried out on the inner face of one forearm, the other unstripped forearm serving as control. At 2, 6, 18, 30 and 72 h after stripping, TEWL measurements were taken, and epidermis samples were collected. Total RNA was extracted and analyzed using DermArray(®) cDNA microarrays. The results highlighted that barrier function recovery and overall kinetics of gene expression were delayed following stripping in aged skin. Indeed, the TEWL measurements showed that barrier recovery in the young group appeared to be dramatically significant during the overall kinetics, while there were no significant evolution in the aged group until 30 h. Moreover, gene expression analysis revealed that the number of modulated genes following tape stripping increased as a function of time and reached a peak at 6 h after tape stripping in young skin, while it was at 30 h in aged skin, showing that cellular activity linked to the repair process may be engaged earlier in young epidermis than in aged epidermis. A total of 370 genes were modulated in the young group. In the aged group, 382 genes were modulated, whose 184 were also modulated in the young group. Only eight genes that were modulated in both groups were significantly differently modulated. The characterization of these genes into 15 functional families helped to draw a scenario for the aging process affecting epidermal repair capacity. PMID:25740152

  19. Damage formation and repair efficiency in the p53 gene of cell lines and blood lymphocytes assayed by multiplex long quantitative polymerase chain reaction.

    PubMed

    Wang, Yu-Chieh; Lee, Pei-Jung; Shih, Chuen-Ming; Chen, Hsing-Yu; Lee, Chin-Chu; Chang, Yuan-Yen; Hsu, Yu-Ting; Liang, Ying-Ju; Wang, Li-Ya; Han, Wen-Hua; Wang, Yi-Ching

    2003-08-15

    We examined ultraviolet (UV) irradiation and cisplatin treatment damage formation and repair efficiency in the p53 tumor suppressor gene of various cultured cell lines and lymphocytes using a nonradioactive multiplex long quantitative polymerase chain reaction (QPCR) assay, which amplified a 7-kb fragment of the target gene and a 500-bp fragment of the template control to successfully increase the sensitivity and reliability of the assay. The multiplex long QPCR detected a lesion frequency of 0.63 lesions/10kb/10J/m(2) in the p53 gene of fibroblast cells. In addition, the multiplex long QPCR assay detected pronounced differences in the repair of UV damage in the p53 gene among repair-proficient CRL-1475 cells and repair-deficient XP-A and XP-C cells. The multiplex long QPCR assay was also evaluated as a sensitive assay for the detection of DNA damage induced by cisplatin. The data indicated that the lesion frequency in the p53 gene was 1.27-1.75 times higher in the H23 cisplatin-sensitive cell than in the H1435 cisplatin-resistant cell at the IC(70) dose. After 8-h and 24-h repair periods, only 13 and 75% of cisplatin-induced damage had been removed in the H23 cells, whereas these values were 92 and 100% in the H1435 cells. In addition, our data indicate that multiplex long QPCR is a sensitive method for validly estimating repair in freshly isolated lymphocytes. The results suggest that the current protocol of the multiplex long QPCR method can be used to assess the damage formation and repair efficiency of various agents at biologically relevant doses and to allow a more precise determination of gene-specific repair in disease susceptibility and drug resistance in epidemiological studies. PMID:12871714

  20. A mutation in the XPB/ERCC3 DNA repair transcription gene, associated with trichothiodystrophy

    SciTech Connect

    Weeda, G.; Donker, I.; Vermeulen, W.

    1997-02-01

    Trichothiodystrophy (TTD) is a rare, autosomal recessive disorder characterized by sulfur-deficient brittle hair and nails, mental retardation, impaired sexual development, and ichthyosis. Photosensitivity has been reported in {approximately}50% of the cases, but no skin cancer is associated with TTD. Virtually all photosensitive TTD patients have a deficiency in the nucleotide excision repair (NER) of UV-induced DNA damage that is indistinguishable from that of xeroderma pigmentosum (XP) complementation group D (XP-D) patients. DNA repair defects in XP-D are associated with two additional, quite different diseases; XP, a sun-sensitive and cancer-prone repair disorder, and Cockayne syndrome (CS), a photosensitive condition characterized by physical and mental retardation and wizened facial appearance. One photosensitive TTD case constitutes a new repair-deficient complementation group, TTD-A. Remarkably, both TTD-A and XP-D defects are associated with subunits of TFIIH, a basal transcription factor with a second function in DNA repair. Thus, mutations in TFIIH components may, on top of a repair defect, also cause transcriptional insufficiency, which may explain part of the non-XP clinical features of TTD. To date, three patients with the remarkable conjunction of XP and CS but not TM have been assigned to XP complementation group B (XP-B). Here we present the characterization of the NER defect in two mild TTD patients (TTD6VI and TTD4VI) and confirm the assignment to X-PB. The causative mutation was found to be a single base substitution resulting in a missense mutation (T119P) in a region of the XPB protein. These findings define a third TTD complementation group, extend the clinical heterogeneity associated with XP-B, stress the exclusive relationship between TTD and mutations in subunits of repair/transcription factor TFIIH, and strongly support the concept of {open_quotes}transcription syndromes.{close_quotes} 46 refs., 6 figs., 2 tabs.

  1. Cloning, comparative mapping, and RNA expression of the mouse homologues of the Saccharomyces cerevisiae nucleotide excision repair gene RAD23.

    PubMed

    van der Spek, P J; Visser, C E; Hanaoka, F; Smit, B; Hagemeijer, A; Bootsma, D; Hoeijmakers, J H

    1996-01-01

    The Saccharomyces cerevisiae RAD23 gene is involved in nucleotide excision repair (NER). Two human homologs of RAD23, HHR23A and HHR23B (HGMW-approved symbols RAD23A and RAD23B), were previously isolated. The HHR23B protein is complexed with the protein defective in the cancer-prone repair syndrome xeroderma pigmentosum, complementation group C, and is specifically involved in the global genome NER subpathway. The cloning of both mouse homologs (designated MHR23A and MHR23B) and detailed sequence comparison permitted the deduction of the following overall structure for all RAD23 homologs: an ubiquitin-like N-terminus followed by a strongly conserved 50-amino-acid domain that is repeated at the C-terminus. We also found this domain as a specific C-terminal extension of one of the ubiquitin-conjugating enzymes, providing a second link with the ubiquitin pathway. By means of in situ hybridization, MHR23A was assigned to mouse chromosome 8C3 and MHR23B to 4B3. Because of the close chromosomal proximity of human XPC and HHR23B, the mouse XPC chromosomal location was determined (6D). Physical disconnection of the genes in mouse argues against a functional significance of the colocalization of these genes in human. Northern blot analysis revealed constitutive expression of both MHR23 genes in all tissues examined. Elevated RNA expression of both MHR23 genes was observed in testis. Although the RAD23 equivalents are well conserved during evolution, the mammalian genes did not express the UV-inducible phenotype of their yeast counterpart. This may point to a fundamental difference between the UV responses of yeast and human. No stage-specific mRNA expression during the cell cycle was observed for the mammalian RAD23 homologs. PMID:8808275

  2. Isolation and Characterization of Two Saccharomyces Cerevisiae Genes Encoding Homologs of the Bacterial Hexa and Muts Mismatch Repair Proteins

    PubMed Central

    Reenan, R. A.; Kolodner, R. D.

    1992-01-01

    Homologs of the Escherichia coli (mutL, S and uvrD) and Streptococcus pneumoniae (hexA, B) genes involved in mismatch repair are known in several distantly related organisms. Degenerate oligonucleotide primers based on conserved regions of E. coli MutS protein and its homologs from Salmonella typhimurium, S. pneumoniae and human were used in the polymerase chain reaction (PCR) to amplify and clone mutS/hexA homologs from Saccharomyces cerevisiae. Two DNA sequences were amplified whose deduced amino acid sequences both shared a high degree of homology with MutS. These sequences were then used to clone the full-length genes from a yeast genomic library. Sequence analysis of the two MSH genes (MSH = mutS homolog), MSH1 and MSH2, revealed open reading frames of 2877 bp and 2898 bp. The deduced amino acid sequences predict polypeptides of 109.3 kD and 109.1 kD, respectively. The overall amino acid sequence identity with the E. coli MutS protein is 28.6% for MSH1 and 25.2% for MSH2. Features previously found to be shared by MutS homologs, such as the nucleotide binding site and the helix-turn-helix DNA binding motif as well as other highly conserved regions whose function remain unknown, were also found in the two yeast homologs. Evidence presented in this and a companion study suggest that MSH1 is involved in repair of mitochondrial DNA and that MSH2 is involved in nuclear DNA repair. PMID:1459447

  3. Radiation-Induced Upregulation of Gene Expression From Adenoviral Vectors Mediated by DNA Damage Repair and Regulation

    SciTech Connect

    Nokisalmi, Petri; Rajecki, Maria; Pesonen, Sari; Escutenaire, Sophie; Soliymani, Rabah; Tenhunen, Mikko; Ahtiainen, Laura; Hemminki, Akseli

    2012-05-01

    Purpose: In the present study, we evaluated the combination of replication-deficient adenoviruses and radiotherapy in vitro. The purpose of the present study was to analyze the mechanism of radiation-mediated upregulation of adenoviral transgene expression. Methods and Materials: Adenoviral transgene expression (luciferase or green fluorescent protein) was studied with and without radiation in three cell lines: breast cancer M4A4-LM3, prostate cancer PC-3MM2, and lung cancer LNM35/enhanced green fluorescent protein. The effect of the radiation dose, modification of the viral capsid, and five different transgene promoters were studied. The cellular responses were studied using mass spectrometry and immunofluorescence analysis. Double strand break repair was modulated by inhibitors of heat shock protein 90, topoisomerase-I, and DNA protein kinase, and transgene expression was measured. Results: We found that a wide range of radiation doses increased adenoviral transgene expression regardless of the cell line, transgene, promoter, or viral capsid modification. Treatment with adenovirus, radiation, and double strand break repair inhibitors resulted in persistence of double strand breaks and subsequent increases in adenovirus transgene expression. Conclusions: Radiation-induced enhancement of adenoviral transgene expression is linked to DNA damage recognition and repair. Radiation induces a global cellular response that results in increased production of RNA and proteins, including adenoviral transgene products. This study provides a mechanistic rationale for combining radiation with adenoviral gene delivery.

  4. Cloning of the E. coli O6-methylguanine and methylphosphotriester methyltransferase gene using a functional DNA repair assay.

    PubMed Central

    Margison, G P; Cooper, D P; Brennand, J

    1985-01-01

    Alkylating agents react with various nitrogen and oxygen atoms in DNA and many of the products are substrates for repair processes. Oxygen atom derivatives such as O6-methylguanine (O6-meG) O4-methylthymine and methylphosphotriesters (MP) have been shown to undergo repair by methyl group removal. The proteins involved in the latter reaction can be considered to be methyltransferases (MT) because their action results in the transfer of the methyl group to a cysteine residue within a polypeptide. A rapid and sensitive assay for MT activity has been developed and used to screen extracts of bacteria harbouring an E. coli genomic DNA library carried in a plasmid vector. We report here the cloning of an E. coli gene coding for O6-meG and MP MT repair functions. These two activities reside on a 37Kd protein that can undergo a host-dependent cleavage to produce an 18Kd protein which contains only O6-meG MT and a 13Kd protein which contains only MP MT. Images PMID:3889845

  5. Impaired Cytogenetic Damage Repair and Cell Cycle Regulation in Response to Ionizing Radiation in Human Fibroblast Cells with Individual Knock-down of 25 Genes

    NASA Technical Reports Server (NTRS)

    Zhang, Ye; Rohde, Larry; Emami, Kamal; Hammond, Dianne; Casey, Rachael; Mehta, Satish; Jeevarajan, Antony; Pierson, Duane; Wu, Honglu

    2008-01-01

    Changes of gene expression profile are one of the most important biological responses in living cells after ionizing radiation (IR) exposure. Although some studies have demonstrated that genes with upregulated expression induced by IR may play important roles in DNA damage sensing, cell cycle checkpoint and chromosomal repair, the relationship between the regulation of gene expression by IR and its impact on cytogenetic responses to ionizing radiation has not been systematically studied. In our present study, the expression of 25 genes selected based on their transcriptional changes in response to IR or from their known DNA repair roles were individually knocked down by siRNA transfection in human fibroblast cells. Chromosome aberrations (CA) and micronuclei (MN) formation were measured as the cytogenetic endpoints. Our results showed that the yield of MN and/or CA formation were significantly increased by suppressed expression of 5 genes that included Ku70 in the DSB repair pathway; XPA in the NER pathway; RPA1 in the MMR pathway; RAD17 and RBBP8 in cell cycle control. Knocked-down expression of 4 genes including MRE11A, RAD51 in the DSB pathway, and SESN1 and SUMO1 showed significant inhibition of cell cycle progression, possibly because of severe impairment of DNA damage repair. Furthermore, loss of XPA, p21 and MLH1 expression resulted in both enhanced cell cycle progression and significantly higher yield of cytogenetic damage, indicating the involvement of these gene products in both cell cycle control and DNA damage repair. Of these 11 genes that affected the cytogenetic response, 9 were up-regulated in the cells exposed to gamma radiation, suggesting that genes transcriptionally modulated by IR were critical to regulating the biological consequences after IR. Failure to express these IR-responsive genes, such as by gene mutation, could seriously change the outcome of the post IR scenario and lead to carcinogenesis.

  6. Molecular cloning of the human XRCC1 gene, which corrects defective DNA strand break repair and sister chromatid exchange.

    PubMed Central

    Thompson, L H; Brookman, K W; Jones, N J; Allen, S A; Carrano, A V

    1990-01-01

    We describe the cloning and function of the human XRCC1 gene, which is the first mammalian gene isolated that affects cellular sensitivity to ionizing radiation. The CHO mutant EM9 has 10-fold-higher sensitivity to ethyl methanesulfonate, 1.8-fold-higher sensitivity to ionizing radiation, a reduced capacity to rejoin single-strand DNA breaks, and a 10-fold-elevated level of sister chromatid exchange compared with the CHO parental cells. The complementing human gene was cloned from a cosmid library of a tertiary transformant. Two cosmid clones produced transformants that showed approximately 100% correction of the repair defect in EM9 cells, as determined by the kinetics of strand break repair, cell survival, and the level of sister chromatid exchange. A nearly full-length clone obtained from the pcD2 human cDNA expression library gave approximately 80% correction of EM9, as determined by the level of sister chromatid exchange. Based on an analysis of the nucleotide sequence of the cDNA insert compared with that of the 5' end of the gene from a cosmid clone, the cDNA clone appeared to be missing approximately 100 bp of transcribed sequence, including 26 nucleotides of coding sequence. The cDNA probe detected a single transcript of approximately 2.2 kb in HeLa polyadenylated RNA by Northern (RNA) blot hybridization. From the open reading frame and the positions of likely start sites for transcription and translation, the size of the putative XRCC1 protein is 633 amino acids (69.5 kDa). The size of the XRCC1 gene is 33 kb, as determined by localizing the endpoints on a restriction endonuclease site map of one cosmid clone. The deduced amino acid sequence did not show significant homology with any protein in the protein sequence data bases examined. Images PMID:2247054

  7. Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum.

    PubMed Central

    Tanaka, K; Satokata, I; Ogita, Z; Uchida, T; Okada, Y

    1989-01-01

    For isolation of the gene responsible for xeroderma pigmentosum (XP) complementation group A, plasmid pSV2gpt and genomic DNA from a mouse embryo were cotransfected into XP2OSSV cells, a group-A XP cell line. Two primary UV-resistant XP transfectants were isolated from about 1.6 X 10(5) pSV2gpt-transformed XP colonies. pSV2gpt and genomic DNA from the primary transfectants were again cotransfected into XP2OSSV cells and a secondary UV-resistant XP transfectant was obtained by screening about 4.8 X 10(5) pSV2gpt-transformed XP colonies. The secondary transfectant retained fewer mouse repetitive sequences. A mouse gene that complements the defect of XP2OSSV cells was cloned into an EMBL3 vector from the genome of a secondary transfectant. Transfections of the cloned DNA also conferred UV resistance on another group-A XP cell line but not on XP cell lines of group C, D, F, or G. Northern blot analysis of poly(A)+ RNA with a subfragment of cloned mouse DNA repair gene as the probe revealed that an approximately 1.0 kilobase mRNA was transcribed in the donor mouse embryo and secondary transfectant, and approximately 1.0- and approximately 1.3-kilobase mRNAs were transcribed in normal human cells, but none of these mRNAs was detected in three strains of group-A XP cells. These results suggest that the cloned DNA repair gene is specific for group-A XP and may be the mouse homologue of the group-A XP human gene. Images PMID:2748601

  8. Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum

    SciTech Connect

    Tanaka, K.; Satokata, I.; Ogita, Z.; Uchida, T.; Okada, Y.

    1989-07-01

    For isolation of the gene responsible for xeroderma pigmentosum (XP) complementation group A, plasmid pSV2gpt and genomic DNA from a mouse embryo were cotransfected into XP2OSSV cells, a group-A XP cell line. Two primary UV-resistant XP transfectants were isolated from about 1.6 X 10(5) pSV2gpt-transformed XP colonies. pSV2gpt and genomic DNA from the primary transfectants were again cotransfected into XP2OSSV cells and a secondary UV-resistant XP transfectant was obtained by screening about 4.8 X 10(5) pSV2gpt-transformed XP colonies. The secondary transfectant retained fewer mouse repetitive sequences. A mouse gene that complements the defect of XP2OSSV cells was cloned into an EMBL3 vector from the genome of a secondary transfectant. Transfections of the cloned DNA also conferred UV resistance on another group-A XP cell line but not on XP cell lines of group C, D, F, or G. Northern blot analysis of poly(A)+ RNA with a subfragment of cloned mouse DNA repair gene as the probe revealed that an approximately 1.0 kilobase mRNA was transcribed in the donor mouse embryo and secondary transfectant, and approximately 1.0- and approximately 1.3-kilobase mRNAs were transcribed in normal human cells, but none of these mRNAs was detected in three strains of group-A XP cells. These results suggest that the cloned DNA repair gene is specific for group-A XP and may be the mouse homologue of the group-A XP human gene.

  9. POLYMORPHISMS IN THE DNA NUCLEOTIDE EXCISION REPAIR GENES AND LUNG CANCER RISK IN XUAN WEI, CHINA

    EPA Science Inventory

    The lung cancer mortality rate in Xuan Wei County, China is among the highest in the country and has been etiologically attributed to exposure to indoor smoky coal emissions that contain very high levels of polycyclic aromatic hydrocarbons (PAHs). Nucleotide excision repair (NE...

  10. Antioxidative Dietary Compounds Modulate Gene Expression Associated with Apoptosis, DNA Repair, Inhibition of Cell Proliferation and Migration

    PubMed Central

    Wang, Likui; Gao, Shijuan; Jiang, Wei; Luo, Cheng; Xu, Maonian; Bohlin, Lars; Rosendahl, Markus; Huang, Wenlin

    2014-01-01

    Many dietary compounds are known to have health benefits owing to their antioxidative and anti-inflammatory properties. To determine the molecular mechanism of these food-derived compounds, we analyzed their effect on various genes related to cell apoptosis, DNA damage and repair, oxidation and inflammation using in vitro cell culture assays. This review further tests the hypothesis proposed previously that downstream products of COX-2 (cyclooxygenase-2) called electrophilic oxo-derivatives induce antioxidant responsive elements (ARE), which leads to cell proliferation under antioxidative conditions. Our findings support this hypothesis and show that cell proliferation was inhibited when COX-2 was down-regulated by polyphenols and polysaccharides. Flattened macrophage morphology was also observed following the induction of cytokine production by polysaccharides extracted from viili, a traditional Nordic fermented dairy product. Coix lacryma-jobi (coix) polysaccharides were found to reduce mitochondrial membrane potential and induce caspase-3- and 9-mediated apoptosis. In contrast, polyphenols from blueberries were involved in the ultraviolet-activated p53/Gadd45/MDM2 DNA repair system by restoring the cell membrane potential. Inhibition of hypoxia-inducible factor-1 by saponin extracts of ginsenoside (Ginsen) and Gynostemma and inhibition of S100A4 by coix polysaccharides inhibited cancer cell migration and invasion. These observations suggest that antioxidants and changes in cell membrane potential are the major driving forces that transfer signals through the cell membrane into the cytosol and nucleus, triggering gene expression, changes in cell proliferation and the induction of apoptosis or DNA repair. PMID:25226533

  11. Decreased DNA repair gene XRCC1 expression is associated with radiotherapy-induced acute side effects in breast cancer patients.

    PubMed

    Batar, Bahadir; Guven, Gulgun; Eroz, Seda; Bese, Nuran Senel; Guven, Mehmet

    2016-05-10

    DNA repair plays a critical role in response to ionizing radiation (IR) and developing of radiotherapy induced normal tissue reactions. In our study, we investigated the association of radiotherapy related acute side effects, with X-ray repair cross complementing group 1 (XRCC1) and Poly (ADP-ribose) polymerase 1 (PARP1) DNA repair gene expression levels, their changes in protein expression and DNA damage levels in breast cancer patients. The study included 40 women with newly diagnosed breast cancer; an experimental case group (n=20) with acute side effects and the control group (n=20) without side effects. For gene and protein expression analysis, lymphocytes were cultured for 72 h and followed by in vitro 2 Gray (Gy) gamma-irradiation. For detection of DNA damage levels, lymphocytes were irradiated with in vitro 2 Gy gamma-rays and followed by incubation for 72 h. XRCC1 mRNA and protein expression levels were significantly higher in controls than in experimental cases (P=0.020). In terms of DNA damage levels, an increased frequency of micronucleus (MN) was observed in experimental cases versus controls, but this association was not significant (P=0.206). We also observed a significant negative correlation between MN frequency and XRCC1 protein levels in experimental (r=-0.469, P=0.037) vs control (r=-0.734, P<0.001). Our results suggested that decreased XRCC1 expression levels might be associated with the increased risk of therapeutic IR-related acute side effects in patients with breast cancer. PMID:26826460

  12. Predisposing factors and prevention of frostbite.

    PubMed

    Rintamäki, H

    2000-04-01

    This review focuses on the physiological, behavioural and environmental factors which predispose to frostbite. Also prevention of frostbite is summarised. Predisposing factors may increase heat loss, decrease heat production, decrease the insulation of the clothing, make people especially susceptible to cold or make them to behave inadequately. Marked increase in convective or conductive heat loss is often the immediate reason for frostbite. Wind (as described by wind chill index) increases convective heat loss and touching of metal objects increases conductive cooling. Poor insulation of the clothing is also a common reason of frostbite. The insulation can be insufficient when clothing is wet, tight, permeable to wind or does not cover the cold sensitive body parts. Individual factors predisposing to frostbite are inadequate behaviour, low physical fitness, fatigue, dehydration, earlier cold injuries, sickness or poor circulation in peripheral parts of the body. Frostbite is often associated with the use of alcohol. To prevent frostbite, it is necessary to recognise cold risks, practise tasks in the cold, eat and drink well, have physical exercise, have sufficient clothing (also spare clothing), change into dry clothing if necessary and take care of companions. In the cold it is not advisable to get fatigued until exhaustion, sweat excessively, use tight and/or wet clothing, drink alcohol, smoke and expose oneself unnecessarily to wind, metals or fluids. PMID:10998828

  13. [Chromosomal aberrations and genetic polymorphism in genes of the xenobiotic detoxification and DNA repair enzymes in thermoelectric power plant employers].

    PubMed

    Savchenko, Ia A; Minina, V I; Bakanova, M L

    2012-01-01

    The results of the investigation of the interrelationship between frequency of chromosomal aberrations and detoxification enzymes (GSTM1, GSTT1) and DNA repair (hOGG1, XPD) genes in the employees of fuel energy complex in Kemerovo are presented In the group of the workers frequency of metaphases with aberrations (3,9 +/- 0,2%: n = 288) was shown to be significantly higher than in the comparison group (2,1 0, 2%: n = +/- 141). In the group of workers and control donors statistically significant differences were revealed in the frequency of distribution of the GSTT1 and hOGG1 genes. The level of chromosomal aberrations was established to be higher in patients with GSTM1 genotype "0/0" in the group of control donors. PMID:23458003

  14. TGFβ Induces ‘BRCAness’ and Sensitivity to PARP Inhibition in Breast Cancer by Regulating DNA Repair Genes

    PubMed Central

    Liu, Liang; Zhou, Weiying; Cheng, Chun-Ting; Ren, Xiubao; Somlo, George; Fong, Miranda Y.; Chin, Andrew R.; Li, Hui; Yu, Yang; Xu, Yang; O'Connor, Sean Timothy Francis; O'Connor, Timothy R.; Ann, David K.; Stark, Jeremy M.; Wang, Shizhen Emily

    2014-01-01

    Transforming growth factor β (TGFβ) proteins are multitasking cytokines, whose high levels at tumor sites generally correlate with poor prognosis in human cancer patients. Previously it was reported that TGFβ downregulates the expression of ataxia telangiectasia mutated (ATM) and mutS homolog 2 (MSH2) in breast cancer (BC) cells through a miRNA-mediated mechanism. In this study, expression of a panel of DNA repair genes was examined, identifying breast cancer 1, early onset (BRCA1) as a target downregulated by TGFβ through the miR-181 family. Correlations between the expression levels of TGFβ1 and the miR-181/BRCA1 axis were observed in primary breast tumor specimens. By downregulating BRCA1, ATM, and MSH2, TGFβ orchestrates DNA damage response (DDR) in certain BC cells to induce a ‘BRCAness’ phenotype, including impaired DNA repair efficiency and synthetic lethality to the inhibition of poly (ADP-ribose) polymerase (PARP). Xenograft tumors with active TGFβ signaling exhibited resistance to the DNA-damaging agent doxorubicin but increased sensitivity to the PARP inhibitor ABT-888. Combination of doxorubicin with ABT-888 significantly improved the treatment efficacy in TGFβ-active tumors. Thus, TGFβ can induce ‘BRCAness’ in certain BCs carrying wild-type BRCA genes and enhance the responsiveness to PARP inhibition, and the molecular mechanism behind this is characterized. Implications: These findings enable better selection of sporadic breast cancer patients for PARP interventions, which have exhibited beneficial effects in patients carrying BRCA mutations. PMID:25103497

  15. Three-dimensionally specific inhibition of DNA repair-related genes by activated KRAS in colon crypt model.

    PubMed

    Tsunoda, Toshiyuki; Takashima, Yasuo; Fujimoto, Takahiro; Koyanagi, Midori; Yoshida, Yasuhiro; Doi, Keiko; Tanaka, Yoko; Kuroki, Masahide; Sasazuki, Takehiko; Shirasawa, Senji

    2010-05-01

    Growth and differentiation of colonic epithelium are regulated in the three-dimensional (3D) physiological architecture, colonic crypt, and deregulation of 3D interactions is involved in tumorigenesis. Cell-based 3D culture systems provide a suitable approach bridging the gap between two-dimensional (2D) culture and animal models. KRAS mutations are found at high frequencies in human colorectal cancer (CRC); however, KRAS-targeted cancer therapy has not been developed. Here, we have established a 3D cell culture model resembling the colonic crypt by use of HKe3 cells, human CRC HCT116 cells disrupted at activated KRAS. In this 3D colonic crypt model, HKe3 cells showed the features of time course-dependent transit-amplifying and terminal-differentiated stages, which are characteristic of normal colonic crypt. On the basis of the features of HCT116 cells, activated KRAS inhibited normal cell polarity and apoptosis in 3D culture. The expression of DNA repair-related tumor suppressor genes including TP53, BRCA1, BRCA2, and EXO-1 was markedly suppressed by activated KRAS in 3D culture but not in 2D culture. These results together suggest that activated KRAS plays critical roles in the accumulation of genetic alterations through inhibition of DNA repair genes and apoptosis and that this 3D culture model will provide a useful tool for investigating the molecular mechanisms of CRC development. PMID:20454511

  16. Three-dimensionally Specific Inhibition of DNA Repair-Related Genes by Activated KRAS in Colon Crypt Model1 2

    PubMed Central

    Tsunoda, Toshiyuki; Takashima, Yasuo; Fujimoto, Takahiro; Koyanagi, Midori; Yoshida, Yasuhiro; Doi, Keiko; Tanaka, Yoko; Kuroki, Masahide; Sasazuki, Takehiko; Shirasawa, Senji

    2010-01-01

    Growth and differentiation of colonic epithelium are regulated in the three-dimensional (3D) physiological architecture, colonic crypt, and deregulation of 3D interactions is involved in tumorigenesis. Cell-based 3D culture systems provide a suitable approach bridging the gap between two-dimensional (2D) culture and animal models. KRAS mutations are found at high frequencies in human colorectal cancer (CRC); however, KRAS-targeted cancer therapy has not been developed. Here, we have established a 3D cell culture model resembling the colonic crypt by use of HKe3 cells, human CRC HCT116 cells disrupted at activated KRAS. In this 3D colonic crypt model, HKe3 cells showed the features of time course-dependent transit-amplifying and terminal-differentiated stages, which are characteristic of normal colonic crypt. On the basis of the features of HCT116 cells, activated KRAS inhibited normal cell polarity and apoptosis in 3D culture. The expression of DNA repair-related tumor suppressor genes including TP53, BRCA1, BRCA2, and EXO-1 was markedly suppressed by activated KRAS in 3D culture but not in 2D culture. These results together suggest that activated KRAS plays critical roles in the accumulation of genetic alterations through inhibition of DNA repair genes and apoptosis and that this 3D culture model will provide a useful tool for investigating the molecular mechanisms of CRC development. PMID:20454511

  17. Trichostatin A enhances vascular repair by injected human endothelial progenitors through increasing the expression of TAL1-dependent genes.

    PubMed

    Palii, Carmen G; Vulesevic, Branka; Fraineau, Sylvain; Pranckeviciene, Erinija; Griffith, Alexander J; Chu, Alphonse; Faralli, Hervé; Li, Yuhua; McNeill, Brian; Sun, Jie; Perkins, Theodore J; Dilworth, F Jeffrey; Perez-Iratxeta, Carol; Suuronen, Erik J; Allan, David S; Brand, Marjorie

    2014-05-01

    A major goal of cell therapy for vascular diseases is to promote revascularization through the injection of endothelial stem/progenitor cells. The gene regulatory mechanisms that underlie endothelial progenitor-mediated vascular repair, however, remain elusive. Here, we identify the transcription factor TAL1/SCL as a key mediator of the vascular repair function of primary human endothelial colony-forming cells (ECFCs). Genome-wide analyses in ECFCs demonstrate that TAL1 activates a transcriptional program that promotes cell adhesion and migration. At the mechanistic level, we show that TAL1 upregulates the expression of migratory and adhesion genes through recruitment of the histone acetyltransferase p300. Based on these findings, we establish a strategy that enhances the revascularization efficiency of ECFCs after ischemia through ex vivo priming with the histone deacetylase inhibitor TSA. Thus, small molecule epigenetics drugs are effective tools for modifying the epigenome of stem/progenitor cells prior to transplantation as a means to enhance their therapeutic potential. PMID:24792117

  18. SNPs in DNA repair or oxidative stress genes and late subcutaneous fibrosis in patients following single shot partial breast irradiation

    PubMed Central

    2012-01-01

    Background The aim of this study was to evaluate the potential association between single nucleotide polymorphisms related response to radiotherapy injury, such as genes related to DNA repair or enzymes involved in anti-oxidative activities. The paper aims to identify marker genes able to predict an increased risk of late toxicity studying our group of patients who underwent a Single Shot 3D-CRT PBI (SSPBI) after BCS (breast conserving surgery). Methods A total of 57 breast cancer patients who underwent SSPBI were genotyped for SNPs (single nucleotide polymorphisms) in XRCC1, XRCC3, GST and RAD51 by Pyrosequencing technology. Univariate analysis (ORs and 95% CI) was performed to correlate SNPs with the risk of developing ≥ G2 fibrosis or fat necrosis. Results A higher significant risk of developing ≥ G2 fibrosis or fat necrosis in patients with: polymorphic variant GSTP1 (Ile105Val) (OR = 2.9; 95%CI, 0.88-10.14, p = 0.047). Conclusions The presence of some SNPs involved in DNA repair or response to oxidative stress seem to be able to predict late toxicity. Trial Registration ClinicalTrials.gov: NCT01316328 PMID:22272830

  19. Ovarian steroids regulate gene expression related to DNA repair and neurodegenerative diseases in serotonin neurons of macaques.

    PubMed

    Bethea, C L; Reddy, A P

    2015-12-01

    Depression often accompanies the perimenopausal transition and it often precedes overt symptomology in common neurodegenerative diseases (NDDs, such as Alzheimer's, Parkinson's, Huntington, amyotrophic lateral sclerosis). Serotonin dysfunction is frequently found in the different etiologies of depression. We have shown that ovariectomized (Ovx) monkeys treated with estradiol (E) for 28 days supplemented with placebo or progesterone (P) on days 14-28 had reduced DNA fragmentation in serotonin neurons of the dorsal raphe nucleus, and long-term Ovx monkeys had fewer serotonin neurons than intact controls. We questioned the effect of E alone or E+P (estradiol supplemented with progesterone) on gene expression related to DNA repair, protein folding (chaperones), the ubiquitin-proteosome, axon transport and NDD-specific genes in serotonin neurons. Ovx macaques were treated with placebo, E or E+P (n=3 per group) for 1 month. Serotonin neurons were laser captured and subjected to microarray analysis and quantitative real-time PCR (qRT-PCR). Increases were confirmed with qRT-PCR in five genes that code for proteins involved in repair of strand breaks and nucleotide excision. NBN1, PCNA (proliferating nuclear antigen), GADD45A (DNA damage-inducible), RAD23A (DNA damage recognition) and GTF2H5 (gene transcription factor 2H5) significantly increased with E or E+P treatment (all analysis of variance (ANOVA), P<0.01). Chaperone genes HSP70 (heat-shock protein 70), HSP60 and HSP27 significantly increased with E or E+P treatment (all ANOVA, P<0.05). HSP90 showed a similar trend. Ubiquinase coding genes UBEA5, UBE2D3 and UBE3A (Parkin) increased with E or E+P (all ANOVA, P<0.003). Transport-related genes coding kinesin, dynein and dynactin increased with E or E+P treatment (all ANOVA, P<0.03). SCNA (α-synuclein) and ADAM10 (α-secretase) increased (both ANOVA, P<0.02) but PSEN1 (presenilin1) decreased (ANOVA, P<0.02) with treatment. APP decreased 10-fold with E or E

  20. Mismatch Repair in Schizosaccharomyces Pombe Requires the Mutl Homologous Gene Pms1: Molecular Cloning and Functional Analysis

    PubMed Central

    Schar, P.; Baur, M.; Schneider, C.; Kohli, J.

    1997-01-01

    Homologues of the bacterial mutS and mutL genes involved in DNA mismatch repair have been found in organisms from bacteria to humans. Here, we describe the structure and function of a newly identified Schizosaccharomyces pombe gene that encodes a predicted amino acid sequence of 794 residues with a high degree of homology to MutL related proteins. On the basis of its closer relationship to the eukaryotic ``PMS'' genes than to the ``MLH'' genes, we have designated the S. pombe homologue pms1. Disruption of the pms1 gene causes a significant increase of spontaneous mutagenesis as documented by reversion rate measurements. Tetrad analyses of crosses homozygous for the pms1 mutation reveal a reduction of spore viability from >92% to 80% associated with a low proportion (~50%) of meioses producing four viable spores and a significant, allele-dependent increase of the level of post-meiotic segregation of genetic marker allele pairs. The mutant phenotypes are consistent with a general function of pms1 in correction of mismatched base pairs arising as a consequence of DNA polymerase errors during DNA synthesis, or of hybrid DNA formation between homologous but not perfectly complementary DNA strands during meiotic recombination. PMID:9258673

  1. Heterogeneity of excision repair cross-complementation group 1 gene expression in non-small-cell lung cancer patients

    PubMed Central

    SMIRNOV, SERHEY; PASHKEVICH, ANASTASIYA; LIUNDYSHEVA, VALERIYA; BABENKO, ANDREY; SMOLYAKOVA, RAISA

    2015-01-01

    Excision repair cross-complementation group 1 (ERCC1) gene expression analysis is currently used widely in the molecular diagnosis of cancer. According to numerous studies, ERCC1 gene expression correlates with overall survival and effectiveness of chemotherapy with platinum agents. However, the degree of this correlation differs among various studies, with certain authors reporting a complete lack of such a correlation. These contradictions may be attributed to a number of factors, including the heterogeneity of the tumor tissue. In this study, we attempted to assess the degree of genetic heterogeneity exhibited by tissue samples obtained from non-small-cell lung cancer (NSCLC) through the expression of the ERCC1 gene. This study included 25 samples of tumor tissue from patients with a morphologically confirmed NSCLC diagnosis. A total of three randomized sections of each specimen were used. The ERCC1 gene expression was assessed by quantitative polymerase chain reaction (qPCR) in the TaqMan format. When planning the experiment and analysis of qPCR data, the MIQE guidelines were taken into consideration. We established that the coefficient of variation of the relative level of ERCC1 gene expression in the majority of the samples exceeded 33% (P<0.05), indicating the significant heterogeneity of the sample. We also demonstrated that the degree of heterogeneity of the tumor tissue is largely dependent on disease stage. PMID:25469300

  2. A Novel Cell Death Gene Acts to Repair Patterning Defects in Drosophila melanogaster

    PubMed Central

    Tanaka, Kentaro M.; Takahashi, Aya; Fuse, Naoyuki; Takano-Shimizu-Kouno, Toshiyuki

    2014-01-01

    Cell death is a mechanism utilized by organisms to eliminate excess cells during development. Here, we describe a novel regulator of caspase-independent cell death, Mabiki (Mabi), that is involved in the repair of the head patterning defects caused by extra copies of bicoid in Drosophila melanogaster. Mabiki functions together with caspase-dependent cell death mechanisms to provide robustness during development. PMID:24671768

  3. A novel cell death gene acts to repair patterning defects in Drosophila melanogaster.

    PubMed

    Tanaka, Kentaro M; Takahashi, Aya; Fuse, Naoyuki; Takano-Shimizu-Kouno, Toshiyuki

    2014-06-01

    Cell death is a mechanism utilized by organisms to eliminate excess cells during development. Here, we describe a novel regulator of caspase-independent cell death, Mabiki (Mabi), that is involved in the repair of the head patterning defects caused by extra copies of bicoid in Drosophila melanogaster. Mabiki functions together with caspase-dependent cell death mechanisms to provide robustness during development. PMID:24671768

  4. Beryllium chloride-induced oxidative DNA damage and alteration in the expression patterns of DNA repair-related genes.

    PubMed

    Attia, Sabry M; Harisa, Gamaleldin I; Hassan, Memy H; Bakheet, Saleh A

    2013-09-01

    Beryllium metal has physical properties that make its use essential for very specific applications, such as medical diagnostics, nuclear/fusion reactors and aerospace applications. Because of the widespread human exposure to beryllium metals and the discrepancy of the genotoxic results in the reported literature, detail assessments of the genetic damage of beryllium are warranted. Mice exposed to beryllium chloride at an oral dose of 23mg/kg for seven consecutive days exhibited a significant increase in the level of DNA-strand breaking and micronuclei formation as detected by a bone marrow standard comet assay and micronucleus test. Whereas slight beryllium chloride-induced oxidative DNA damage was detected following formamidopyrimidine DNA glycosylase digestion, digestion with endonuclease III resulted in considerable increases in oxidative DNA damage after the 11.5 and 23mg/kg/day treatment as detected by enzyme-modified comet assays. Increased 8-hydroxydeoxyguanosine was also directly correlated with increased bone marrow micronuclei formation and DNA strand breaks, which further confirm the involvement of oxidative stress in the induction of bone marrow genetic damage after exposure to beryllium chloride. Gene expression analysis on the bone marrow cells from beryllium chloride-exposed mice showed significant alterations in genes associated with DNA damage repair. Therefore, beryllium chloride may cause genetic damage to bone marrow cells due to the oxidative stress and the induced unrepaired DNA damage is probably due to the down-regulation in the expression of DNA repair genes, which may lead to genotoxicity and eventually cause carcinogenicity. PMID:23793613

  5. XRCC1 and XPD DNA repair gene polymorphisms: A potential risk factor for glaucoma in the Pakistani population

    PubMed Central

    Yousaf, Sajeela; Khan, Muhammad Imran; Micheal, Shazia; Akhtar, Farah; Ali, Syeda Hafiza Benish; Riaz, Moeen; Ali, Mahmood; Lall, Pramila; Waheed, Nadia Khalida; den Hollander, Anneke I.; Ahmed, Asifa

    2011-01-01

    Purpose The present study was designed to determine the association of polymorphisms of the DNA repair genes X-ray cross-complementing group 1 (XRCC1) (c.1316G>A [rs25487]) and xeroderma pigmentosum complementation group D (XPD) (c.2298A>C [rs13181]) with primary open-angle glaucoma (POAG) and primary closed-angle glaucoma (PCAG). Methods In this prospective case-control study, polymerase chain reaction-restriction fragment length polymorphism analysis was used to study the association of XRCC1 and XPD with 160 POAG patients, 163 PCAG patients, and 193 unaffected controls. Results XRCC1 rs25487 was found to be significantly associated specifically with male POAG patients (χ2=13.2 [p=0.001]), only for the dominant model (odds ratio [OR]=2.65 [95% confidence interval [CI]=1.44–4.85], p<0.005). In addition XPD rs13181 was also found to be associated with male POAG patients (χ2=12.1 [p<0.005]), for both dominant (OR=2.44 [95% CI=1.33–4.47], p<0.005) as well as recessive model (OR=3.62 [95% CI=1.45–9.01], p<0.01). Combined genotypes of both the genes revealed that the heterozygote AC/GA was significantly associated with the male POAG patients (z=3.00 [p<0.001]). The AA/GG genotype was present at a higher frequency in the male controls and the AA/GA in the female controls and could thus have a protective role in males and females, respectively. Conclusions We postulate that defects in the DNA repair genes XRCC1 and XPD may possibly be associated with the progression of POAG in male patients of Pakistani origin. PMID:21617750

  6. Dependence of Colorectal Cancer Risk on the Parent-of-Origin of Mutations in DNA Mismatch Repair Genes

    PubMed Central

    van Vliet, Christine M.; Dowty, James G.; van Vliet, Jane L.; Smith, Letitia; Mead, Leeanne J.; Macrae, Finlay A.; St. John, D. James B.; Giles, Graham G.; Southey, Melissa C.; Jenkins, Mark A.; Velan, Gary M.; Hopper, John L.

    2011-01-01

    Genetic diseases associated with dynamic mutations in microsatellite DNA often display parent-of-origin effects (POEs) in which the risk of disease depends on the sex of the parent from whom the disease allele was inherited. Carriers of germline mutations in mismatch repair (MMR) genes have high risks of colorectal carcinoma (CRC). We investigated whether these risks depend on the parent-of-origin of the mutation. We studied 422 subjects, including 89 MMR gene mutation carriers, from 17 population-based families who were each recruited via a CRC case diagnosed before age 45 years and found to carry a MMR gene mutation. The POE hazard ratio (HRPOE), defined to be the CRC incidence for carriers with maternally derived mutations divided by the corresponding paternal incidence, was estimated using a novel application of modified segregation analysis. HRPOE (95% confidence interval) was estimated to be 3.2 (1.1–9.8) for males (P=0.03) and 0.8 (0.2–2.8) for females (P=0.5) and the corresponding cumulative risks to age 80 years were 88% (54%–100%) for male carriers with maternally derived mutations and 38–48% for all other carriers. If confirmed by larger studies, these results will have important implications for the etiology of CRC and for the clinical management of MMR gene mutation carriers. PMID:21120946

  7. Dependence of colorectal cancer risk on the parent-of-origin of mutations in DNA mismatch repair genes.

    PubMed

    van Vliet, Christine M; Dowty, James G; van Vliet, Jane L; Smith, Letitia; Mead, Leeanne J; Macrae, Finlay A; St John, D James B; Giles, Graham G; Southey, Melissa C; Jenkins, Mark A; Velan, Gary M; Hopper, John L

    2011-02-01

    Genetic diseases associated with dynamic mutations in microsatellite DNA often display parent-of-origin effects (POEs) in which the risk of disease depends on the sex of the parent from whom the disease allele was inherited. Carriers of germline mutations in mismatch repair (MMR) genes have high risks of colorectal carcinoma (CRC). We investigated whether these risks depend on the parent-of-origin of the mutation. We studied 422 subjects, including 89 MMR gene mutation carriers, from 17 population-based families who were each recruited via a CRC case diagnosed before age 45 years and found to carry a MMR gene mutation. The POE hazard ratio (HR(POE)), defined to be the CRC incidence for carriers with maternally derived mutations divided by the corresponding paternal incidence, was estimated using a novel application of modified segregation analysis. HR(POE) (95% confidence interval) was estimated to be 3.2 (1.1-9.8) for males (P = 0.03) and 0.8 (0.2-2.8) for females (P = 0.5) and the corresponding cumulative risks to age 80 years were 88% (54%-100%) for male carriers with maternally derived mutations and 38-48% for all other carriers. If confirmed by larger studies, these results will have important implications for the etiology of CRC and for the clinical management of MMR gene mutation carriers. PMID:21120946

  8. DNA repair genes implicated in triple negative familial non-BRCA1/2 breast cancer predisposition

    PubMed Central

    Ollier, Marie; Radosevic-Robin, Nina; Kwiatkowski, Fabrice; Ponelle, Flora; Viala, Sandrine; Privat, Maud; Uhrhammer, Nancy; Bernard-Gallon, Dominique; Penault-Llorca, Frédérique; Bignon, Yves-Jean; Bidet, Yannick

    2015-01-01

    Among breast cancers, 10 to 15% of cases would be due to hereditary risk. In these familial cases, mutations in BRCA1 and BRCA2 are found in only 15% to 20%, meaning that new susceptibility genes remain to be found. Triple-negative breast cancers represent 15% of all breast cancers, and are generally aggressive tumours without targeted therapies available. Our hypothesis is that some patients with triple negative breast cancer could share a genetic susceptibility different from other types of breast cancers. We screened 36 candidate genes, using pyrosequencing, in all the 50 triple negative breast cancer patients with familial history of cancer but no BRCA1 or BRCA2 mutation of a population of 3000 families who had consulted for a familial breast cancer between 2005 and 2013. Any mutations were also sequenced in available relatives of cases. Protein expression and loss of heterozygosity were explored in tumours. Seven deleterious mutations in 6 different genes (RAD51D, MRE11A, CHEK2, MLH1, MSH6, PALB2) were observed in one patient each, except the RAD51D mutation found in two cases. Loss of heterozygosity in the tumour was found for 2 of the 7 mutations. Protein expression was absent in tumour tissue for 5 mutations. Taking into consideration a specific subtype of tumour has revealed susceptibility genes, most of them in the homologous recombination DNA repair pathway. This may provide new possibilities for targeted therapies, along with better screening and care of patients. PMID:26328243

  9. A Mutation in a Saccharomyces Cerevisiae Gene (Rad3) Required for Nucleotide Excision Repair and Transcription Increases the Efficiency of Mismatch Correction

    PubMed Central

    Yang, Y.; Johnson, A. L.; Johnston, L. H.; Siede, W.; Friedberg, E. C.; Ramachandran, K.; Kunz, B. A.

    1996-01-01

    RAD3 functions in DNA repair and transcription in Saccharomyces cerevisiae and particular rad3 alleles confer a mutator phenotype, possibly as a consequence of defective mismatch correction. We assessed the potential involvement of the Rad3 protein in mismatch correction by comparing heteroduplex repair in isogenic rad3-1 and wild-type strains. The rad3-1 allele increased the spontaneous mutation rate but did not prevent heteroduplex repair or bias its directionality. Instead, the efficiency of mismatch correction was enhanced in the rad3-1 strain. This surprising result prompted us to examine expression of yeast mismatch repair genes. We determined that MSH2, but not MLH1, is transcriptionally regulated during the cell-cycle like PMS1, and that rad3-1 does not increase the transcript levels for these genes in log phase cells. These observations suggest that the rad3-1 mutation gives rise to an enhanced efficiency of mismatch correction via a process that does not involve transcriptional regulation of mismatch repair. Interestingly, mismatch repair also was more efficient when error-editing by yeast DNA polymerase δ was eliminated. We discuss our results in relation to possible mechanisms that may link the rad3-1 mutation to mismatch correction efficiency. PMID:8889512

  10. Retinal detachments after Boston Keratoprosthesis: incidence, predisposing factors, and visual outcomes

    PubMed Central

    Jardeleza, Maria Stephanie R.; Rheaume, Marc-Andre; Chodosh, James; Lane, Anne Marie; Dohlman, Claes H.

    2015-01-01

    Purpose To determine the rates, predisposing factors, and visual outcomes of retinal detachment (RD) after Boston Keratoprosthesis (KPro) implantation. Methods In this noncomparative, interventional case series, the medical records of 170 patients (205 eyes) who underwent Boston type 1 and type 2 KPro implantation at the Massachusetts Eye and Ear Infirmary between April 1993 and June 2009 were retrospectively reviewed. Incidence and annual rates of RD were calculated, and the roles of possible predictive factors for RD after KPro were investigated. Main outcome measures were rates of and risk factors for RD, visual acuity after RD, and surgical outcomes after repair. Results Sterile vitritis and autoimmune systemic disease significantly predisposed patients to RD after KPro placement. Of patients who developed RD after implantation, 50% progressed to visual acuity of no light perception despite surgical repair. Conclusions Inflammation plays a major role in RD development after KPro implantation. Patients with predisposing factors should be advised of the high rates of RD and comanaged with a vitreoretinal specialist.

  11. The RAD7 and RAD16 genes, which are essential for pyrimidine dimer removal from the silent mating type loci, are also required for repair of the nontranscribed strand of an active gene in Saccharomyces cerevisiae.

    PubMed Central

    Verhage, R; Zeeman, A M; de Groot, N; Gleig, F; Bang, D D; van de Putte, P; Brouwer, J

    1994-01-01

    The rad16 mutant of Saccharomyces cerevisiae was previously shown to be impaired in removal of UV-induced pyrimidine dimers from the silent mating-type loci (D. D. Bang, R. A. Verhage, N. Goosen, J. Brouwer, and P. van de Putte, Nucleic Acids Res. 20:3925-3931, 1992). Here we show that rad7 as well as rad7 rad16 double mutants have the same repair phenotype, indicating that the RAD7 and RAD16 gene products might operate in the same nucleotide excision repair subpathway. Dimer removal from the genome overall is essentially incomplete in these mutants, leaving about 20 to 30% of the DNA unrepaired. Repair analysis of the transcribed RPB2 gene shows that the nontranscribed strand is not repaired at all in rad7 and rad16 mutants, whereas the transcribed strand is repaired in these mutants at a fast rate similar to that in RAD+ cells. When the results obtained with the RPB2 gene can be generalized, the RAD7 and RAD16 proteins not only are essential for repair of silenced regions but also function in repair of nontranscribed strands of active genes in S. cerevisiae. The phenotype of rad7 and rad16 mutants closely resembles that of human xeroderma pigmentosum complementation group C (XP-C) cells, suggesting that RAD7 and RAD16 in S. cerevisiae function in the same pathway as the XPC gene in human cells. RAD4, which on the basis of sequence homology has been proposed to be the yeast XPC counterpart, seems to be involved in repair of both inactive and active yeast DNA, challenging the hypothesis that RAD4 and XPC are functional homologs. Images PMID:8065346

  12. Variants in DNA double-strand break repair genes and risk of familial breast cancer in a South American population.

    PubMed

    Jara, Lilian; Dubois, Karen; Gaete, Daniel; de Mayo, Tomas; Ratkevicius, Nikalai; Bravo, Teresa; Margarit, Sonia; Blanco, Rafael; Gómez, Fernando; Waugh, Enrique; Peralta, Octavio; Reyes, Jose M; Ibáñez, Gladys; González-Hormazábal, Patricio

    2010-08-01

    The double-strand break (DSB) DNA repair pathway has been implicated in breast cancer (BC). RAD51 and its paralogs XRCC3 and RAD51D play an important role in the repair of DSB through homologous recombination (HR). Some polymorphisms including XRCC3-Thr241Met, RAD51-135G>C, and RAD51D-E233G have been found to confer increased BC susceptibility. In order to detect novel mutations that may contribute to BC susceptibility, 150 patients belonging to 150 Chilean BRCA1/2-negative families were screened for mutations in XRCC3. No mutations were detected in the XRCC3 gene. In addition, using a case-control design we studied the XRCC3-Thr241Met, and RAD51D-E233G polymorphisms in 267 BC cases and 500 controls to evaluate their possible association with BC susceptibility. The XRCC3 Met/Met genotype was associated with an increased BC risk (P = 0.003, OR = 2.44 [95%CI 1.34-4.43]). We did not find an association between E233G polymorphism and BC risk. We also analyzed the effect of combined genotypes among RAD51-135G>C, Thr241Met, and E233G polymorphisms on BC risk. No interaction was observed between Thr241Met and 135G>C. The combined genotype Thr/Met-E/G was associated with an increased BC risk among women who (a) have a family history of BC, (b) are BRCA1/2-negative, and (c) were <50 years at onset (n = 195) (P = 0.037, OR = 10.5 [95%CI 1.16-94.5]). Our results suggested that the variability of the DNA HR repair genes XRCC3 and RAD51D may play a role in BC risk, but this role may be underlined by a mutual interaction between these genes. These findings should be confirmed in other populations. PMID:20054644

  13. Low-level laser irradiation alters mRNA expression from genes involved in DNA repair and genomic stabilization in myoblasts

    NASA Astrophysics Data System (ADS)

    Trajano, L. A. S. N.; Sergio, L. P. S.; Silva, C. L.; Carvalho, L.; Mencalha, A. L.; Stumbo, A. C.; Fonseca, A. S.

    2016-07-01

    Low-level lasers are used for the treatment of diseases in soft and bone tissues, but few data are available regarding their effects on genomic stability. In this study, we investigated mRNA expression from genes involved in DNA repair and genomic stabilization in myoblasts exposed to low-level infrared laser. C2C12 myoblast cultures in different fetal bovine serum concentrations were exposed to low-level infrared laser (10, 35 and 70 J cm‑2), and collected for the evaluation of DNA repair gene expression. Laser exposure increased gene expression related to base excision repair (8-oxoguanine DNA glycosylase and apurinic/apyrimidinic endonuclease 1), nucleotide excision repair (excision repair cross-complementation group 1 and xeroderma pigmentosum C protein) and genomic stabilization (ATM serine/threonine kinase and tumor protein p53) in normal and low fetal bovine serum concentrations. Results suggest that genomic stability could be part of a biostimulation effect of low-level laser therapy in injured muscles.

  14. Correction of point mutations at the endogenous locus of the dihydrofolate reductase gene using repair-PolyPurine Reverse Hoogsteen hairpins in mammalian cells.

    PubMed

    Solé, Anna; Ciudad, Carlos J; Chasin, Lawrence A; Noé, Véronique

    2016-06-15

    Correction of point mutations that lead to aberrant transcripts, often with pathological consequences, has been the focus of considerable research. In this work, repair-PPRHs are shown to be a new powerful tool for gene correction. A repair-PPRH consists of a PolyPurine Reverse Hoogsteen hairpin core bearing an extension sequence at one end, homologous to the DNA strand to be repaired but containing the wild type nucleotide instead of the mutation. Previously, we had corrected a single-point mutation with repair-PPRHs using a mutated version of a dihydrofolate reductase (dhfr) minigene. To further evaluate the utility of these molecules, different repair-PPRHs were designed to correct insertions, deletions, substitutions and a double substitution present in a collection of mutants at the endogenous locus of the dhfr gene, the product of which is the target of the chemotherapeutic agent methotrexate. We also describe an approach to use when the point mutation is far away from the homopyrimidine target domain. This strategy consists in designing Long-Distance- and Short-Distance-Repair-PPRHs where the PPRH core is bound to the repair tail by a five-thymidine linker. Surviving colonies in a DHFR selective medium, lacking glycine and sources of purines and thymidine, were analyzed by DNA sequencing, and by mRNA, protein and enzymatic measurements, confirming that all the dhfr mutants had been corrected. These results show that repair-PPRHs can be effective tools to accomplish a permanent correction of point mutations in the DNA sequence of mutant mammalian cells. PMID:27063945

  15. Many amino acid substitution variants identified in DNA repair genes during human population screenings are predicted to impact protein function

    SciTech Connect

    Xi, T; Jones, I M; Mohrenweiser, H W

    2003-11-03

    Over 520 different amino acid substitution variants have been previously identified in the systematic screening of 91 human DNA repair genes for sequence variation. Two algorithms were employed to predict the impact of these amino acid substitutions on protein activity. Sorting Intolerant From Tolerant (SIFT) classified 226 of 508 variants (44%) as ''Intolerant''. Polymorphism Phenotyping (PolyPhen) classed 165 of 489 amino acid substitutions (34%) as ''Probably or Possibly Damaging''. Another 9-15% of the variants were classed as ''Potentially Intolerant or Damaging''. The results from the two algorithms are highly associated, with concordance in predicted impact observed for {approx}62% of the variants. Twenty one to thirty one percent of the variant proteins are predicted to exhibit reduced activity by both algorithms. These variants occur at slightly lower individual allele frequency than do the variants classified as ''Tolerant'' or ''Benign''. Both algorithms correctly predicted the impact of 26 functionally characterized amino acid substitutions in the APE1 protein on biochemical activity, with one exception. It is concluded that a substantial fraction of the missense variants observed in the general human population are functionally relevant. These variants are expected to be the molecular genetic and biochemical basis for the associations of reduced DNA repair capacity phenotypes with elevated cancer risk.

  16. Microarray-based expression of DNA repair genes does not correlate with growth inhibition of cancer cells by natural products derived from traditional Chinese medicine.

    PubMed

    Konkimalla, V S Badireenath; Wang, Gan; Kaina, Bernd; Efferth, Thomas

    2008-01-01

    Drug resistance represents a major obstacle in cancer chemotherapy. As chemically characterized compounds derived from plants used in traditional Chinese medicine (TCM) may have molecular targets different from those of standard antitumor drugs, they might be attractive candidates for novel therapeutics with improved pharmacological features. DNA repair is frequently involved in the development of resistance to established anticancer drugs, e.g. alkylating agents. Using a database of 531 chemically characterized TCM compounds from medicinal plants recently established by us, the IC50 values of 60 N.C.I. tumor cell lines for these 531 natural products were tested for correlation with the microarray-based mRNA expression of six genes involved in nucleotide excision repair (ERCC1, XPA, XPC, DDB2, ERCC4, ERCC5). No compound correlated with the expression of these genes, indicating that mRNA expression of these genes is not associated with resistance of the cell lines to these TCM compounds. The same is true for another six genes of the base excision repair pathway (MPG, APEX1, OGG1, XRCC1, LIG3, POLB). Microarray-based COMPARE analyses were performed to identify other candidate genes that are able to predict responsiveness of tumor cells to TCM-derived natural products. As an example, diallyl disulfide from garlic (Allium sativum L., Chinese name: dashuan) was chosen. Eighteen genes were identified whose mRNA expression predicted sensitivity or resistance to diallyl disulfide in hierarchical cluster analyses. Apart from some genes with still unknown function, genes were identified from different functional groups, e.g. signal transducers, regulators of GTPase activity, those associated with cytoskeleton formation and regulation, constituents of the ribosome. Remarkably, none of these genes have been described to be involved in DNA repair. In conclusion, our data indicate that TCM-derived natural products are worth being further investigated as novel compounds to

  17. What aspects of autism predispose to talent?

    PubMed Central

    Happé, Francesca; Vital, Pedro

    2009-01-01

    In this paper, we explore the question, why are striking special skills so much more common in autism spectrum conditions (ASC) than in other groups? Current cognitive accounts of ASC are briefly reviewed in relation to special skills. Difficulties in ‘theory of mind’ may contribute to originality in ASC, since individuals who do not automatically ‘read other minds’ may be better able to think outside prevailing fashions and popular theories. However, originality alone does not confer talent. Executive dysfunction has been suggested as the ‘releasing’ mechanism for special skills in ASC, but other groups with executive difficulties do not show raised incidence of talents. Detail-focused processing bias (‘weak coherence’, ‘enhanced perceptual functioning’) appears to be the most promising predisposing characteristic, or ‘starting engine’, for talent development. In support of this notion, we summarize data from a population-based twin study in which parents reported on their 8-year-olds' talents and their ASC-like traits. Across the whole sample, ASC-like traits, and specifically ‘restricted and repetitive behaviours and interests’ related to detail focus, were more pronounced in children reported to have talents outstripping older children. We suggest that detail-focused cognitive style predisposes to talent in savant domains in, and beyond, autism spectrum disorders. PMID:19528019

  18. Enrichment of G2/M cell cycle phase in human pluripotent stem cells enhances HDR-mediated gene repair with customizable endonucleases.

    PubMed

    Yang, Diane; Scavuzzo, Marissa A; Chmielowiec, Jolanta; Sharp, Robert; Bajic, Aleksandar; Borowiak, Malgorzata

    2016-01-01

    Efficient gene editing is essential to fully utilize human pluripotent stem cells (hPSCs) in regenerative medicine. Custom endonuclease-based gene targeting involves two mechanisms of DNA repair: homology directed repair (HDR) and non-homologous end joining (NHEJ). HDR is the preferred mechanism for common applications such knock-in, knock-out or precise mutagenesis, but remains inefficient in hPSCs. Here, we demonstrate that synchronizing synchronizing hPSCs in G2/M with ABT phase increases on-target gene editing, defined as correct targeting cassette integration, 3 to 6 fold. We observed improved efficiency using ZFNs, TALENs, two CRISPR/Cas9, and CRISPR/Cas9 nickase to target five genes in three hPSC lines: three human embryonic stem cell lines, neural progenitors and diabetic iPSCs. neural progenitors and diabetic iPSCs. Reversible synchronization has no effect on pluripotency or differentiation. The increase in on-target gene editing is locus-independent and specific to the cell cycle phase as G2/M phase enriched cells show a 6-fold increase in targeting efficiency compared to cells in G1 phase. Concurrently inhibiting NHEJ with SCR7 does not increase HDR or improve gene targeting efficiency further, indicating that HR is the major DNA repair mechanism after G2/M phase arrest. The approach outlined here makes gene editing in hPSCs a more viable tool for disease modeling, regenerative medicine and cell-based therapies. PMID:26887909

  19. Enrichment of G2/M cell cycle phase in human pluripotent stem cells enhances HDR-mediated gene repair with customizable endonucleases

    PubMed Central

    Yang, Diane; Scavuzzo, Marissa A; Chmielowiec, Jolanta; Sharp, Robert; Bajic, Aleksandar; Borowiak, Malgorzata

    2016-01-01

    Efficient gene editing is essential to fully utilize human pluripotent stem cells (hPSCs) in regenerative medicine. Custom endonuclease-based gene targeting involves two mechanisms of DNA repair: homology directed repair (HDR) and non-homologous end joining (NHEJ). HDR is the preferred mechanism for common applications such knock-in, knock-out or precise mutagenesis, but remains inefficient in hPSCs. Here, we demonstrate that synchronizing synchronizing hPSCs in G2/M with ABT phase increases on-target gene editing, defined as correct targeting cassette integration, 3 to 6 fold. We observed improved efficiency using ZFNs, TALENs, two CRISPR/Cas9, and CRISPR/Cas9 nickase to target five genes in three hPSC lines: three human embryonic stem cell lines, neural progenitors and diabetic iPSCs. neural progenitors and diabetic iPSCs. Reversible synchronization has no effect on pluripotency or differentiation. The increase in on-target gene editing is locus-independent and specific to the cell cycle phase as G2/M phase enriched cells show a 6-fold increase in targeting efficiency compared to cells in G1 phase. Concurrently inhibiting NHEJ with SCR7 does not increase HDR or improve gene targeting efficiency further, indicating that HR is the major DNA repair mechanism after G2/M phase arrest. The approach outlined here makes gene editing in hPSCs a more viable tool for disease modeling, regenerative medicine and cell-based therapies. PMID:26887909

  20. Genetic re-engineering of Saccharomyces cerevisiae RAD51 leads to a significant increase in the frequency of gene repair in vivo

    PubMed Central

    Liu, Li; Maguire, Katie K.; Kmiec, Eric B.

    2004-01-01

    Oligonucleotides can be used to direct the alteration of single nucleotides in chromosomal genes in yeast. Rad51 protein appears to play a central role in catalyzing the reaction, most likely through its DNA pairing function. Here, we re-engineer the RAD51 gene in order to produce proteins bearing altered levels of known activities. Overexpression of wild-type ScRAD51 elevates the correction of an integrated, mutant hygromycin resistance gene ∼3-fold. Overexpression of an altered RAD51 gene, which encodes a protein that has a higher affinity for ScRad54, enhances the targeting frequency nearly 100-fold. Another mutation which increases the affinity of Rad51 for DNA was also found to increase gene repair when overexpressed in the cell. Other mutations in the Rad51 protein, such as one that reduces interaction with Rad52, has little or no effect on the frequency of gene repair. These data provide the first evidence that the Rad51 protein can be modified so as to increase the frequency of gene repair in yeast. PMID:15087488

  1. Evaluation of CADD Scores in Curated Mismatch Repair Gene Variants Yields a Model for Clinical Validation and Prioritization

    PubMed Central

    van der Velde, K. Joeri; Kuiper, Joël; Thompson, Bryony A.; Plazzer, John‐Paul; van Valkenhoef, Gert; de Haan, Mark; Jongbloed, Jan D.H.; Wijmenga, Cisca; de Koning, Tom J.; Abbott, Kristin M.; Sinke, Richard; Spurdle, Amanda B.; Macrae, Finlay; Genuardi, Maurizio; Sijmons, Rolf H.

    2015-01-01

    ABSTRACT Next‐generation sequencing in clinical diagnostics is providing valuable genomic variant data, which can be used to support healthcare decisions. In silico tools to predict pathogenicity are crucial to assess such variants and we have evaluated a new tool, Combined Annotation Dependent Depletion (CADD), and its classification of gene variants in Lynch syndrome by using a set of 2,210 DNA mismatch repair gene variants. These had already been classified by experts from InSiGHT's Variant Interpretation Committee. Overall, we found CADD scores do predict pathogenicity (Spearman's ρ = 0.595, P < 0.001). However, we discovered 31 major discrepancies between the InSiGHT classification and the CADD scores; these were explained in favor of the expert classification using population allele frequencies, cosegregation analyses, disease association studies, or a second‐tier test. Of 751 variants that could not be clinically classified by InSiGHT, CADD indicated that 47 variants were worth further study to confirm their putative pathogenicity. We demonstrate CADD is valuable in prioritizing variants in clinically relevant genes for further assessment by expert classification teams. PMID:25871441

  2. LNA modification of single-stranded DNA oligonucleotides allows subtle gene modification in mismatch-repair-proficient cells.

    PubMed

    van Ravesteyn, Thomas W; Dekker, Marleen; Fish, Alexander; Sixma, Titia K; Wolters, Astrid; Dekker, Rob J; Te Riele, Hein P J

    2016-04-12

    Synthetic single-stranded DNA oligonucleotides (ssODNs) can be used to generate subtle genetic modifications in eukaryotic and prokaryotic cells without the requirement for prior generation of DNA double-stranded breaks. However, DNA mismatch repair (MMR) suppresses the efficiency of gene modification by >100-fold. Here we present a commercially available ssODN design that evades MMR and enables subtle gene modification in MMR-proficient cells. The presence of locked nucleic acids (LNAs) in the ssODNs at mismatching bases, or also at directly adjacent bases, allowed 1-, 2-, or 3-bp substitutions in MMR-proficient mouse embryonic stem cells as effectively as in MMR-deficient cells. Additionally, in MMR-proficient Escherichia coli, LNA modification of the ssODNs enabled effective single-base-pair substitution. In vitro, LNA modification of mismatches precluded binding of purified E. coli MMR protein MutS. These findings make ssODN-directed gene modification particularly well suited for applications that require the evaluation of a large number of sequence variants with an easy selectable phenotype. PMID:26951689

  3. Male infertility, genetic analysis of the DAZ genes on the human Y chromosome and genetic analysis of DNA repair.

    PubMed

    Fox, M S; Reijo Pera, R A

    2001-11-26

    Many genes that are required for fertility have been identified in model organisms (). Mutations in these genes cause infertility due to defects in development of the germ cell lineage, but the organism is otherwise healthy. Although human reproduction is undoubtedly as complex as that of other organisms, very few fertility loci have been mapped (). This is in spite of the prevalence of human infertility, the lack of effective treatments to remedy germ cell defects, and the cost to couples and society of assisted reproductive techniques. Fifteen percent of couples are infertile and half of all cases can be traced to the male partner. Aside from defects in sperm production, most infertile men are otherwise healthy. This review is divided into two distinct parts to discuss work that: (i) led to the identification of several genes on the Y chromosome that likely function in sperm production; and (ii) implicates DNA repair in male infertility via increased frequency of mutations in DNA from men with meiotic arrest. PMID:11694340

  4. In GERD patients, mucosal repair associated genes are upregulated in non-inflamed oesophageal epithelium

    PubMed Central

    De Vries, D R; Ter Linde, J J M; Van Herwaarden, M A; Schwartz, M P; Shephard, P; Geng, M M; Smout, A J P M; Samsom, M

    2009-01-01

    Abstract Previous studies addressing the effects of acid reflux and PPI therapy on gene expression in oesophageal epithelium concentrated on inflamed tissue. We aimed to determine changes in gene expression in non-inflamed oesophageal epithelium of GERD patients. Therefore, we included 20 GERD patients with pathological total 24-hr acid exposure of 6–12% and SAP ≥ 95%. Ten patients discontinued PPI treatment (PPI-), 10 took pantoprazole 40 mg bid (PPI+). Ten age/sex-matched healthy controls were recruited. Biopsies were taken from non-inflamed mucosa 6 cm and 16 cm proximal to the squamocolumnar junction (SCJ). Gene expression profiling of biopsies from 6 cm was performed on Human Genome U133 Plus 2.0 arrays (Affymetrix). Genes exhibiting a fold change >1.4 (t-test P-value < 1E– 4) were considered differentially expressed. Results were confirmed by real-time RT-PCR. In PPI- patients, 92 microarray probesets were deregulated. The majority of the corresponding genes were associated with cell–cell contacts, cytoskeletal reorganization and cellular motility, suggesting facilitation of a migratory phenotype. Genes encoding proteins with anti-apoptotic or anti-proliferative functions or stress-protective functions were also deregulated. No probesets were deregulated in PPI+ patients. QPCR analysis of 20 selected genes confirmed most of the deregulations in PPI- patients, and showed several deregulated genes in PPI+ patients as well. In the biopsies taken at 16 cm QPCR revealed no deregulations of the selected genes. We conclude that upon acid exposure, oesophageal epithelial cells activate a process globally known as epithelial restitution: up-regulation of anti-apoptotic, anti-oxidant and migration associated genes. Possibly this process helps maintaining barrier function. PMID:19413890

  5. Tissue-Specific Effects of Valproic Acid on DNA Repair Genes and Apoptosis in Postimplantation Mouse Embryos

    PubMed Central

    Lamparter, Christina; Winn, Louise M.

    2014-01-01

    Exposure to the anticonvulsant drug valproic acid (VPA) is associated with an increased risk of congenital malformations. Although the mechanisms contributing to its teratogenicity are poorly understood, VPA has been shown to induce DNA double strand breaks (DSB) and to increase homologous recombination in vitro. The objective of the present study was to determine whether in utero exposure to VPA alters the frequency of intrachromosomal recombination and the expression of several genes involved in DSB repair in pKZ1 mouse embryos. Pregnant pKZ1 transgenic mice (GD 9.0) were administered VPA (500 mg/kg s.c.) and embryos were extracted and microdissected into the head, heart, and trunk regions 1, 3, 6, and 24 h after injection. Quantitative PCR was used to measure the tissue-specific expression of lacZ, a surrogate measure of recombination, Xrcc4, Rad51, Brca1, and Brca2, with Western blotting used to quantify Rad51, cleaved caspase-3 and cleaved-PARP protein. Increased recombination was only observed in the embryonic head following 6-h VPA exposure. VPA had no effect on Xrcc4 expression. Rad51, Brca1, and Brca2 expression rapidly decreased in head and trunk tissues after 1-h VPA exposure, followed by a subsequent increase in all tissues, although it was generally attenuated in the head and not due to differences in endogenous levels. Cleaved caspase-3 and cleaved-PARP expression was increased in all tissues 3 h following VPA exposure. This study indicates that the tissue-specific expression of several genes involved in DSB repair is altered following exposure to VPA and may be contributing to increased apoptosis. PMID:24913804

  6. Meningocele repair

    MedlinePlus

    ... dysraphism repair; Meningomyelocele repair; Neural tube defect repair; Spina bifida repair ... a medical team with experience in children with spina bifida. Your baby will likely have an MRI (magnetic ...

  7. Challenges and complexities in estimating both the functional impact and the disease risk associated with the extensive genetic variation in human DNA repair genes.

    PubMed

    Mohrenweiser, Harvey W; Wilson, David M; Jones, Irene M

    2003-05-15

    Individual risk and the population incidence of disease result from the interaction of genetic susceptibility and exposure. DNA repair is an example of a cellular process where genetic variation in families with extreme predisposition is documented to be associated with high disease likelihood, including syndromes of premature aging and cancer. Although the identification and characterization of new genes or variants in cancer families continues to be important, the focus of this paper is the current status of efforts to define the impact of polymorphic amino acid substitutions in DNA repair genes on individual and population cancer risk. There is increasing evidence that mild reductions in DNA repair capacity, assumed to be the consequence of common genetic variation, affect cancer predisposition. The extensive variation being found in the coding regions of DNA repair genes and the large number of genes in each of the major repair pathways results in complex genotypes with potential to impact cancer risk in the general population. The implications of this complexity for molecular epidemiology studies, as well as concepts that may make these challenges more manageable, are discussed. The concepts include both experimental and computational approaches that could be employed to develop predictors of disease susceptibility based on DNA repair genotype, focusing initially on studies to assess functional impact on individual proteins and pathways and then on molecular epidemiology studies to assess exposure-dependent health risk. In closing, we raise some of the non-technical challenges to the utilization of the full richness of the genetic variation to reduce disease occurrence and ultimately improve health care. PMID:12714187

  8. Selection-free gene repair after adenoviral vector transduction of designer nucleases: rescue of dystrophin synthesis in DMD muscle cell populations.

    PubMed

    Maggio, Ignazio; Stefanucci, Luca; Janssen, Josephine M; Liu, Jin; Chen, Xiaoyu; Mouly, Vincent; Gonçalves, Manuel A F V

    2016-02-18

    Duchenne muscular dystrophy (DMD) is a fatal X-linked muscle-wasting disorder caused by mutations in the 2.4 Mb dystrophin-encoding DMD gene. The integration of gene delivery and gene editing technologies based on viral vectors and sequence-specific designer nucleases, respectively, constitutes a potential therapeutic modality for permanently repairing defective DMD alleles in patient-derived myogenic cells. Therefore, we sought to investigate the feasibility of combining adenoviral vectors (AdVs) with CRISPR/Cas9 RNA-guided nucleases (RGNs) alone or together with transcriptional activator-like effector nucleases (TALENs), for endogenous DMD repair through non-homologous end-joining (NHEJ). The strategies tested involved; incorporating small insertions or deletions at out-of-frame sequences for reading frame resetting, splice acceptor knockout for DNA-level exon skipping, and RGN-RGN or RGN-TALEN multiplexing for targeted exon(s) removal. We demonstrate that genome editing based on the activation and recruitment of the NHEJ DNA repair pathway after AdV delivery of designer nuclease genes, is a versatile and robust approach for repairing DMD mutations in bulk populations of patient-derived muscle progenitor cells (up to 37% of corrected DMD templates). These results open up a DNA-level genetic medicine strategy in which viral vector-mediated transient designer nuclease expression leads to permanent and regulated dystrophin synthesis from corrected native DMD alleles. PMID:26762977

  9. Selection-free gene repair after adenoviral vector transduction of designer nucleases: rescue of dystrophin synthesis in DMD muscle cell populations

    PubMed Central

    Maggio, Ignazio; Stefanucci, Luca; Janssen, Josephine M.; Liu, Jin; Chen, Xiaoyu; Mouly, Vincent; Gonçalves, Manuel A.F.V.

    2016-01-01

    Duchenne muscular dystrophy (DMD) is a fatal X-linked muscle-wasting disorder caused by mutations in the 2.4 Mb dystrophin-encoding DMD gene. The integration of gene delivery and gene editing technologies based on viral vectors and sequence-specific designer nucleases, respectively, constitutes a potential therapeutic modality for permanently repairing defective DMD alleles in patient-derived myogenic cells. Therefore, we sought to investigate the feasibility of combining adenoviral vectors (AdVs) with CRISPR/Cas9 RNA-guided nucleases (RGNs) alone or together with transcriptional activator-like effector nucleases (TALENs), for endogenous DMD repair through non-homologous end-joining (NHEJ). The strategies tested involved; incorporating small insertions or deletions at out-of-frame sequences for reading frame resetting, splice acceptor knockout for DNA-level exon skipping, and RGN-RGN or RGN-TALEN multiplexing for targeted exon(s) removal. We demonstrate that genome editing based on the activation and recruitment of the NHEJ DNA repair pathway after AdV delivery of designer nuclease genes, is a versatile and robust approach for repairing DMD mutations in bulk populations of patient-derived muscle progenitor cells (up to 37% of corrected DMD templates). These results open up a DNA-level genetic medicine strategy in which viral vector-mediated transient designer nuclease expression leads to permanent and regulated dystrophin synthesis from corrected native DMD alleles. PMID:26762977

  10. A Study of Molecular Signals Deregulating Mismatch Repair Genes in Prostate Cancer Compared to Benign Prostatic Hyperplasia

    PubMed Central

    Basu, Sanmitra; Majumder, Subhadipa; Bhowal, Ankur; Ghosh, Alip; Naskar, Sukla; Nandy, Sumit; Mukherjee, Subhabrata; Sinha, Rajan Kumar; Basu, Keya; Karmakar, Dilip; Banerjee, Soma; Sengupta, Sanghamitra

    2015-01-01

    Prostate cancer is one of the leading causes of mortality among aging males. There is an unmet requirement of clinically useful biomarkers for early detection of prostate cancer to reduce the liabilities of overtreatment and accompanying morbidity. The present population-based study investigates the factors disrupting expression of multiple functionally related genes of DNA mismatch repair pathway in prostate cancer patients to identify molecular attributes distinguishing adenocarcinoma from benign hyperplasia of prostate. Gene expression was compared between tissue samples from prostate cancer and benign prostatic hyperplasia using real-time-PCR, western blot and immunohistochemistry. Assessment of genotypes of seven single-nucleotide-polymorphisms of three MMR genes was conducted using PCR-coupled RFLP and sequencing. Promoter methylation was interrogated by methylation-specific-PCR and bisulfite-sequencing. Interaction between microRNAs and MMR genes was verified by 3'UTR-based dual luciferase assays. Concurrent reduction of three MMR genes namely hMLH1, hMSH6 and hMSH2 (34-85%, P<0.05) was observed in prostate cancer tissues. hMSH6 polymorphism rs1800932(Pro92Pro) conferred a borderline protection in cancer patients (OR = 0.33, 95% CI = 0.15-0.75). Relative transcript level of hMLH1 was inversely related (r = -0.59, P<0.05) with methylation quotient of its promoter which showed a significantly higher methylation density (P = 0.008, Z = -2.649) in cancer patients. hsa-miR-155, hsa-miR-141 and hsa-miR-21 gene expressions were significantly elevated (66-85%, P<0.05) in tumor specimens and negatively correlated (r = -0.602 to -0.527, P<0.05) with that of MMR genes. hsa-miR-155 & hsa-miR-141 and hsa-miR-155 & hsa-miR-21 were demonstrated to bind to their putative seed sequences in hMLH1 and hMSH6 3’UTRs respectively. Relatively higher expression of DNA methyl-transferases (DNMT1 and DNMT3b) and HIF-1α genes (34-50%, P<0.05) were also detected in tumor tissues

  11. A study of molecular signals deregulating mismatch repair genes in prostate cancer compared to benign prostatic hyperplasia.

    PubMed

    Basu, Sanmitra; Majumder, Subhadipa; Bhowal, Ankur; Ghosh, Alip; Naskar, Sukla; Nandy, Sumit; Mukherjee, Subhabrata; Sinha, Rajan Kumar; Basu, Keya; Karmakar, Dilip; Banerjee, Soma; Sengupta, Sanghamitra

    2015-01-01

    Prostate cancer is one of the leading causes of mortality among aging males. There is an unmet requirement of clinically useful biomarkers for early detection of prostate cancer to reduce the liabilities of overtreatment and accompanying morbidity. The present population-based study investigates the factors disrupting expression of multiple functionally related genes of DNA mismatch repair pathway in prostate cancer patients to identify molecular attributes distinguishing adenocarcinoma from benign hyperplasia of prostate. Gene expression was compared between tissue samples from prostate cancer and benign prostatic hyperplasia using real-time-PCR, western blot and immunohistochemistry. Assessment of genotypes of seven single-nucleotide-polymorphisms of three MMR genes was conducted using PCR-coupled RFLP and sequencing. Promoter methylation was interrogated by methylation-specific-PCR and bisulfite-sequencing. Interaction between microRNAs and MMR genes was verified by 3'UTR-based dual luciferase assays. Concurrent reduction of three MMR genes namely hMLH1, hMSH6 and hMSH2 (34-85%, P<0.05) was observed in prostate cancer tissues. hMSH6 polymorphism rs1800932(Pro92Pro) conferred a borderline protection in cancer patients (OR = 0.33, 95% CI = 0.15-0.75). Relative transcript level of hMLH1 was inversely related (r = -0.59, P<0.05) with methylation quotient of its promoter which showed a significantly higher methylation density (P = 0.008, Z = -2.649) in cancer patients. hsa-miR-155, hsa-miR-141 and hsa-miR-21 gene expressions were significantly elevated (66-85%, P<0.05) in tumor specimens and negatively correlated (r = -0.602 to -0.527, P<0.05) with that of MMR genes. hsa-miR-155 & hsa-miR-141 and hsa-miR-155 & hsa-miR-21 were demonstrated to bind to their putative seed sequences in hMLH1 and hMSH6 3'UTRs respectively. Relatively higher expression of DNA methyl-transferases (DNMT1 and DNMT3b) and HIF-1α genes (34-50%, P<0.05) were also detected in tumor tissues. This

  12. The Cellular Response to Oxidatively Induced DNA Damage and Polymorphism of Some DNA Repair Genes Associated with Clinicopathological Features of Bladder Cancer

    PubMed Central

    Savina, Nataliya V.; Nikitchenko, Nataliya V.; Kuzhir, Tatyana D.; Rolevich, Alexander I.; Krasny, Sergei A.; Goncharova, Roza I.

    2016-01-01

    Genome instability and impaired DNA repair are hallmarks of carcinogenesis. The study was aimed at evaluating the DNA damage response in H2O2-treated lymphocytes using the alkaline comet assay in bladder cancer (BC) patients as compared to clinically healthy controls, elderly persons, and individuals with chronic inflammations. Polymorphism in DNA repair genes involved in nucleotide excision repair (NER) and base excision repair (BER) was studied using the PCR-RFLP method in the Belarusian population to elucidate the possible association of their variations with both bladder cancer risk and clinicopathological features of tumors. The increased level of H2O2-induced DNA damage and a higher proportion of individuals sensitive to oxidative stress were found among BC patients as compared to other groups under study. Heterozygosity in the XPD gene (codon 751) increased cancer risk: OR (95% CI) = 1.36 (1.03–1.81), p = 0.031. The frequency of the XPD 312Asn allele was significantly higher in T ≥ 2 high grade than in T ≥ 2 low grade tumors (p = 0.036); the ERCC6 1097Val/Val genotype was strongly associated with muscle-invasive tumors. Combinations of homozygous wild type alleles occurred with the increased frequency in patients with non-muscle-invasive tumors suggesting that the maintenance of normal DNA repair activity may prevent cancer progression. PMID:26649138

  13. Cox4i2, Ifit2, and Prdm11 Mutant Mice: Effective Selection of Genes Predisposing to an Altered Airway Inflammatory Response from a Large Compendium of Mutant Mouse Lines

    PubMed Central

    Bönisch, Clemens; Côme, Christophe; Kolster-Fog, Cathrine; Jensen, Klaus T.; Lund, Anders H.; Lee, Icksoo; Grossman, Lawrence I.; Sinkler, Christopher; Hüttemann, Maik; Bohn, Erwin; Fuchs, Helmut; Ollert, Markus; Gailus-Durner, Valérie; Hrabĕ de Angelis, Martin; Beckers, Johannes

    2015-01-01

    We established a selection strategy to identify new models for an altered airway inflammatory response from a large compendium of mutant mouse lines that were systemically phenotyped in the German Mouse Clinic (GMC). As selection criteria we included published gene functional data, as well as immunological and transcriptome data from GMC phenotyping screens under standard conditions. Applying these criteria we identified a few from several hundred mutant mouse lines and further characterized the Cox4i2tm1Hutt, Ifit2tm1.1Ebsb, and Prdm11tm1.1ahl lines following ovalbumin (OVA) sensitization and repeated OVA airway challenge. Challenged Prdm11tm1.1ahl mice exhibited changes in B cell counts, CD4+ T cell counts, and in the number of neutrophils in bronchoalveolar lavages, whereas challenged Ifit2tm1.1Ebsb mice displayed alterations in plasma IgE, IgG1, IgG3, and IgM levels compared to the challenged wild type littermates. In contrast, challenged Cox4i2tm1Hutt mutant mice did not show alterations in the humoral or cellular immune response compared to challenged wild type mice. Transcriptome analyses from lungs of the challenged mutant mouse lines showed extensive changes in gene expression in Prdm11tm1.1ahl mice. Functional annotations of regulated genes of all three mutant mouse lines were primarily related to inflammation and airway smooth muscle (ASM) remodeling. We were thus able to define an effective selection strategy to identify new candidate genes for the predisposition to an altered airway inflammatory response under OVA challenge conditions. Similar selection strategies may be used for the analysis of additional genotype – envirotype interactions for other diseases. PMID:26263558

  14. Genetic variants in DNA double-strand break repair genes and risk of salivary gland carcinoma: a case-control study.

    PubMed

    Xu, Li; Tang, Hongwei; El-Naggar, Adel K; Wei, Peng; Sturgis, Erich M

    2015-01-01

    DNA double strand break (DSB) repair is the primary defense mechanism against ionizing radiation-induced DNA damage. Ionizing radiation is the only established risk factor for salivary gland carcinoma (SGC). We hypothesized that genetic variants in DSB repair genes contribute to individual variation in susceptibility to SGC. To test this hypothesis, we conducted a case-control study in which we analyzed 415 single nucleotide polymorphisms (SNPs) in 45 DSB repair genes in 352 SGC cases and 598 controls. Multivariate logistic regression analysis was performed to calculate odds ratios (ORs) and 95% confidence intervals (CIs). Rs3748522 in RAD52 and rs13180356 in XRCC4 were significantly associated with SGC after Bonferroni adjustment; ORs (95% CIs) for the variant alleles of these SNPs were 1.71 (1.40-2.09, P = 1.70 × 10(-7)) and 0.58 (0.45-0.74, P = 2.00 × 10(-5)) respectively. The genetic effects were modulated by histological subtype. The association of RAD52-rs3748522 with SGC was strongest for mucoepidermoid carcinoma (OR = 2.21, 95% CI: 1.55-3.15, P = 1.25 × 10(-5), n = 74), and the association of XRCC4-rs13180356 with SGC was strongest for adenoid cystic carcinoma (OR = 0.60, 95% CI: 0.42-0.87, P = 6.91 × 10(-3), n = 123). Gene-level association analysis revealed one gene, PRKDC, with a marginally significant association with SGC risk in non-Hispanic whites. To our knowledge, this study is the first to comprehensively evaluate the genetic effect of DSB repair genes on SGC risk. Our results indicate that genetic variants in the DSB repair pathways contribute to inter-individual differences in susceptibility to SGC and show that the impact of genetic variants differs by histological subtype. Independent studies are warranted to confirm these findings. PMID:26035306

  15. Repair of uv damaged DNA: Genes and proteins of yeast and human. Progress report, November 1, 1991--April 15, 1992

    SciTech Connect

    Prakash, L.

    1992-04-01

    Our objectives are to determine the molecular mechanism of the incision step of excision repair of ultraviolet (UV) light damaged DNA in eukaryotic organisms, using the yeast Saccharomyces cerevisiae as a model system, and to study the human homologs of yeast excision repair and postreplication repair proteins progress is described.

  16. Mismatch repair gene defects contribute to the genetic basis of double primary cancers of the colorectum and endometrium.

    PubMed

    Millar, A L; Pal, T; Madlensky, L; Sherman, C; Temple, L; Mitri, A; Cheng, H; Marcus, V; Gallinger, S; Redston, M; Bapat, B; Narod, S

    1999-05-01

    Hereditary non-polyposis colorectal cancer (HNPCC) is a dominantly inherited cancer syndrome caused by germline defects of mismatch repair (MMR) genes. Endometrial cancer is the most common extracolonic neoplasm in HNPCC and is the primary clinical manifestation of the syndrome in some families. The cumulative incidence of endometrial cancer among HNPCC mutation carriers is high, estimated to be from 22 to 43%. We hypothesized that women with double primary cancers of the colorectum and endometrium are likely to be members of HNPCC families. In order to determine how frequently HNPCC manifests in the context of double primary cancers, we examined alterations of two MMR genes, hMSH2 and hMLH1, in 40 unrelated women affected with double primary cancers. These cases were identified using hospital-based and population-based cancer registries in Ontario, Canada. MMR gene mutations were screened by single-strand conformation polymorphism analysis and confirmed by direct sequencing. Eighteen percent (seven of 40) were found to harbor mutations of one of the two MMR genes. Analysis of colorectal and/or endometrial tumors of mutation-negative probands found microsatellite instability in seven of 20 cases. Six of seven mutation-positive probands had strong family histories suggestive of HNPCC. First degree relatives of mutation-positive probands had a very high relative risk (RR) of colorectal cancer (RR = 8.1, CI 3. 5-15.9) and endometrial cancer (RR = 23.8, CI 6.4-61.0). The relative risk of mutation-negative cases was 2.8 (CI 1.7-4.5) for colorectal cancer and 5.4 (CI 2.0-11.7) for endometrial cancer. We recommend that all double primary patients with cancers at these sites should have a genetic evaluation, including molecular analysis for HNPCC where appropriate. PMID:10196371

  17. Recent developments with the human repair genes ERCC2, ERCC4, and XRCC1

    SciTech Connect

    Thompson, L.H.; Caldecott, K.W.; Brookman, K.W.; Weber, C.A.; Salazar, E.S.; Takayama, K.; Fornace, A.J.

    1992-11-06

    ERCC2 was first identified as a gene on human chromosome 19 that complemented the UV sensitivity of CHO UV5 cells in somatic cell hybrids. Subsequent studies localized ERCC2 to the same chromosomal region (19q13.2--13.3) as the ERCC1 gene and showed that the two genes were less than 250 kb apart. Cloning of ERCC2 was accomplished by transfection of genomic DNA into UV5 cells and rescue of the gene from a secondary transformant. Recovery of the gene was aided by the presence of repetitive sequences that were detected on Southern blots with a probe for Alu-family repeats. ERCC2, which is 19 kb in size, quantitatively corrected the UV sensitivity and incision defect in UV5 cells upon transfection. An ERCC2 CDNA clone was recovered from the pcD2 expression library. Although this clone was truncated at the 5 in. end, it conferred transient, but not stable, correction to UV5 cells upon transfection. Based on genomic sequence, this clone was extended by oligonucleotide addition to obtain minigene constructs in which the complete open reading frame (ORF) was present. Translation of the ERCC2 ORF gives an amino acid sequence that has 72% similarity with the S. cerevisiae RAD3 protein, which encodes a DNA helicase.

  18. Candidate driver genes involved in genome maintenance and DNA repair in Sézary syndrome.

    PubMed

    Woollard, Wesley J; Pullabhatla, Venu; Lorenc, Anna; Patel, Varsha M; Butler, Rosie M; Bayega, Anthony; Begum, Nelema; Bakr, Farrah; Dedhia, Kiran; Fisher, Joshua; Aguilar-Duran, Silvia; Flanagan, Charlotte; Ghasemi, Aria A; Hoffmann, Ricarda M; Castillo-Mosquera, Nubia; Nuttall, Elisabeth A; Paul, Arisa; Roberts, Ceri A; Solomonidis, Emmanouil G; Tarrant, Rebecca; Yoxall, Antoinette; Beyers, Carl Z; Ferreira, Silvia; Tosi, Isabella; Simpson, Michael A; de Rinaldis, Emanuele; Mitchell, Tracey J; Whittaker, Sean J

    2016-06-30

    Sézary syndrome (SS) is a leukemic variant of cutaneous T-cell lymphoma (CTCL) and represents an ideal model for study of T-cell transformation. We describe whole-exome and single-nucleotide polymorphism array-based copy number analyses of CD4(+) tumor cells from untreated patients at diagnosis and targeted resequencing of 101 SS cases. A total of 824 somatic nonsynonymous gene variants were identified including indels, stop-gain/loss, splice variants, and recurrent gene variants indicative of considerable molecular heterogeneity. Driver genes identified using MutSigCV include POT1, which has not been previously reported in CTCL; and TP53 and DNMT3A, which were also identified consistent with previous reports. Mutations in PLCG1 were detected in 11% of tumors including novel variants not previously described in SS. This study is also the first to show BRCA2 defects in a significant proportion (14%) of SS tumors. Aberrations in PRKCQ were found to occur in 20% of tumors highlighting selection for activation of T-cell receptor/NF-κB signaling. A complex but consistent pattern of copy number variants (CNVs) was detected and many CNVs involved genes identified as putative drivers. Frequent defects involving the POT1 and ATM genes responsible for telomere maintenance were detected and may contribute to genomic instability in SS. Genomic aberrations identified were enriched for genes implicated in cell survival and fate, specifically PDGFR, ERK, JAK STAT, MAPK, and TCR/NF-κB signaling; epigenetic regulation (DNMT3A, ASLX3, TET1-3); and homologous recombination (RAD51C, BRCA2, POLD1). This study now provides the basis for a detailed functional analysis of malignant transformation of mature T cells and improved patient stratification and treatment. PMID:27121473

  19. Mutations in the transcriptional repressor REST predispose to Wilms tumor.

    PubMed

    Mahamdallie, Shazia S; Hanks, Sandra; Karlin, Kristen L; Zachariou, Anna; Perdeaux, Elizabeth R; Ruark, Elise; Shaw, Chad A; Renwick, Alexander; Ramsay, Emma; Yost, Shawn; Elliott, Anna; Birch, Jillian; Capra, Michael; Gray, Juliet; Hale, Juliet; Kingston, Judith; Levitt, Gill; McLean, Thomas; Sheridan, Eamonn; Renwick, Anthony; Seal, Sheila; Stiller, Charles; Sebire, Neil; Westbrook, Thomas F; Rahman, Nazneen

    2015-12-01

    Wilms tumor is the most common childhood renal cancer. To identify mutations that predispose to Wilms tumor, we are conducting exome sequencing studies. Here we describe 11 different inactivating mutations in the REST gene (encoding RE1-silencing transcription factor) in four familial Wilms tumor pedigrees and nine non-familial cases. Notably, no similar mutations were identified in the ICR1000 control series (13/558 versus 0/993; P < 0.0001) or in the ExAC series (13/558 versus 0/61,312; P < 0.0001). We identified a second mutational event in two tumors, suggesting that REST may act as a tumor-suppressor gene in Wilms tumor pathogenesis. REST is a zinc-finger transcription factor that functions in cellular differentiation and embryonic development. Notably, ten of 11 mutations clustered within the portion of REST encoding the DNA-binding domain, and functional analyses showed that these mutations compromise REST transcriptional repression. These data establish REST as a Wilms tumor predisposition gene accounting for ∼2% of Wilms tumor. PMID:26551668

  20. Defects in the DNA repair and transcription gene ERCC2(XPD) in trichothiodystrophy

    SciTech Connect

    Takayama, K.; Salazar, E.P.; Thompson, L.H.

    1996-02-01

    Trichothiodystrophy (TTD) is a rare autosomal recessive disorder characterized by brittle hair with reduced sulfur content, ichthyosis, peculiar face, and mental and growth retardation. Clinical photosensitivity is present in {approximately}50% of TTD patients but is not associated with an elevated frequency of cancers. Previous complementation studies show that the photosensitivity in nearly all of the studied patients is due to a defect in the same genetic locus that underlies the cancer-prone genetic disorder xeroderma pigmentosum group D (XP-D). Nucleotide-sequence analysis of the ERCC2 cDNA from three TTD cell strains (TTD1VI, TTD3VI, and TTD1RO) revealed mutations within the region from amino acid 713-730 and within previously identified helicase functional domains. The various clinical presentations and DNA repair characteristics of the cell strains can be correlated with the particular mutations found in the ERCC2 locus. Mutations of Arg658 to either His or Cys correlate with TTD cell strains with intermediate UV-sensitivity, mutation of Arg722 to Trp correlates with highly UV-sensitive TTD cell strains, and mutation of Arg683 to Trp correlates with XP-D. Alleles with mutation of Arg616 to Pro or with the combined mutation of Leu461 to Val and deletion of 716-730 are found in both XP-D and TTD cell strains. 39 refs., 2 figs., 3 tabs.

  1. Induction of resistance to alkylating agents in E. coli: the ada+ gene product serves both as a regulatory protein and as an enzyme for repair of mutagenic damage.

    PubMed Central

    Teo, I; Sedgwick, B; Demple, B; Li, B; Lindahl, T

    1984-01-01

    The expression of several inducible enzymes for repair of alkylated DNA in Escherichia coli is controlled by the ada+ gene. This regulatory gene has been cloned into a multicopy plasmid and shown to code for a 37-kd protein. Antibodies raised against homogeneous O6-methylguanine-DNA methyltransferase (the main repair activity for mutagenic damage in alkylated DNA) were found to cross-react with this 37-kd protein. Cell extracts from several independently derived ada mutants contain variable amounts of an altered 37-kd protein after an inducing alkylation treatment. In addition, an 18-kd protein identical with the previously isolated O6-methyl-guanine-DNA methyltransferase has been identified as a product of the ada+ gene. The smaller polypeptide is derived from the 37-kd protein by proteolytic processing. Images Fig. 1. Fig. 2. Fig. 4. Fig. 5. Fig. 6. Fig. 7. Fig. 8. PMID:6092060

  2. Structure and expression of the human XPBC/ERCC-3 gene involved in DNA repair disorders xeroderma pigmentosum and Cockayne's syndrome.

    PubMed Central

    Weeda, G; Ma, L B; van Ham, R C; van der Eb, A J; Hoeijmakers, J H

    1991-01-01

    The human XPBC/ERCC-3 was cloned by virtue of its ability to correct the excision repair defect of UV-sensitive rodent mutants of complementation group 3. The gene appeared to be in addition implicated in the human, cancer prone repair disorder xeroderma pigmentosum group B, which is also associated with Cockayne's syndrome. Here we present the genomic architecture of the gene and its expression. The XPBC/ERCC-3 gene consists of at least 14 exons spread over approximately 45 kb. Notably, the donor splice site of the third exon contains a GC instead of the canonical GT dinucleotide. The promoter region, first exon and intron comprise a CpG island with several putative GC boxes. The promoter was confined to a region of 260 bp upstream of the presumed cap site and acts bidirectionally. Like the promoter of another excision repair gene, ERCC-1, it lacks classical promoter elements such as CAAT and TATA boxes, but it shares with ERCC-1 a hitherto unknown 12 nucleotide sequence element, preceding a polypyrimidine track. Despite the presence of (AU)-rich elements in the 3'-untranslated region, which are thought to be associated with short mRNA half-life actinomycin-D experiments indicate that the mRNA is very stable (t 1/2 greater than 3h). Southern blot analysis revealed the presence of XPBC/ERCC-3 cross-hybridizing fragments elsewhere in the genome, which may belong to a related gene. Images PMID:1956789

  3. DNA Damage Repair Genes Controlling Human Papillomavirus (HPV) Episome Levels under Conditions of Stability and Extreme Instability

    PubMed Central

    Edwards, Terri G.; Vidmar, Thomas J.; Koeller, Kevin; Bashkin, James K.; Fisher, Chris

    2013-01-01

    DNA damage response (DDR) genes and pathways controlling the stability of HPV episomal DNA are reported here. We set out to understand the mechanism by which a DNA-binding, N-methylpyrrole-imidazole hairpin polyamide (PA25) acts to cause the dramatic loss of HPV DNA from cells. Southern blots revealed that PA25 alters HPV episomes within 5 hours of treatment. Gene expression arrays identified numerous DDR genes that were specifically altered in HPV16 episome-containing cells (W12E) by PA25, but not in HPV-negative (C33A) cells or in cells with integrated HPV16 (SiHa). A siRNA screen of 240 DDR genes was then conducted to identify enhancers and repressors of PA25 activity. Serendipitously, the screen also identified many novel genes, such as TDP1 and TDP2, regulating normal HPV episome stability. MRN and 9-1-1 complexes emerged as important for PA25-mediated episome destruction and were selected for follow-up studies. Mre11, along with other homologous recombination and dsDNA break repair genes, was among the highly significant PA25 repressors. The Mre11 inhibitor Mirin was found to sensitize HPV episomes to PA25 resulting in a ∼5-fold reduction of the PA25 IC50. A novel assay that couples end-labeling of DNA to Q-PCR showed that PA25 causes strand breaks within HPV DNA, and that Mirin greatly enhances this activity. The 9-1-1 complex member Rad9, a representative PA25 enhancer, was transiently phosphorylated in response to PA25 treatment suggesting that it has a role in detecting and signaling episome damage by PA25 to the cell. These results establish that DNA-targeted compounds enter cells and specifically target the HPV episome. This action leads to the activation of numerous DDR pathways and the massive elimination of episomal DNA from cells. Our findings demonstrate that viral episomes can be targeted for elimination from cells by minor groove binding agents, and implicate DDR pathways as important mediators of this process. PMID:24098381

  4. Loss of genes for DNA recombination and repair in the reductive genome evolution of thioautotrophic symbionts of Calyptogena clams

    PubMed Central

    2011-01-01

    Background Two Calyptogena clam intracellular obligate symbionts, Ca. Vesicomyosocius okutanii (Vok; C. okutanii symbiont) and Ca. Ruthia magnifica (Rma; C. magnifica symbiont), have small genomes (1.02 and 1.16 Mb, respectively) with low G+C contents (31.6% and 34.0%, respectively) and are thought to be in an ongoing stage of reductive genome evolution (RGE). They lack recA and some genes for DNA repair, including mutY. The loss of recA and mutY is thought to contribute to the stabilization of their genome architectures and GC bias, respectively. To understand how these genes were lost from the symbiont genomes, we surveyed these genes in the genomes from 10 other Calyptogena clam symbionts using the polymerase chain reaction (PCR). Results Phylogenetic trees reconstructed using concatenated 16S and 23S rRNA gene sequences showed that the symbionts formed two clades, clade I (symbionts of C. kawamurai, C. laubieri, C. kilmeri, C. okutanii and C. soyoae) and clade II (those of C. pacifica, C. fausta, C. nautilei, C. stearnsii, C. magnifica, C. fossajaponica and C. phaseoliformis). recA was detected by PCR with consensus primers for recA in the symbiont of C. phaseoliformis. A detailed homology search revealed a remnant recA in the Rma genome. Using PCR with a newly designed primer set, intact recA or its remnant was detected in clade II symbionts. In clade I symbionts, the recA coding region was found to be mostly deleted. In the Rma genome, a pseudogene of mutY was found. Using PCR with newly designed primer sets, mutY was not found in clade I symbionts but was found in clade II symbionts. The G+C content of 16S and 23S rRNA genes in symbionts lacking mutY was significantly lower than in those with mutY. Conclusions The extant Calyptogena clam symbionts in clade II were shown to have recA and mutY or their remnants, while those in clade I did not. The present results indicate that the extant symbionts are losing these genes in RGE, and that the loss of mut

  5. Reduced XPC DNA repair gene mRNA levels in clinically normal parents of xeroderma pigmentosum patients.

    PubMed

    Khan, Sikandar G; Oh, Kyu-Seon; Shahlavi, Tala; Ueda, Takahiro; Busch, David B; Inui, Hiroki; Emmert, Steffen; Imoto, Kyoko; Muniz-Medina, Vanessa; Baker, Carl C; DiGiovanna, John J; Schmidt, Deborah; Khadavi, Arash; Metin, Ahmet; Gozukara, Engin; Slor, Hanoch; Sarasin, Alain; Kraemer, Kenneth H

    2006-01-01

    Xeroderma pigmentosum group C (XP-C) is a rare autosomal recessive disorder. Patients with two mutant alleles of the XPC DNA repair gene have sun sensitivity and a 1000-fold increase in skin cancers. Clinically normal parents of XP-C patients have one mutant allele and one normal allele. As a step toward evaluating cancer risk in these XPC heterozygotes we characterized cells from 16 XP families. We identified 15 causative mutations (5 frameshift, 6 nonsense and 4 splicing) in the XPC gene in cells from 16 XP probands. All had premature termination codons (PTC) and absence of normal XPC protein on western blotting. The cell lines from 26 parents were heterozygous for the same mutations. We employed a real-time quantitative reverse transcriptase-PCR assay as a rapid and sensitive method to measure XPC mRNA levels. The mean XPC mRNA levels in the cell lines from the XP-C probands were 24% (P<10(-7)) of that in 10 normal controls. This reduced XPC mRNA level in cells from XP-C patients was caused by the PTC that induces nonsense-mediated mRNA decay. The mean XPC mRNA levels in cell lines from the heterozygous XP-C carriers were intermediate (59%, P=10(-4)) between the values for the XP patients and the normal controls. This study demonstrates reduced XPC mRNA levels in XP-C patients and heterozygotes. Thus, XPC mRNA levels may be evaluated as a marker of cancer susceptibility in carriers of mutations in the XPC gene. PMID:16081512

  6. DNA repair gene ERCC1 polymorphisms and glioma susceptibility among Chinese population: a meta-analysis

    PubMed Central

    Jiang, Chunming; Shen, Fang; Du, Jianmin; Wang, Xiaohua; Su, Jin; Liu, Zhanli; Huang, Xianmei

    2015-01-01

    Background: Excision repair cross complementation group 1 (ERCC1) has been shown to be involved in the progression of glioma susceptibility. However, the results remain conflict. The aim of this study was to systematically review and evaluate the role of ERCC1 C118T and C8092A polymorphisms in glioma risk among Chinese population. Methods: Related case-control studies were searched in online electronic databases. Odds ratio (OR) with its 95% confidence interval (CI) were employed to calculate the extracted data. Results: Total seven articles were retrieved, including 4426 subjects (1926 were glioma patients and 2500 were matched controls). No significant heterogeneity was found between studies (I2=0%, P>0.01). Our results demonstrated that A allele and AA genotype of ERCC1 C8092A polymorphism have a positive association with increasing the risk of glioma in the fixed-effect model (A vs. C: OR=1.13, 95% CI=1.02-1.25, P=0.02; AA vs. CC: OR=1.29, 95% CI=1.04-1.61, P=0.02; AA vs. CA+CC: OR=1.25, 95% CI=1.01-1.55, P=0.04). However, no significant relationship was found between C118T variant and glioma susceptibility. Conclusions: Our results indicated that ERCC1 C8092A, not C118T polymorphism might be a biomarker for patients with glioma among Chinese population. Future studies with more ethnicities are needed to explore the precise association. PMID:26379816

  7. Injectable graphene oxide/hydrogel-based angiogenic gene delivery system for vasculogenesis and cardiac repair.

    PubMed

    Paul, Arghya; Hasan, Anwarul; Kindi, Hamood Al; Gaharwar, Akhilesh K; Rao, Vijayaraghava T S; Nikkhah, Mehdi; Shin, Su Ryon; Krafft, Dorothee; Dokmeci, Mehmet R; Shum-Tim, Dominique; Khademhosseini, Ali

    2014-08-26

    The objective of this study was to develop an injectable and biocompatible hydrogel which can efficiently deliver a nanocomplex of graphene oxide (GO) and vascular endothelial growth factor-165 (VEGF) pro-angiogenic gene for myocardial therapy. For the study, an efficient nonviral gene delivery system using polyethylenimine (PEI) functionalized GO nanosheets (fGO) complexed with DNAVEGF was formulated and incorporated in the low-modulus methacrylated gelatin (GelMA) hydrogel to promote controlled and localized gene therapy. It was hypothesized that the fGOVEGF/GelMA nanocomposite hydrogels can efficiently transfect myocardial tissues and induce favorable therapeutic effects without invoking cytotoxic effects. To evaluate this hypothesis, a rat model with acute myocardial infarction was used, and the therapeutic hydrogels were injected intramyocardially in the peri-infarct regions. The secreted VEGF from in vitro transfected cardiomyocytes demonstrated profound mitotic activities on endothelial cells. A significant increase in myocardial capillary density at the injected peri-infarct region and reduction in scar area were noted in the infarcted hearts with fGOVEGF/GelMA treatment compared to infarcted hearts treated with untreated sham, GelMA and DNAVEGF/GelMA groups. Furthermore, the fGOVEGF/GelMA group showed significantly higher (p < 0.05, n = 7) cardiac performance in echocardiography compared to other groups, 14 days postinjection. In addition, no significant differences were noticed between GO/GelMA and non-GO groups in the serum cytokine levels and quantitative PCR based inflammatory microRNA (miRNA) marker expressions at the injected sites. Collectively, the current findings suggest the feasibility of a combined hydrogel-based gene therapy system for ischemic heart diseases using nonviral hybrid complex of fGO and DNA. PMID:24988275

  8. Injectable Graphene Oxide/Hydrogel-Based Angiogenic Gene Delivery System for Vasculogenesis and Cardiac Repair

    PubMed Central

    2015-01-01

    The objective of this study was to develop an injectable and biocompatible hydrogel which can efficiently deliver a nanocomplex of graphene oxide (GO) and vascular endothelial growth factor-165 (VEGF) pro-angiogenic gene for myocardial therapy. For the study, an efficient nonviral gene delivery system using polyethylenimine (PEI) functionalized GO nanosheets (fGO) complexed with DNAVEGF was formulated and incorporated in the low-modulus methacrylated gelatin (GelMA) hydrogel to promote controlled and localized gene therapy. It was hypothesized that the fGOVEGF/GelMA nanocomposite hydrogels can efficiently transfect myocardial tissues and induce favorable therapeutic effects without invoking cytotoxic effects. To evaluate this hypothesis, a rat model with acute myocardial infarction was used, and the therapeutic hydrogels were injected intramyocardially in the peri-infarct regions. The secreted VEGF from in vitro transfected cardiomyocytes demonstrated profound mitotic activities on endothelial cells. A significant increase in myocardial capillary density at the injected peri-infarct region and reduction in scar area were noted in the infarcted hearts with fGOVEGF/GelMA treatment compared to infarcted hearts treated with untreated sham, GelMA and DNAVEGF/GelMA groups. Furthermore, the fGOVEGF/GelMA group showed significantly higher (p < 0.05, n = 7) cardiac performance in echocardiography compared to other groups, 14 days postinjection. In addition, no significant differences were noticed between GO/GelMA and non-GO groups in the serum cytokine levels and quantitative PCR based inflammatory microRNA (miRNA) marker expressions at the injected sites. Collectively, the current findings suggest the feasibility of a combined hydrogel-based gene therapy system for ischemic heart diseases using nonviral hybrid complex of fGO and DNA. PMID:24988275

  9. Effects of expression level of DNA repair-related genes involved in the NHEJ pathway on radiation-induced cognitive impairment

    PubMed Central

    Zhang, Li-Yuan; Chen, Lie-Song; Sun, Rui; JI, Sheng-Jun; Ding, Yan-Yan; Wu, Jia; Tian, Ye

    2013-01-01

    Cranial radiation therapy can induce cognitive decline. Impairments of hippocampal neurogenesis are thought to be a paramountly important mechanism underlying radiation-induced cognitive dysfunction. In the mature nervous system, DNA double-strand breaks (DSBs) are mainly repaired by non-homologous end-joining (NHEJ) pathways. It has been demonstrated that NHEJ deficiencies are associated with impaired neurogenesis. In our study, rats were randomly divided into five groups to be irradiated by single doses of 0 (control), 0 (anesthesia control), 2, 10, and 20 Gy, respectively. The cognitive function of the irradiated rats was measured by open field, Morris water maze and passive avoidance tests. Real-time PCR was also used to detect the expression level of DNA DSB repair-related genes involved in the NHEJ pathway, such as XRCC4, XRCC5and XRCC6, in the hippocampus. The influence of different radiation doses on cognitive function in rats was investigated. From the results of the behavior tests, we found that rats receiving 20 Gy irradiation revealed poorer learning and memory, while no significant loss of learning and memory existed in rats receiving irradiation from 0–10 Gy. The real-time PCR and Western blot results showed no significant difference in the expression level of DNA repair-related genes between the 10 and 20 Gy groups, which may help to explain the behavioral results, i.e. DNA damage caused by 0–10 Gy exposure was appropriately repaired, however, damage induced by 20 Gy exceeded the body's maximum DSB repair ability. Ionizing radiation-induced cognitive impairments depend on the radiation dose, and more directly on the body's own ability to repair DNA DSBs via the NHEJ pathway. PMID:23135157

  10. DNA Repair Gene Polymorphisms and Their Relation With DNA Damage, DNA Repair, and Total Antioxidant Capacity in Childhood Acute Lymphoblastic Leukemia Survivors.

    PubMed

    Dincer, Yildiz; Yüksel, Selin; Batar, Bahadir; Güven, Mehmet; Onaran, Ilhan; Celkan, Tiraje

    2015-07-01

    Oxidative stress and defective DNA repair are major contributory factors in the initiation and progression of carcinogenesis. Chemotherapy and radiotherapy cause oxidative DNA damage, consume antioxidant capacity, and impair DNA repair activity. These effects of chemotherapy and radiotherapy may be contributory factors in the development of secondary malignancy in cancer survivors. Basal, H2O2-induced, and postrepair DNA damage; urinary 8-hydroxydeoxyguanosine level as a marker of oxidatively damaged DNA; and serum total antioxidant capacity were measured; XPD Lys751Gln, XRCC1 Arg399Gln, and XRCC1 Arg194Trp polymorphisms were analyzed in childhood acute lymphoblastic leukemia (ALL) survivors. Basal and H2O2-induced DNA damage were found to be higher in the ALL survivor group versus the control group, however, there was no significant difference between the other parameters. No association was found between the examined parameters and polymorphisms of XPD 751 and XRCC1 399 and both the groups. XRCC1 194Trp allele was found to be associated with a low level of postrepair DNA damage in the ALL survivors. In conclusion, basal DNA damage and susceptibility to oxidation are high in childhood ALL survivors. This situation which may easily lead to occurrence of a secondary cancer does not seem to be a result of deficient DNA repair. PMID:24577548

  11. Interactive effects of ultraviolet-B radiation and pesticide exposure on DNA photo-adduct accumulation and expression of DNA damage and repair genes in Xenopus laevis embryos.

    PubMed

    Yu, Shuangying; Tang, Song; Mayer, Gregory D; Cobb, George P; Maul, Jonathan D

    2015-02-01

    Pesticide use and ultraviolet-B (UVB) radiation have both been suggested to adversely affect amphibians; however, little is known about their interactive effects. One potential adverse interaction could involve pesticide-induced dysregulation of DNA repair pathways, resulting in greater numbers of DNA photo-adducts from UVB exposure. In the present study, we investigated the interactive effects of UVB radiation and two common pesticides (endosulfan and α-cypermethrin) on induction of DNA photo-adducts and expression of DNA damage and repair related genes in African clawed frog (Xenopus laevis) embryos. We examined 13 genes that are, collectively, involved in stress defense, cell cycle arrest, nucleotide excision repair (NER), base excision repair, mismatch repair, DNA repair regulation, and apoptosis. We exposed X. laevis embryos to 0, 25, and 50 μg/L endosulfan or 0, 2.5, and 5.0 μg/L α-cypermethrin for 96 h, with environmentally relevant exposures of UVB radiation during the last 7 h of the 96 h exposure. We measured the amount of cyclobutane pyrimidine dimers (CPDs) and mRNA abundance of the 13 genes among treatments including control, pesticide only, UVB only, and UVB and pesticide co-exposures. Each of the co-exposure scenarios resulted in elevated CPD levels compared to UVB exposure alone, suggesting an inhibitory effect of endosulfan and α-cypermethrin on CPD repair. This is attributed to results indicating that α-cypermethrin and endosulfan reduced mRNA abundance of XPA and HR23B, respectively, to levels that may affect the initial recognition of DNA lesions. In contrast, both pesticides increased transcript abundance of CSA and MUTL. In addition, mRNA abundance of HSP70 and GADD45α were increased by endosulfan and mRNA abundance of XPG was increased by α-cypermethrin. XPC, HR23B, XPG, and GADD45α exhibited elevated mRNA concentrations whereas there was a reduction in MUTL transcript concentrations in UVB-alone treatments. It appeared that even

  12. A new gene involved in DNA double-strand break repair and V(D)J recombination is located on human chromosome 10p.

    PubMed

    Moshous, D; Li, L; Chasseval, R; Philippe, N; Jabado, N; Cowan, M J; Fischer, A; de Villartay, J P

    2000-03-01

    V(D)J recombination, accountable for the diversity of T cell receptor- and immunoglobulin-encoding genes, is initiated by a lymphoid-specific DNA double-strand break. The general DNA repair machinery is responsible for the resolution of this break. Any defect in one of the known components of the DNA repair/V(D)J recombination machinery (Ku70, Ku80, DNA-PKcs, XRCC4 and DNA ligase IV) leads to abortion of the V(D)J rearrangement process, early block in both T and B cell maturation, and ultimately to severe combined immune deficiency (SCID) in several animal models. A human SCID condition is also characterized by an absence of mature T and B lymphocytes, and is associated with an increase in sensitivity to DNA-damaging agents (RS-SCID). None of the above-mentioned genes are defective in these patients, arguing for the likelihood of the existence of yet another unknown component of the V(D)J recombination/DNA repair apparatus. Athabascan-speaking (SCIDA) Navajo and Apache Native Americans have a very high incidence of T(-)B(-)SCID. The SCIDA locus is highly linked with markers on chromosome 10p, although the exact molecular defect has not been recognized in these patients. We show here that cells with the SCIDA defect are impaired in the DNA repair phase of V(D)J recombination similarly to RS-SCID, precisely an absence of V(D)J coding joint formation. Moreover, genotyping analysis in several RS-SCID families corroborates a linkage of the RS-SCID locus to the SCIDA region on chromosome 10p. These results demonstrate the presence of a new essential DNA repair/V(D)J recombination gene in this region, the mutation of which causes RS-SCID in humans. PMID:10699181

  13. Xeroderma pigmentosum and the role of DNA repair in oncogenesis.

    PubMed

    Giannelli, F

    1978-01-01

    Biochemical and genetic information on xeroderma pigmentosum (XP) has been briefly reviewed. This indicates that 80 to 90 percent of all XP patients are defective in the excision repair of pyrimidine dimers and are unable to perform the first step of this process as shown, for example, by their inability to undergo the DNA superhelical changes which accompany the initiation of excision repair in normal cells. However, in spite of its apparent biochemical homogeneity, XP is genetically heterogeneous and many genes appear to be responsible for the function of the factor defective in XP. Ten to 20 percent of all XP patients (called XP-variants) are capable of "dimer excision repair" but have difficulties in replicating UV-damaged DNA. The defects of XP and XP-variant affect also the repair of DNA damage caused by a number of chemical mutagens and carcinogens. This has important theoretical and practical implications since it indicates, for example, that the repair systems defective in XP must have broad specificity and that even XP cells not exposed to the harmful effect of light may suffer from poor repair of DNA damage. With regard to cancer, two questions have been considered. Namely, does XP provide a valid general model for UV-carcinogenesis in man and does it show how DNA damage leads to malignant transformation? The first question was answered in the affirmative in view of some clinical but, mainly, of cell biological data indicating that normal and excision defective XP cells differ, more quantitatively than qualitatively, in their response to UV-light. With regard to the second question XP seems to provide some support for various theories on carcinogenesis and, DNA repair defects may favour actinic carcinogenesis in a complex, non-univocous manner. Possibly the most important lesson imparted by XP is that, in man, the stability of the genetic material is dependent on the function of repair systems whose failure may predispose to cancer. In addition, the

  14. [Predisposing factors and prevention of emergence agitation].

    PubMed

    Mizuno, Ju; Nakata, Yoshinori; Morita, Shigeho; Arita, Hideko; Hanaoka, Kazuo

    2011-04-01

    Agitation during the emergence from general anesthesia is a great post-operative problem that often injures the patients themselves and requires the medical staff to restrain and calm the patients. The predisposing factors for emergence agitation include anesthesia, operation, and patient. Sevoflurane anesthesia results in higher incidence of emergence agitation than halothane, because of the rapid emergence, and its effects on central nervous system inducing convulsion and post-operative behavioral changes. The otorhinolaryngologic and ophthalmologic surgeries, post-operative pain, young age, pre-operative anxiety, no past surgical history, and adjustment disorder of patients are risk factors for emergence agitation. The change from sevoflurane to propofol during anesthesia maintenance is a contributing factor to reduce incidence of emergence agitation. The medications including opioids, midazolam, alpha-2 agonists, ketamine, non-steroidal anti-inflammatory drugs, nitrous oxide, and propofol, and aggressive nerve block such as caudal epidural block for post-operative sedation and analgesia are effective to avoid incidence of emergence agitation. The calm emergence following general anesthesia would decrease the self-injuring behavior, and enhance the parent and caregiver satisfaction in general anesthesia and surgery. PMID:21520589

  15. Ultrasound-induced hyperthermia for the spatio-temporal control of gene expression in bone repair

    NASA Astrophysics Data System (ADS)

    Wilson, Christopher; Padilla, Frédéric; Zhang, Man; Vilaboa, Nuria; Kripfgans, Oliver; Fowlkes, Brian; Franceschi, Renny

    2012-10-01

    Spatial and temporal control over the expression of growth/differentiation factors is of great interest for regeneration of bone, but technologies capable of providing tight and active control over gene expression remain elusive. We propose the use of focused ultrasound for the targeted activation of heat shock-sensitive expression systems in engineered bone. We report in vitro results with cells that express firefly luciferase (fLuc) under the control of a heat shock protein promoter. Cells were embedded in fibrin scaffolds and exposed to focused ultrasound, using a custom 3.3MHz transducer (focal length 4", f-number 1.33", focal dimension 1.2mm lateral FWHM) in CW mode for 2-20 minutes at intensities ISPTA=120-440 W/cm2. The kinetics of ultrasound-mediated activation of the cells was compared with that of strictly thermal activation. Bioluminescence imaging revealed fLuc expression in an area ≥2.5mm in diameter at the position of the ultrasound focus, and the diameter and intensity of the signal increased with the amplitude of the acoustic energy. We also found that ultrasound activated fLuc expression with substantially shorter exposures than thermal activation. Our results demonstrate the potential for focused ultrasound to selectively activate the expression of a gene of interest in an engineered tissue and suggest that focused ultrasound activates the heat shock pathway by a combination of thermal and non-thermal mechanisms.

  16. DNA repair

    SciTech Connect

    Friedberg, E.C.; Hanawalt, P.C. )

    1988-01-01

    Topics covered in this book included: Eukaryote model systems for DNA repair study; Sensitive detection of DNA lesions and their repair; and Defined DNA sequence probes for analysis of mutagenesis and repair.

  17. Gene polymorphism of IL-6 and MMP-3 decreases passive range of motion after rotator cuff repair

    PubMed Central

    Ling, Yan; Peng, Cheng; Liu, Chenguang; Zhang, Na; Yue, Shouwei

    2015-01-01

    Objectives: Post-operative stiffness is common after rotator cuff repair, given the difference in susceptibility and severity, the genetic factors may be involved. Interleukin 6 (IL-6) and Matrix metalloproteinases 3 (MMP-3) were previous found as key cytokines in the pathologies of adhesive capsulitis. The present study aims to investigate whether variants within the IL-6 and MMP-3 gene contributed to post-operative stiffness in a Chinese Han population. Methods: A total of 188 patients diagnosed with rotator cuff tears treated with mini-open surgery were enrolled in this study, among which 87 patients were diagnosed as post-operative stiffness and the remaining 101 patients as controls. All subjects were genotyped for IL-6 and MMP-3 SNPs. Results: The rs1800796 of IL-6 and rs679620 of MMP-3 were found significantly associated with increased susceptibility and severity of post-operative stiffness. Conclusion: The rs1800796 SNP of IL-6 and rs650108 SNP of MMP-3 were associated with increased risk of post-operative stiffness susceptibility and severity. This finding can be used in guiding the rehabilitation procedure after rotator cuff surgery, in another word, those with the genetic susceptibility factors should receive a more radical rehabilitation procedure and those without the susceptibility factors can be more conservative. PMID:26191285

  18. Histone Sprocket Arginine Residues Are Important for Gene Expression, DNA Repair, and Cell Viability in Saccharomyces cerevisiae.

    PubMed

    Hodges, Amelia J; Gallegos, Isaura J; Laughery, Marian F; Meas, Rithy; Tran, Linh; Wyrick, John J

    2015-07-01

    A critical feature of the intermolecular contacts that bind DNA to the histone octamer is the series of histone arginine residues that insert into the DNA minor groove at each superhelical location where the minor groove faces the histone octamer. One of these "sprocket" arginine residues, histone H4 R45, significantly affects chromatin structure in vivo and is lethal when mutated to alanine or cysteine in Saccharomyces cerevisiae (budding yeast). However, the roles of the remaining sprocket arginine residues (H3 R63, H3 R83, H2A R43, H2B R36, H2A R78, H3 R49) in chromatin structure and other cellular processes have not been well characterized. We have genetically characterized mutations in each of these histone residues when introduced either singly or in combination to yeast cells. We find that pairs of arginine residues that bind DNA adjacent to the DNA exit/entry sites in the nucleosome are lethal in yeast when mutated in combination and cause a defect in histone occupancy. Furthermore, mutations in individual residues compromise repair of UV-induced DNA lesions and affect gene expression and cryptic transcription. This study reveals simple rules for how the location and structural mode of DNA binding influence the biological function of each histone sprocket arginine residue. PMID:25971662

  19. Nucleotide excision repair genes and risk of lung cancer among San Francisco Bay Area Latinos and African Americans.

    PubMed

    Chang, Jeffrey S; Wrensch, Margaret R; Hansen, Helen M; Sison, Jennette D; Aldrich, Melinda C; Quesenberry, Charles P; Seldin, Michael F; Kelsey, Karl T; Kittles, Rick A; Silva, Gabriel; Wiencke, John K

    2008-11-01

    Few studies on the association between nucleotide excision repair (NER) variants and lung cancer risk have included Latinos and African Americans. We examine variants in 6 NER genes (ERCC2, ERCC4, ERCC5, LIG1, RAD23B and XPC) in association with primary lung cancer risk among 113 Latino and 255 African American subjects newly diagnosed with primary lung cancer from 1998 to 2003 in the San Francisco Bay Area and 579 healthy controls (299 Latinos and 280 African Americans). Individual single nucleotide polymorphism and haplotype analyses, multifactor dimensionality reduction (MDR) and principal components analysis (PCA) were performed to assess the association between 6 genes in the NER pathway and lung cancer risk. Among Latinos, ERCC2 haplotype CGA (rs238406, rs11878644, rs6966) was associated with reduced lung cancer risk [odds ratio (OR) of 0.65 and 95% confidence interval (CI): 0.44-0.97], especially among nonsmokers (OR = 0.29; 95% CI: 0.12-0.67). From MDR analysis, in Latinos, smoking and 3 SNPs (ERCC2 rs171140, ERCC5 rs17655 and LIG1 rs20581) together had a prediction accuracy of 67.4% (p = 0.001) for lung cancer. Among African Americans, His/His genotype of ERCC5 His1104Asp (rs17655) was associated with increased lung cancer risk (OR = 1.78; 95% CI: 1.09-2.91), and LIG1 haplotype GGGAA (rs20581, rs156641, rs3730931, rs20579 and rs439132) was associated with reduced lung cancer risk (OR = 0.61; 95% CI: 0.42-0.88). Our study suggests different elements of the NER pathway may be important in the different ethnic groups resulting either from different linkage relationship, genetic backgrounds and/or exposure histories. PMID:18709642

  20. Repair of bone defect by using vascular bundle implantation combined with Runx II gene-transfected adipose-derived stem cells and a biodegradable matrix.

    PubMed

    Han, Dong; Li, Jianjun

    2013-06-01

    A large hurdle in orthopedics today is the difficulty of dealing with the non-union of fractured bones. We therefore evaluated the effects of runt-related transcription factor II (Runx II), a factor used to create gene-modified tissue-engineered bone, combined with vascular bundle implantation for repairing segmental bone defects. Adenovirus Runx II gene (Ad-Runx II)-modified rabbit adipose-derived stem cells (ADSCs) were seeded onto polylactic acid/polycaprolacton (PLA/PCL) scaffolds to construct gene-modified tissue-engineered bone. The following four methods were used for repair in rabbit radial-defect (1.5 cm long) models: gene-modified tissue-engineered bone with vascular bundle (Group A), gene-modified tissue-engineered bone (Group B), non-gene-modified tissue-engineered bone with vascular bundle (Group C), and PLA/PCL scaffolds only (Group D). X-ray, histological examination, biomechanics analysis, and micro-angiography were conducted 4, 8, and 12 weeks later to determine angiogenesis and osteogenesis. The volume and speed of production of newly formed bones in Group A were significantly superior to those in other groups, and de-novo vascular network circulation from the vessel bundle through newly formed bone tissue was observed, with the defect being completely repaired. Group B showed a slightly better effect in terms of speed and quality of bone formation than Group C, whereas the bone defect in Group D was replaced by fibrous tissue. The maximal anti-bending strength in Group A was significantly higher than that in the other groups. Runx II gene therapy combined with vascular bundle implantation thus displays excellent abilities for osteoinduction and vascularization and is a promising method for the treatment of bone non-union and defect. PMID:23604755

  1. Inhibition of DNA-repair genes Ercc1 and Mgmt enhances temozolomide efficacy in gliomas treatment: a pre-clinical study

    PubMed Central

    Boccard, Sandra G.; Marand, Sandie V.; Geraci, Sandra; Pycroft, Laurie; Berger, François R.; Pelletier, Laurent A.

    2015-01-01

    Gliomas are the most common primary brain tumors. To date, therapies do not allow curing patients, and glioblastomas (GBMs) are associated with remarkably poor prognosis. This situation is at least partly due to intrinsic or acquired resistance to treatment, especially to chemotherapy. In 2005, temozolomide (TMZ) has become the first chemotherapeutic drug validated for GBM. Nevertheless TMZ efficacy depends on Mgmt status. While the methylation of Mgmt promoter was considered so far as a prognostic marker, its targeting is becoming an effective therapeutic opportunity. Thus, arrival of both TMZ and Mgmt illustrated that considerable progress can still be realized by optimizing adjuvant chemotherapy. A part of this progress could be accomplished in the future by overcoming residual resistance. The aim of the present study was to investigate the involvement of a set of other DNA-repair genes in glioma resistance to temozolomide. We focused on DNA-repair genes located in the commonly deleted chromosomal region in oligodendroglioma (1p/19q) highly correlated with patient response to chemotherapy. We measured effects of inhibition of ten DNA-repair genes expression using siRNAs on astrocytoma cell response to cisplatin (CDDP) and TMZ. SiRNAs targeting ercc1, ercc2, mutyh, and pnkp significantly sensitized cells to chemotherapy, increasing cell death by up to 25%. In vivo we observed a decrease of subcutaneous glioma tumor growth after injection of siRNA in conjunction with absorption of TMZ. We demonstrated in this pre-clinical study that targeting of DNA-repair genes such as Ercc1 could be used as an adjuvant chemosensitization treatment, similarly to Mgmt inhibition. PMID:26336131

  2. Inhibition of DNA-repair genes Ercc1 and Mgmt enhances temozolomide efficacy in gliomas treatment: a pre-clinical study.

    PubMed

    Boccard, Sandra G; Marand, Sandie V; Geraci, Sandra; Pycroft, Laurie; Berger, François R; Pelletier, Laurent A

    2015-10-01

    Gliomas are the most common primary brain tumors. To date, therapies do not allow curing patients, and glioblastomas (GBMs) are associated with remarkably poor prognosis. This situation is at least partly due to intrinsic or acquired resistance to treatment, especially to chemotherapy. In 2005, temozolomide (TMZ) has become the first chemotherapeutic drug validated for GBM. Nevertheless TMZ efficacy depends on Mgmt status. While the methylation of Mgmt promoter was considered so far as a prognostic marker, its targeting is becoming an effective therapeutic opportunity. Thus, arrival of both TMZ and Mgmt illustrated that considerable progress can still be realized by optimizing adjuvant chemotherapy. A part of this progress could be accomplished in the future by overcoming residual resistance. The aim of the present study was to investigate the involvement of a set of other DNA-repair genes in glioma resistance to temozolomide. We focused on DNA-repair genes located in the commonly deleted chromosomal region in oligodendroglioma (1p/19q) highly correlated with patient response to chemotherapy. We measured effects of inhibition of ten DNA-repair genes expression using siRNAs on astrocytoma cell response to cisplatin (CDDP) and TMZ. SiRNAs targeting ercc1, ercc2, mutyh, and pnkp significantly sensitized cells to chemotherapy, increasing cell death by up to 25%. In vivo we observed a decrease of subcutaneous glioma tumor growth after injection of siRNA in conjunction with absorption of TMZ. We demonstrated in this pre-clinical study that targeting of DNA-repair genes such as Ercc1 could be used as an adjuvant chemosensitization treatment, similarly to Mgmt inhibition. PMID:26336131

  3. Association of nucleotide excision repair pathway gene polymorphisms with gastric cancer and atrophic gastritis risks

    PubMed Central

    Liu, Jingwei; Sun, Liping; Xu, Qian; Tu, Huakang; He, Caiyun; Xing, Chengzhong; Yuan, Yuan

    2016-01-01

    Polymorphisms of NER genes could change NER ability, thereby altering individual susceptibility to GC. We systematically analyzed 39 SNPs of 8 key genes of NER pathway in 2686 subjects including 898 gastric cancer (GC), 851 atrophic gastritis (AG) and 937 controls (CON) in northern Chinese. SNP genotyping were performed using Sequenom MassARRAY platform. The results demonstrated that DDB2 rs830083 GG genotype was significantly associated with increased GC risk compared with wildtype CC (OR=2.32, P = 6.62 × 10−9); XPC rs2607775 CG genotype conferred a 1.73 increased odds of GC risk than non-cancer subjects compared with wild-type CC (OR=1.73, P= 3.04 × 10−4). The combined detection of these two polymorphisms demonstrated even higher GC risk (OR=3.05). Haplotype analysis suggested that DDB2 rs2029298-rs326222-rs3781619-rs830083 GTAG haplotype was significantly associated with disease risk in each step of CON→AG→GC development (AG vs. CON: OR=2.88, P= 7.51 × 10−7; GC vs. AG: OR=2.90, P=5.68 × 10−15; GC vs. CON: OR=8.42, P=2.22 × 10−15); DDB2 GTAC haplotype was associated with reduced risk of GC compared with CON (OR=0.63, P= 8.31 × 10−12). XPC rs1870134-rs2228000- rs2228001-rs2470352-rs2607775 GCAAG haplotype conferred increased risk of GC compared with AG (OR=1.88, P= 6.98 × 10−4). XPA rs2808668 and drinking, DDB2 rs326222, rs3781619, rs830083 and smoking demonstrated significant interactions in AG; XPC rs2607775 had significant interaction with smoking in GC. In conclusion, NER pathway polymorphisms especially in “damage incision” step were significantly associated with GC risk and had interactions with environment factors. The detection of NER pathway polymorphisms such as DDB2 and XPC might be applied in the prediction of GC risk and personalized prevention in the future. NOVELTY & IMPACT STATEMENTS NER pathway polymorphisms especially in “damage incision” step were significantly associated with GC risk and had interactions with

  4. Association of nucleotide excision repair pathway gene polymorphisms with gastric cancer and atrophic gastritis risks.

    PubMed

    Liu, Jingwei; Sun, Liping; Xu, Qian; Tu, Huakang; He, Caiyun; Xing, Chengzhong; Yuan, Yuan

    2016-02-01

    Polymorphisms of NER genes could change NER ability, thereby altering individual susceptibility to GC. We systematically analyzed 39 SNPs of 8 key genes of NER pathway in 2686 subjects including 898 gastric cancer (GC), 851 atrophic gastritis (AG) and 937 controls (CON) in northern Chinese. SNP genotyping were performed using Sequenom MassARRAY platform. The results demonstrated that DDB2 rs830083 GG genotype was significantly associated with increased GC risk compared with wild-type CC (OR=2.32, P= 6.62 × 10-9); XPC rs2607775 CG genotype conferred a 1.73 increased odds of GC risk than non-cancer subjects compared with wild-type CC (OR=1.73, P= 3.04 × 10-4). The combined detection of these two polymorphisms demonstrated even higher GC risk (OR=3.05). Haplotype analysis suggested that DDB2 rs2029298-rs326222-rs3781619-rs830083 GTAG haplotype was significantly associated with disease risk in each step of CON→AG→GC development (AG vs. CON: OR=2.88, P= 7.51 × 10-7; GC vs. AG: OR=2.90, P=5.68 × 10-15; GC vs. CON: OR=8.42, P=2.22 × 10-15); DDB2 GTAC haplotype was associated with reduced risk of GC compared with CON (OR=0.63, P= 8.31 × 10-12). XPC rs1870134-rs2228000-rs2228001-rs2470352-rs2607775 GCAAG haplotype conferred increased risk of GC compared with AG (OR=1.88, P= 6.98 × 10-4). XPA rs2808668 and drinking, DDB2 rs326222, rs3781619, rs830083 and smoking demonstrated significant interactions in AG; XPC rs2607775 had significant interaction with smoking in GC. In conclusion, NER pathway polymorphisms especially in "damage incision" step were significantly associated with GC risk and had interactions with environment factors. The detection of NER pathway polymorphisms such as DDB2 and XPC might be applied in the prediction of GC risk and personalized prevention in the future. NER pathway polymorphisms especially in "damage incision" step were significantly associated with GC risk and had interactions with environment factors, which might be applied in the

  5. Genetic variability of genes involved in DNA repair influence treatment outcome in osteosarcoma.

    PubMed

    Wang, M J; Zhu, Y; Guo, X J; Tian, Z Z

    2015-01-01

    We conducted a perspective study to investigate the role of ERCC1 (rs11615), ERCC2 (rs13181 and rs1799793), ERCC4 (rs1800067), and ERCC5 (rs17655) in NER pathway in the prognosis of osteosarcoma patients. In total, 146 osteosarcoma patients were recruited between 2008 and 2013. ERCC1 rs11615, ERCC2 rs13181 and rs1799793, ERCC4 rs1800067, and ERCC5 rs17655 gene polymorphisms were assessed by polymerase chain reaction-restriction fragment length polymorphism assay. By multivariate Cox proportional hazards models, we found that carriers of ERCC1 rs11615 TT genotype showed significantly favorable survival compared to wide-type CC genotype, and the adjusted OR (95%CI) was 0.24 (0.08-0.96). Moreover, we found that subjects with ERCC2 rs1799793 AA genotype were associated with decreased hazards of death in multivariate analysis (HR = 0.22, 95%CI = 0.12-0.93). In conclusion, our results suggest that ERCC1 rs11615 and ERCC2 rs1799793 may be useful genetic prognostic markers for osteosarcoma in a Chinese population. PMID:26436406

  6. Polymorphisms in DNA Repair Gene XRCC3 and Susceptibility to Breast Cancer in Saudi Females

    PubMed Central

    Ali, Alaa Mohammed; AbdulKareem, Huda; Al Anazi, Mohammad; Reddy Parine, Narasimha; Shaik, Jilani Purusottapatnam; Alamri, Abdullah; Ali Khan Pathan, Akbar

    2016-01-01

    We investigated three common polymorphisms (SNPs) in the XRCC3 gene (rs861539, rs1799794, and rs1799796) in 143 Saudi females suffering from breast cancer (median age = 51.4 years) and 145 age matched normal healthy controls. DNA was extracted from whole blood and genotyping was conducted using PCR-RFLP. rs1799794 showed significant association, where AA and AA+AG occurred at a significantly higher frequency in the cancer patients compared to the control group (OR: 28.1; 95% CI: 3.76–21.12; χ2: 22.82; p < 0.0001). The G allele was protective and presented with a dominant model. The genotype and allele frequencies of rs861539 C>T and rs1799796 A>G did not show a significant difference when the results in the patients and controls were compared. However, the frequency of rs1799796 differed significantly in patients with different age of diagnosis, tumor grade, and ER and HER2 status. The wild type A allele occurred at a higher frequency in the ER− and HER2− group. Our results among Saudis suggest that some variations in XRCC3 may contribute to breast cancer susceptibility. In conclusion, the results obtained during this study suggest that rs1799794 in XRCC3 shows strong association with breast cancer development in Saudi females. PMID:26881229

  7. The function of the Periaxin gene during nerve repair in a model of CMT4F *

    PubMed Central

    Williams, Anna C; Brophy, Peter J

    2002-01-01

    Mutations in the Periaxin (PRX) gene are known to cause autosomal recessive demyelinating Charcot-Marie-Tooth (CMT4F) and Dejerine-Sottas disease. The pathogenesis of these diseases is not fully understood. However, progress is being made by studying both the periaxin-null mouse, a mouse model of the disease, and the protein–protein interactions of periaxin. L-periaxin is a constituent of the dystroglycan–dystrophin-related protein-2 complex linking the Schwann cell cytoskeleton to the extracellular matrix. Although periaxin-null mice myelinate normally, they develop a demyelinating peripheral neuropathy later in life. This suggests that periaxin is required for the stable maintenance of a normal myelin sheath. We carried out sciatic nerve crushes in 6-week-old periaxin-null mice, and, 6 weeks later, found that although the number of myelinated axons had returned to normal, the axon diameters remained smaller than in the contralateral uncrushed nerve. Not only do periaxin-null mice have more hypermyelinated axons than their wild-type counterparts but they also recapitulate this hypermyelination during regeneration. Therefore, periaxin-null mice can undergo peripheral nerve remyelination, but the regulation of peripheral myelin thickness is disrupted. PMID:12090399

  8. The yeast TFB1 and SSL1 genes, which encode subunits of transcription factor IIH, are required for nucleotide excision repair and RNA polymerase II transcription.

    PubMed Central

    Wang, Z; Buratowski, S; Svejstrup, J Q; Feaver, W J; Wu, X; Kornberg, R D; Donahue, T F; Friedberg, E C

    1995-01-01

    The essential TFB1 and SSL1 genes of the yeast Saccharomyces cerevisiae encode two subunits of the RNA polymerase II transcription factor TFIIH (factor b). Here we show that extracts of temperature-sensitive mutants carrying mutations in both genes (tfb1-101 and ssl1-1) are defective in nucleotide excision repair (NER) and RNA polymerase II transcription but are proficient for base excision repair. RNA polymerase II-dependent transcription at the CYC1 promoter was normal at permissive temperatures but defective in extracts preincubated at a restrictive temperature. In contrast, defective NER was observed at temperatures that are permissive for growth. Additionally, both mutants manifested increased sensitivity to UV radiation at permissive temperatures. The extent of this sensitivity was not increased in a tfb1-101 strain and was only slightly increased in a ssl1-1 strain at temperatures that are semipermissive for growth. Purified factor TFIIH complemented defective NER in both tfb1-101 and ssl1-1 mutant extracts. These results define TFB1 and SSL1 as bona fide NER genes and indicate that, as is the case with the yeast Rad3 and Ss12 (Rad25) proteins, Tfb1 and Ssl1 are required for both RNA polymerase II basal transcription and NER. Our results also suggest that the repair and transcription functions of Tfb1 and Ssl1 are separable. PMID:7891722

  9. In vivo effects of UV radiation on multiple endpoints and expression profiles of DNA repair and heat shock protein (Hsp) genes in the cycloid copepod Paracyclopina nana.

    PubMed

    Won, Eun-Ji; Han, Jeonghoon; Lee, Yeonjung; Kumar, K Suresh; Shin, Kyung-Hoon; Lee, Su-Jae; Park, Heum Gi; Lee, Jae-Seong

    2015-08-01

    To evaluate the effects of ultraviolet (UV) radiation on energy acquisition and consumption, the copepod Paracyclopina nana was irradiated with several doses (0-3kJ/m(2)) of UV. After UV radiation, we measured the re-brooding success, growth pattern of newly hatched nauplii, ingestion rate, and assimilation of diet. In addition, we checked the modulated patterns of DNA repair and heat shock protein (hsp) chaperoning genes of P. nana. UV-B radiation induced a significant reduction (7-87%) of the re-brooding rate of ovigerous females, indicating that UV-induced egg sac damage is closely correlated with a reduction in the hatching rate of UV-irradiated ovigerous female offspring. Using chlorophyll a and stable carbon isotope incubation experiments, we found a dose-dependent decrease (P<0.05) in food ingestion and the rate of assimilation to the body in response to UV radiation, implying that P. nana has an underlying ability to shift its balanced-energy status from growth and reproduction to DNA repair and adaptation. Also, expression of P. nana base excision repair (BER)-associated genes and hsp chaperoning genes was significantly increased in response to UV radiation in P. nana. These findings indicate that even 1kJ/m(2) of UV radiation induces a reduction in reproduction and growth patterns, alters the physiological balance and inhibits the ability to cope with UV-induced damage in P. nana. PMID:26001085

  10. The mismatch repair system (mutS, mutL and uvrD genes) in Pseudomonas aeruginosa: molecular characterization of naturally occurring mutants.

    PubMed

    Oliver, Antonio; Baquero, Fernando; Blázquez, Jesús

    2002-03-01

    We have recently described the presence of a high proportion of Pseudomonas aeruginosa isolates (20%) with an increased mutation frequency (mutators) in the lungs of cystic fibrosis (CF) patients. In four out of 11 independent P. aeruginosa strains, the high mutation frequency was found to be complemented with the wild-type mutS gene from P. aeruginosa PAO1. Here, we report the cloning and sequencing of two additional P. aeruginosa mismatch repair genes and the characterization, by complementation of deficient strains, of these two putative P. aeruginosa mismatch repair genes (mutL and uvrD). We also describe the alterations in the mutS, mutL and uvrD genes responsible for the mutator phenotype of hypermutable P. aeruginosa strains isolated from CF patients. Seven out of the 11 mutator strains were found to be defective in the MMR system (four mutS, two mutL and one uvrD). In four cases (three mutS and one mutL), the genes contained frameshift mutations. The fourth mutS strain showed a 3.3 kb insertion after the 10th nucleotide of the mutS gene, and a 54 nucleotide deletion between two eight nucleotide direct repeats. This deletion, involving domain II of MutS, was found to be the main one responsible for mutS inactivation. The second mutL strain presented a K310M mutation, equivalent to K307 in Escherichia coli MutL, a residue known to be essential for its ATPase activity. Finally, the uvrD strain had three amino acid substitutions within the conserved ATP binding site of the deduced UvrD polypeptide, showing defective mismatch repair activity. Interestingly, cells carrying this mutant allele exhibited a fully active UvrABC-mediated excision repair. The results shown here indicate that the putative P. aeruginosa mutS, mutL and uvrD genes are mutator genes and that their alteration results in a mutator phenotype. PMID:11952911

  11. Single-nucleotide polymorphisms in base excision repair, nucleotide excision repair, and double strand break genes as markers for response to radiotherapy in patients with Stage I to II head-and-neck cancer

    SciTech Connect

    Carles, Joan . E-mail: jcarles@imas.imim.es; Monzo, Mariano; Amat, Marta; Jansa, Sonia; Artells, Rosa; Navarro, Alfons; Foro, Palmira; Alameda, Francesc; Gayete, Angel; Gel, Bernat; Miguel, Maribel; Albanell, Joan; Fabregat, Xavier

    2006-11-15

    Purpose: Polymorphisms in DNA repair genes can influence response to radiotherapy. We analyzed single-nucleotide polymorphisms (SNP) in nine DNA repair genes in 108 patients with head-and-neck cancer (HNSCC) who had received radiotherapy only. Methods and Materials: From May 1993 to December 2004, patients with Stage I and II histopathologically confirmed HNSCC underwent radiotherapy. DNA was obtained from paraffin-embedded tissue, and SNP analysis was performed using a real-time polymerase chain reaction allelic discrimination TaqMan assay with minor modifications. Results: Patients were 101 men (93.5%) and 7 (6.5%) women, with a median age of 64 years (range, 40 to 89 years). Of the patients, 76 (70.4%) patients were Stage I and 32 (29.6%) were Stage II. The XPF/ERCC1 SNP at codon 259 and XPG/ERCC5 at codon 46 emerged as significant predictors of progression (p 0.00005 and 0.049, respectively) and survival (p = 0.0089 and 0.0066, respectively). Similarly, when variant alleles of XPF/ERCC1, XPG/ERCC5 and XPA were examined in combination, a greater number of variant alleles was associated with shorter time to progression (p = 0.0003) and survival (p 0.0002). Conclusions: Genetic polymorphisms in XPF/ERCC1, XPG/ERCC5, and XPA may significantly influence response to radiotherapy; large studies are warranted to confirm their role in HNSCC.

  12. Absence of Activation of DNA Repair Genes and Excellent Efficacy of Phosphaplatins against Human Ovarian Cancers: Implications To Treat Resistant Cancers.

    PubMed

    Bose, Rathindra N; Moghaddas, Shadi; Belkacemi, Louiza; Tripathi, Swarnendu; Adams, Nyssa R; Majmudar, Pooja; McCall, Kelly; Dezvareh, Homa; Nislow, Corey

    2015-11-12

    Phosphaplatins, platinum(II) and platinum(IV) complexes coordinated to a pyrophosphate moiety, exhibit excellent antitumor activities against a variety of cancers. To determine whether phosphaplatins trigger resistance to treatment by engaging DNA damage repair genes, a yeast genome-wide fitness assay was used. Treatment of yeast cells with pyrodach-2 (D2) or pyrodach-4 (D4) revealed no particular sensitivity to nucleotide excision repair, homologous recombination repair, or postreplication repair when compared with platin control compounds. Also, TNF receptor superfamily member 6 (FAS) protein was overexpressed in phosphaplatin-treated ovarian tumor cells, and platinum colocalized with FAS protein in lipid rafts. An overactivation of sphingomyelinase (ASMase) was noted in the treated cells, indicating participation of an extrinsic apoptotic mechanism due to increased ceramide release. Our results indicate that DNA is not the target of phosphaplatins and accordingly, that phosphaplatins might not cause resistance to treatment. Activation of ASMase and FAS along with the colocalization of platinum with FAS in lipid rafts support an extrinsic apoptotic signaling mechanism that is mediated by phosphaplatins. PMID:26455832

  13. Single-nucleotide polymorphisms of the Trypanosoma cruzi MSH2 gene support the existence of three phylogenetic lineages presenting differences in mismatch-repair efficiency.

    PubMed Central

    Augusto-Pinto, Luiz; Teixeira, Santuza M R; Pena, Sérgio D J; Machado, Carlos Renato

    2003-01-01

    We have identified single-nucleotide polymorphisms (SNPs) in the mismatch-repair gene TcMSH2 from Trypanosoma cruzi. Phylogenetic inferences based on the SNPs, confirmed by RFLP analysis of 32 strains, showed three distinct haplogroups, denominated A, B, and C. Haplogroups A and C presented strong identity with the previously described T. cruzi lineages I and II, respectively. A third haplogroup (B) was composed of strains presenting hybrid characteristics. All strains from a haplogroup encoded the same specific protein isoform, called, respectively, TcMHS2a, TcMHS2b, and TcMHS2c. The classification into haplogroups A, B, and C correlated with variation in the efficiency of mismatch repair in these cells. When microsatellite loci of strains representative of each haplogroup were analyzed after being cultured in the presence of hydrogen peroxide, new microsatellite alleles were definitely seen in haplogroups B and C, while no evidence of microsatellite instability was found in haplogroup A. Also, cells from haplogroups B and C were considerably more resistant to cisplatin treatment, a characteristic known to be conferred by deficiency of mismatch repair in eukaryotic cells. Altogether, our data suggest that strains belonging to haplogroups B and C may have decreased mismatch-repair ability when compared with strains assigned to the haplogroup A lineage. PMID:12750325

  14. XPD DNA nucleotide excision repair gene polymorphisms associated with DNA repair deficiency predict better treatment outcomes in secondary acute myeloid leukemia

    PubMed Central

    Kuptsova-Clarkson, Nataliya; Ambrosone, Christine B; Weiss, Joli; Baer, Maria R; Sucheston, Lara E; Zirpoli, Gary; Kopecky, Kenneth J; Ford, Laurie; Blanco, Javier; Wetzler, Meir; Moysich, Kirsten B

    2010-01-01

    Pharmacogenetic studies in DNA repair pathway have consistently demonstrated correlations between the XRCC1 Arg399Gln, XPD Lys751Gln and XPD Asp312Gln genotypes, previously associated with suboptimal DNA repair, and differential cancer treatment outcomes. We evaluated these polymorphisms and XPD haplotypes in adult de novo (n=214) and secondary (n=79) acute myeloid leukemia (AML) patients treated with cytarabine and anthracycline chemotherapy. Genotyping was performed by MALDI-TOF mass spectrometry. Logistic and proportional hazards regression models were used to evaluate relationships. Differential responses were observed in secondary, but not de novo, AML. Among secondary AML patients, the odds of achieving complete remission (CR) were higher for the XPD 312Asn/Asn (OR= 11.23; 95% CI, 2.23-56.63) and XPD 751Gln/Gln (OR= 7.07; 95% CI, 1.42-35.18) genotypes. The XPD diplotypes were coded as the combination of two of the following haplotypes: haplotype A=(Lys)751A/(Asp) 312G; B=(Gln)751C/(Asn)312A; C=(Lys)751A/(Asn)312A; and D=(Gln)751C/(Asp)312G. The BB diplotype was associated with CR attainment [OR=18.31; 95% CI: 2.08-283.57] and longer survival [HR=0.31; 95% CI: 0.14-0.73] compared to the referent AA diplotype. The XPD 751 CC, 312GA, 312AA genotypes and the XPD DC diplotype were also associated with longer overall survival (OS).Thus, XPD codon 312 and 751 variant genotypes and haplotypes containing at least one variant allele may predict better treatment responses. If validated, these findings could support stratification of chemotherapy in secondary AML. PMID:21394217

  15. DNA Repair Genes ERCC1 and BRCA1 Expression in Non-Small Cell Lung Cancer Chemotherapy Drug Resistance

    PubMed Central

    Wang, Shuai; Liu, Feng; Zhu, Jingyan; Chen, Peng; Liu, Hongxing; Liu, Qi; Han, Junqing

    2016-01-01

    Background Surgery combined with chemotherapy is an important therapy for non-small cell lung cancer (NSCLC). However, chemotherapy drug resistance seriously hinders the curative effect. Studies show that DNA repair genes ERCC1 and BRCA1 are associated with NSCLC chemotherapy, but their expression and mechanism in NSCLC chemotherapy drug-resistant cells has not been elucidated. Material/Methods NSCLC cell line A549 and drug resistance cell line A549/DDP were cultured. Real-time PCR and Western blot analyses were used to detect ERCC1 and BRCA1 mRNA expression. A549/DDP cells were randomly divided into 3 groups: the control group; the siRNA-negative control group (scramble group); and the siRNA ERCC1 and BRCA1siRNA transfection group. Real-time PCR and Western blot analyses were used to determine ERCC1 and BRCA1 mRNA and protein expression. MTT was used to detect cell proliferation activity. Caspase 3 activity was tested by use of a kit. Western blot analysis was performed to detect PI3K, AKT, phosphorylated PI3K, and phosphorylated AKT protein expression. Results ERCC1 and BRCA1 were overexpressed in A549/DDP compared with A549 (P<0.05). ERCC1 and BRCA1siRNA transfection can significantly reduce ERCC1 and BRCA1 mRNA and protein expression (P<0.05). Downregulating ERCC1 and BRCA1 expression obviously inhibited cell proliferation and increased caspase 3 activity (P<0.05). Downregulating ERCC1 and BRCA1 significantly decreased PI3K and AKT phosphorylation levels (P<0.05). Conclusions ERCC1 and BRCA1 were overexpressed in NSCLC drug-resistant cells, and they regulated lung cancer occurrence and development through the phosphorylating PI3K/AKT signaling pathway. PMID:27289442

  16. DNA Repair Genes ERCC1 and BRCA1 Expression in Non-Small Cell Lung Cancer Chemotherapy Drug Resistance.

    PubMed

    Wang, Shuai; Liu, Feng; Zhu, Jingyan; Chen, Peng; Liu, Hongxing; Liu, Qi; Han, Junqing

    2016-01-01

    BACKGROUND Surgery combined with chemotherapy is an important therapy for non-small cell lung cancer (NSCLC). However, chemotherapy drug resistance seriously hinders the curative effect. Studies show that DNA repair genes ERCC1 and BRCA1 are associated with NSCLC chemotherapy, but their expression and mechanism in NSCLC chemotherapy drug-resistant cells has not been elucidated. MATERIAL AND METHODS NSCLC cell line A549 and drug resistance cell line A549/DDP were cultured. Real-time PCR and Western blot analyses were used to detect ERCC1 and BRCA1 mRNA expression. A549/DDP cells were randomly divided into 3 groups: the control group; the siRNA-negative control group (scramble group); and the siRNA ERCC1 and BRCA1siRNA transfection group. Real-time PCR and Western blot analyses were used to determine ERCC1 and BRCA1 mRNA and protein expression. MTT was used to detect cell proliferation activity. Caspase 3 activity was tested by use of a kit. Western blot analysis was performed to detect PI3K, AKT, phosphorylated PI3K, and phosphorylated AKT protein expression. RESULTS ERCC1 and BRCA1 were overexpressed in A549/DDP compared with A549 (P<0.05). ERCC1 and BRCA1siRNA transfection can significantly reduce ERCC1 and BRCA1 mRNA and protein expression (P<0.05). Downregulating ERCC1 and BRCA1 expression obviously inhibited cell proliferation and increased caspase 3 activity (P<0.05). Downregulating ERCC1 and BRCA1 significantly decreased PI3K and AKT phosphorylation levels (P<0.05). CONCLUSIONS ERCC1 and BRCA1 were overexpressed in NSCLC drug-resistant cells, and they regulated lung cancer occurrence and development through the phosphorylating PI3K/AKT signaling pathway. PMID:27289442

  17. Post-transcriptional Repair of a Split Heat Shock Protein 90 Gene by mRNA trans-Splicing*♦

    PubMed Central

    Nageshan, Rishi Kumar; Roy, Nainita; Hehl, Adrian B.; Tatu, Utpal

    2011-01-01

    Heat shock protein 90 participates in diverse biological processes ranging from protein folding, cell cycle, signal transduction and development to evolution in all eukaryotes. It is also critically involved in regulating growth of protozoa such as Dictyostelium discoideum, Leishmania donovani, Plasmodium falciparum, Trypanosoma cruzi, and Trypanosoma evansi. Selective inhibition of Hsp90 has also been explored as an intervention strategy against important human diseases such as cancer, malaria, or trypanosomiasis. Giardia lamblia, a simple protozoan parasite of humans and animals, is an important cause of diarrheal disease with significant morbidity and some mortality in tropical countries. Here we show that the G. lamblia cytosolic hsp90 (glhsp90) is split in two similar sized fragments located 777 kb apart on the same scaffold. Intrigued by this unique arrangement, which appears to be specific for the Giardiinae, we have investigated the biosynthesis of GlHsp90. We used genome sequencing to confirm the split nature of the giardial hsp90. However, a specific antibody raised against the peptide detected a product with a mass of about 80 kDa, suggesting a post-transcriptional rescue of the genomic defect. We show evidence for the joining of the two independent Hsp90 transcripts in-trans to one long mature mRNA presumably by RNA splicing. The splicing junction carries hallmarks of classical cis-spliced introns, suggesting that the regular cis-splicing machinery may be sufficient for repair of the open reading frame. A complementary 26-nt sequence in the “intron” regions adjacent to the splice sites may assist in positioning the two pre-mRNAs for processing. This is the first example of post-transcriptional rescue of a split gene by trans-splicing. PMID:21209094

  18. Post-transcriptional repair of a split heat shock protein 90 gene by mRNA trans-splicing.

    PubMed

    Nageshan, Rishi Kumar; Roy, Nainita; Hehl, Adrian B; Tatu, Utpal

    2011-03-01

    Heat shock protein 90 participates in diverse biological processes ranging from protein folding, cell cycle, signal transduction and development to evolution in all eukaryotes. It is also critically involved in regulating growth of protozoa such as Dictyostelium discoideum, Leishmania donovani, Plasmodium falciparum, Trypanosoma cruzi, and Trypanosoma evansi. Selective inhibition of Hsp90 has also been explored as an intervention strategy against important human diseases such as cancer, malaria, or trypanosomiasis. Giardia lamblia, a simple protozoan parasite of humans and animals, is an important cause of diarrheal disease with significant morbidity and some mortality in tropical countries. Here we show that the G. lamblia cytosolic hsp90 (glhsp90) is split in two similar sized fragments located 777 kb apart on the same scaffold. Intrigued by this unique arrangement, which appears to be specific for the Giardiinae, we have investigated the biosynthesis of GlHsp90. We used genome sequencing to confirm the split nature of the giardial hsp90. However, a specific antibody raised against the peptide detected a product with a mass of about 80 kDa, suggesting a post-transcriptional rescue of the genomic defect. We show evidence for the joining of the two independent Hsp90 transcripts in-trans to one long mature mRNA presumably by RNA splicing. The splicing junction carries hallmarks of classical cis-spliced introns, suggesting that the regular cis-splicing machinery may be sufficient for repair of the open reading frame. A complementary 26-nt sequence in the "intron" regions adjacent to the splice sites may assist in positioning the two pre-mRNAs for processing. This is the first example of post-transcriptional rescue of a split gene by trans-splicing. PMID:21209094

  19. Sequence conservation of the rad21 Schizosaccharomyces pombe DNA double-strand break repair gene in human and mouse

    SciTech Connect

    McKay, M.J.; Troelstra, C.; Kanaar, R.

    1996-09-01

    The rad21 gene of Schizosaccharomyces pombe is involved in the repair of ionizing radiation-induced DNA double-strand breaks. The isolation of mouse and human putative homologs of rad21 is reported here. Alignment of the predicted amino acid sequence of Rad21 with the mammalian proteins showed that the similarity was distributed across the length of the proteins, with more highly conserved regions at both termini. The mHR21{sup sp} (mouse homolog of Rad21, S. pombe) and hHR21{sup sp} (human homolog of Rad21, S. pombe) predicted proteins were 96% identical, whereas the human and S. pombe proteins were 25% identical and 47% similar. RNA blot analysis showed that mHR21{sup sp} mRNA was abundant in all adult mouse tissues examined, with highest expression in testis and thymus. In addition to a 3.1-kb constitutive mRNA transcript, a 2.2-kb transcript was present at a high level in postmeiotic spermatids, while expression of the 3.1-kb mRNA in testis was confined to the meiotic compartment. hHR21{sup sp} mRNA was cell-cycle regulated in human cells, increasing in late S phase to a peak in G2 phase. The level of hHR21{sup sp} transcripts was not altered by exposure of normal diploid fibroblasts to 10 Gy ionizing radiation. In situ hybridization showed that mHR21{sup sp} resided on chromosome 15D3, whereas hHR21{sup sp} localized to the syntenic 8q24 region. Elevated expression of mHR21{sup sp} in testis and thymus supports a possible role for the rad21 mammalian homologs in V(D)J and meiotic recombination, respectively. Cell cycle regulation of rad21, retained from S. pombe to human, is consistent with a conservation of function between S. pombe and human rad21 genes. 62 refs., 8 figs., 1 tab.

  20. β-Cyclodextrin-Linked Polyethylenimine Nanoparticles Facilitate Gene Transfer and Enhance the Angiogenic Capacity of Mesenchymal Stem Cells for Wound Repair and Regeneration.

    PubMed

    Peng, Li-Hua; Wei, Wei; Shan, Ying-Hui; Zhang, Tian-Yuan; Zhang, Chen-Zhen; Wu, Jia-He; Yu, Lian; Lin, Jun; Liang, Wen-Quan; Khang, Gilson; Gao, Jian-Qing

    2015-04-01

    Repair of deep wounds by cell transplantation strongly depends on angiogenesis and on the regeneration of skin and appendages. In this study, plasmid DNA encoding vascular endothelial growth factor-165 (VEGF-165) was transduced into bone-marrow mesenchymal stem cells (MSCs) using a nonviral vector, β-cyclodextrin-linked polyethylenimine, to enhance angiogenic capacity. The effects of MSCs administered by intradermal injection or transplantation on wound closure were compared in a full-thickness excision wound model. The results showed that the MSC-seeded sponge had significantly stronger acceleration in wound closure than the MSC injection. The effects on wound repair and regeneration of transplanted MSCs and pDNA-VEGF1 65-transfected MSCs (TMSCs) with gelatin/β-tricalcium phosphate scaffold were also investigated. Compared with MSC transplantation, TMSC transplantation showed higher efficacy in stimulating wound closure, promoting dermal collagen synthesis and regulating the deposition of newly formed collagen. In addition, the angiogenic capacity of the TMSCs was higher than that of the MSCs. The results indicate that the nonviral genetic engineering of the MSCs is a promising strategy to enhance the angiogenic capacity of MSCs for wound repair and angiogenesis. Functional gene-activated MSCs may be used as cost-effective and accessible seed cells for skin tissue engineering and as novel carriers for wound gene therapy. PMID:26310074

  1. Differences in Expression of Key DNA Damage Repair Genes after Epigenetic-Induced BRCAness Dictate Synthetic Lethality with PARP1 Inhibition.

    PubMed

    Wiegmans, Adrian P; Yap, Pei-Yi; Ward, Ambber; Lim, Yi Chieh; Khanna, Kum Kum

    2015-10-01

    The triple-negative breast cancer (TNBC) subtype represents a cancer that is highly aggressive with poor patient outcome. Current preclinical success has been gained through synthetic lethality, targeting genome instability with PARP inhibition in breast cancer cells that harbor silencing of the homologous recombination (HR) pathway. Histone deacetylase inhibitors (HDACi) are a class of drugs that mediate epigenetic changes in expression of HR pathway genes. Here, we compare the activity of the pan-HDAC inhibitor suberoylanilide hydroxamic acid (SAHA), the class I/IIa HDAC inhibitor valproic acid (VPA), and the HDAC1/2-specific inhibitor romidepsin (ROMI) for their capability to regulate DNA damage repair gene expression and in sensitizing TNBC to PARPi. We found that two of the HDACis tested, SAHA and ROMI, but not VPA, indeed inhibit HR repair and that RAD51, BARD1, and FANCD2 represent key proteins whose inhibition is required for HDACi-mediated therapy with PARP inhibition in TNBC. We also observed that restoration of BRCA1 function stabilizes the genome compared with mutant BRCA1 that results in enhanced polyploid population after combination treatment with HDACi and PARPi. Furthermore, we found that overexpression of the key HR protein RAD51 represents a mechanism for this resistance, promoting aberrant repair and the enhanced polyploidy observed. These findings highlight the key components of HR in guiding synthetic lethality with PARP inhibition and support the rationale for utilizing the novel combination of HDACi and PARPi against TNBC in the clinical setting. PMID:26294743

  2. Inducible Apoe Gene Repair in Hypomorphic ApoE Mice Deficient in the LDL Receptor Promotes Atheroma Stabilization with a Human-like Lipoprotein Profile

    PubMed Central

    Eberlé, Delphine; Luk, Fu Sang; Kim, Roy Y.; Olivas, Victor R.; Kumar, Nikit; Posada, Jessica M.; Li, Kang; Gaudreault, Nathalie; Rapp, Joseph H.; Raffai, Robert L.

    2013-01-01

    Objective To study atherosclerosis regression in mice following plasma lipid reduction to moderately elevated apolipoprotein B (apoB)-lipoprotein levels. Approach and Results Chow-fed hypomorphic Apoe mice deficient in LDL receptor expression (Apoeh/hLdlr−/−Mx1-cre mice) develop hyperlipidemia and atherosclerosis. These mice were studied before and after inducible cre-mediated Apoe gene repair. By 1 week, induced mice displayed a 2-fold reduction in plasma cholesterol and triglyceride levels and a decrease in the non-HDL:HDL-cholesterol ratio from 87%:13% to 60%:40%. This halted atherosclerotic lesion growth and promoted macrophage loss and accumulation of thick collagen fibers for up to 8 weeks. Concomitantly, blood Ly-6Chi monocytes were decreased by 2-fold but lesional macrophage apoptosis was unchanged. The expression of several genes involved in extra-cellular matrix remodeling and cell migration were changed in lesional macrophages 1 week after Apoe gene repair. However, mRNA levels of numerous genes involved in cholesterol efflux and inflammation were not significantly changed at this time point. Conclusions Restoring apoE expression in Apoeh/hLdlr−/−Mx1-cre mice resulted in lesion stabilization in the context of a human-like ratio of non-HDL:HDL-cholesterol. Our data suggest that macrophage loss derived in part from reduced blood Ly-6Chi monocytes levels and genetic reprogramming of lesional macrophages. PMID:23788760

  3. Analysis of a human DNA excision repair gene involved in group A xeroderma pigmentosum and containing a zinc-finger domain.

    PubMed

    Tanaka, K; Miura, N; Satokata, I; Miyamoto, I; Yoshida, M C; Satoh, Y; Kondo, S; Yasui, A; Okayama, H; Okada, Y

    1990-11-01

    Xeroderma pigmentosum (XP) is an autosomal recessive disease, characterized by a high incidence of sunlight-induced skin cancer. Cells from people with this condition are hypersensitive to ultraviolet because of a defect in DNA repair. There are nine genetic complementation groups of XP, groups A-H and a variant. We have cloned the mouse DNA repair gene that complements the defect of group A, the XPAC gene. Here we report molecular cloning of human and mouse XPAC complementary DNAs. Expression of XPAC cDNA confers ultraviolet-resistance on several group A cell lines, but not on lines of other XP groups. Almost all group A lines tested showed abnormality or absence of XPAC messenger RNAs. These results indicate that a defective XPAC gene causes group A XP. The human and mouse XPAC genes are located on chromosome 9q34.1 and chromosome 4C2, respectively. Human XPAC cDNA encodes a protein of 273 amino acids with a zinc-finger motif. PMID:2234061

  4. Programmed Genetic Instability: A Tumor-Permissive Mechanism for Maintaining the Evolvability of Higher Species through Methylation-Dependent Mutation of DNA Repair Genes in the Male Germ Line

    PubMed Central

    Zhao, Yongzhong

    2008-01-01

    Tumor suppressor genes are classified by their somatic behavior either as caretakers (CTs) that maintain DNA integrity or as gatekeepers (GKs) that regulate cell survival, but the germ line role of these disease-related gene subgroups may differ. To test this hypothesis, we have used genomic data mining to compare the features of human CTs (n = 38), GKs (n = 36), DNA repair genes (n = 165), apoptosis genes (n = 622), and their orthologs. This analysis reveals that repair genes are numerically less common than apoptosis genes in the genomes of multicellular organisms (P < 0.01), whereas CT orthologs are commoner than GK orthologs in unicellular organisms (P < 0.05). Gene targeting data show that CTs are less essential than GKs for survival of multicellular organisms (P < 0.0005) and that CT knockouts often permit offspring viability at the cost of male sterility. Patterns of human familial oncogenic mutations confirm that isolated CT loss is commoner than is isolated GK loss (P < 0.00001). In sexually reproducing species, CTs appear subject to less efficient purifying selection (i.e., higher Ka/Ks) than GKs (P = 0.000003); the faster evolution of CTs seems likely to be mediated by gene methylation and reduced transcription-coupled repair, based on differences in dinucleotide patterns (P = 0.001). These data suggest that germ line CT/repair gene function is relatively dispensable for survival, and imply that milder (e.g., epimutational) male prezygotic repair defects could enhance sperm variation—and hence environmental adaptation and speciation—while sparing fertility. We submit that CTs and repair genes are general targets for epigenetically initiated adaptive evolution, and propose a model in which human cancers arise in part as an evolutionarily programmed side effect of age- and damage-inducible genetic instability affecting both somatic and germ line lineages. PMID:18535014

  5. The set1Delta mutation unveils a novel signaling pathway relayed by the Rad53-dependent hyperphosphorylation of replication protein A that leads to transcriptional activation of repair genes.

    PubMed

    Schramke, V; Neecke, H; Brevet, V; Corda, Y; Lucchini, G; Longhese, M P; Gilson, E; Géli, V

    2001-07-15

    SET domain proteins are present in chromosomal proteins involved in epigenetic control of transcription. The yeast SET domain protein Set1p regulates chromatin structure, DNA repair, and telomeric functions. We investigated the mechanism by which the absence of Set1p increases DNA repair capacities of checkpoint mutants. We show that deletion of SET1 induces a response relayed by the signaling kinase Rad53p that leads to the MEC1/TEL1-independent hyperphosphorylation of replication protein A middle subunit (Rfa2p). Consequently, the binding of Rfa2p to upstream repressing sequences (URS) of repair genes is decreased, thereby leading to their derepression. Our results correlate the set1Delta-dependent phosphorylation of Rfa2p with the transcriptional induction of repair genes. Moreover, we show that the deletion of the amino-terminal region of Rfa2p suppresses the sensitivity to ultraviolet radiation of a mec3Delta checkpoint mutant, abolishes the URS-mediated repression, and increases the expression of repair genes. This work provides an additional link for the role of Rfa2p in the regulation of the repair capacity of the cell and reveals a role for the phosphorylation of Rfa2p and unveils unsuspected connections between chromatin, signaling pathways, telomeres, and DNA repair. PMID:11459833

  6. Human creatine kinase genes on chromosomes 15 and 19, and proximity of the gene for the muscle form to the genes for apolipoprotein C2 and excision repair.

    PubMed Central

    Stallings, R L; Olson, E; Strauss, A W; Thompson, L H; Bachinski, L L; Siciliano, M J

    1988-01-01

    The human chromosomal assignments of genes of the creatine kinase (CK) family--loci for brain (CKBB), muscle (CKMM), and mitochondrial (CKMT) forms--were studied by Southern filter hybridization analysis of DNAs isolated from a human x rodent somatic cell hybrid clone panel. Probes for the 3'-noncoding sequences of human CKBB and CKMM hybridized concordantly only to DNAs from somatic cell hybrids containing chromosomes 14 and 19, respectively. Thus the earlier assignment of the gene coding for the CKBB isozyme to chromosome 14 was confirmed by molecular means, as was the provisional assignment of CKMM to the long arm of chromosome 19. A probe containing canine sequences for CKMM cross-hybridized with human sequences on chromosomes 14 and 19, a result consistent with the assignments of CKBB and CKMM. A probe containing human sequences for CKMT enabled the provisional assignment of CKMT to human chromosome 15. Independent hybrids with portions of the long arm of chromosome 19 missing indicated the order of genes on the long arm of chromosome 19 as being cen-GPI-(TGFB, CYP1)-[CKMM, (APOC2-ERCC1)]-(CGB, FTL). The unexpectedly more distal location of APOC2 among the genes on the long arm--and APOC2's close association with CKMM--is discussed with respect to the close linkage relationship of APOC2 to myotonic muscular dystrophy. Images Figure 1 PMID:3400641

  7. Mismatch repair genes Mlh1 and Mlh3 modify CAG instability in Huntington's disease mice: genome-wide and candidate approaches.

    PubMed

    Pinto, Ricardo Mouro; Dragileva, Ella; Kirby, Andrew; Lloret, Alejandro; Lopez, Edith; St Claire, Jason; Panigrahi, Gagan B; Hou, Caixia; Holloway, Kim; Gillis, Tammy; Guide, Jolene R; Cohen, Paula E; Li, Guo-Min; Pearson, Christopher E; Daly, Mark J; Wheeler, Vanessa C

    2013-10-01

    The Huntington's disease gene (HTT) CAG repeat mutation undergoes somatic expansion that correlates with pathogenesis. Modifiers of somatic expansion may therefore provide routes for therapies targeting the underlying mutation, an approach that is likely applicable to other trinucleotide repeat diseases. Huntington's disease Hdh(Q111) mice exhibit higher levels of somatic HTT CAG expansion on a C57BL/6 genetic background (B6.Hdh(Q111) ) than on a 129 background (129.Hdh(Q111) ). Linkage mapping in (B6x129).Hdh(Q111) F2 intercross animals identified a single quantitative trait locus underlying the strain-specific difference in expansion in the striatum, implicating mismatch repair (MMR) gene Mlh1 as the most likely candidate modifier. Crossing B6.Hdh(Q111) mice onto an Mlh1 null background demonstrated that Mlh1 is essential for somatic CAG expansions and that it is an enhancer of nuclear huntingtin accumulation in striatal neurons. Hdh(Q111) somatic expansion was also abolished in mice deficient in the Mlh3 gene, implicating MutLγ (MLH1-MLH3) complex as a key driver of somatic expansion. Strikingly, Mlh1 and Mlh3 genes encoding MMR effector proteins were as critical to somatic expansion as Msh2 and Msh3 genes encoding DNA mismatch recognition complex MutSβ (MSH2-MSH3). The Mlh1 locus is highly polymorphic between B6 and 129 strains. While we were unable to detect any difference in base-base mismatch or short slipped-repeat repair activity between B6 and 129 MLH1 variants, repair efficiency was MLH1 dose-dependent. MLH1 mRNA and protein levels were significantly decreased in 129 mice compared to B6 mice, consistent with a dose-sensitive MLH1-dependent DNA repair mechanism underlying the somatic expansion difference between these strains. Together, these data identify Mlh1 and Mlh3 as novel critical genetic modifiers of HTT CAG instability, point to Mlh1 genetic variation as the likely source of the instability difference in B6 and 129 strains and suggest that MLH1

  8. Polymorphisms of the DNA repair genes XRCC1, XRCC3, XPD, interaction with environmental exposures, and bladder cancer risk in a case-control study in northern Italy.

    PubMed

    Shen, Min; Hung, Rayjean J; Brennan, Paul; Malaveille, Christian; Donato, Francesco; Placidi, Donatella; Carta, Angela; Hautefeuille, Agnes; Boffetta, Paolo; Porru, Stefano

    2003-11-01

    Tobacco smoking and occupational exposures are the main known risk factors for bladder cancer, causing direct and indirect damage to DNA. Repair of DNA damage is under genetic control, and DNA repair genes may play a key role in maintaining genome integrity and preventing cancer development. Polymorphisms in DNA repair genes resulting in variation of DNA repair efficiency may therefore be associated with bladder cancer risk. A hospital-based case-control study was conducted in Brescia, Italy, to assess the relationship between polymorphisms in DNA repair genes XRCC1 (Arg(399)Gln), XRCC3 (Thr(241)Met), and XPD (Lys(751)Gln) and bladder cancer risk. A total of 201 male incident bladder cancer cases and 214 male controls with urological nonneoplastic diseases were recruited and frequency-matched on age, period, and hospital of recruitment. Detailed information was collected using a semistructured questionnaire on demographic, dietary, environmental, and occupational factors. Genotypes were determined by PCR-RFLP analysis. The XRCC3 codon 241 variant genotype exhibited a protective effect against bladder cancer [odds ratio (OR), 0.63; 95% confidence interval (CI), 0.42-0.93], which was prominent among heavy smokers (OR, 0.49; 95% CI, 0.28-0.88) but not among never and light smokers. No overall impact of the XRCC1 codon 399 polymorphism was found (OR, 0.86; 95% CI, 0.59-1.28), but a protective influence of the homozygous variant was suggested among heavy smokers (OR, 0.38; 95% CI, 0.14-1.02). XPD polymorphisms did not show an association with bladder cancer (OR, 0.92; 95% CI, 0.62-1.37). There was no statistical evidence of an interaction between these three genetic polymorphisms and either tobacco smoking or occupational exposure to polycyclic aromatic hydrocarbons and aromatic amines. The XRCC3 codon 241 polymorphism had an overall protective effect against bladder cancer that was most apparent among heavy smokers. Similarly, the XRCC1 codon 399 polymorphism also had

  9. International congress on DNA damage and repair: Book of abstracts

    SciTech Connect

    Not Available

    1987-01-01

    This document contains the abstracts of 105 papers presented at the Congress. Topics covered include the Escherichia coli nucleotide excision repair system, DNA repair in malignant transformations, defective DNA repair, and gene regulation. (TEM)

  10. BRCA1/BARD1 orthologs required for DNA repair in Caenorhabditis elegans.

    PubMed

    Boulton, Simon J; Martin, Julie S; Polanowska, Jolanta; Hill, David E; Gartner, Anton; Vidal, Marc

    2004-01-01

    Inherited germline mutations in the tumor suppressor gene BRCA1 predispose individuals to early onset breast and ovarian cancer. BRCA1 together with its structurally related partner BARD1 is required for homologous recombination and DNA double-strand break repair, but how they perform these functions remains elusive. As part of a comprehensive search for DNA repair genes in C. elegans, we identified a BARD1 ortholog. In protein interaction screens, Ce-BRD-1 was found to interact with components of the sumoylation pathway, the TACC domain protein TAC-1, and most importantly, a homolog of mammalian BRCA1. We show that animals depleted for either Ce-brc-1 or Ce-brd-1 display similar abnormalities, including a high incidence of males, elevated levels of p53-dependent germ cell death before and after irradiation, and impaired progeny survival and chromosome fragmentation after irradiation. Furthermore, depletion of ubc-9 and tac-1 leads to radiation sensitivity and a high incidence of males, respectively, potentially linking these genes to the C. elegans BRCA1 pathway. Our findings support a shared role for Ce-BRC-1 and Ce-BRD-1 in C. elegans DNA repair processes, and this role will permit studies of the BRCA1 pathway in an organism amenable to rapid genetic and biochemical analysis. PMID:14711411

  11. Clubfoot repair

    MedlinePlus

    ... release; Talipes equinovarus - repair; Tibialis anterior tendon transfer Images Clubfoot repair - series References Kelly DM. Congenital Anomalies ... provided herein should not be used during any medical emergency or for the diagnosis or treatment of ...

  12. Zinc finger transcription factor CASZ1 interacts with histones, DNA repair proteins and recruits NuRD complex to regulate gene transcription.

    PubMed

    Liu, Zhihui; Lam, Norris; Thiele, Carol J

    2015-09-29

    The zinc finger transcription factor CASZ1 has been found to control neural fate-determination in flies, regulate murine and frog cardiac development, control murine retinal cell progenitor expansion and function as a tumor suppressor gene in humans. However, the molecular mechanism by which CASZ1 regulates gene transcription to exert these diverse biological functions has not been described. Here we identify co-factors that are recruited by CASZ1b to regulate gene transcription using co-immunoprecipitation (co-IP) and mass spectrometry assays. We find that CASZ1b binds to the nucleosome remodeling and histone deacetylase (NuRD) complex, histones and DNA repair proteins. Mutagenesis of the CASZ1b protein assay demonstrates that the N-terminus of CASZ1b is required for NuRD binding, and a poly(ADP-ribose) binding motif in the CASZ1b protein is required for histone H3 and DNA repair proteins binding. The N-terminus of CASZ1b fused to an artificial DNA-binding domain (GAL4DBD) causes a significant repression of transcription (5xUAS-luciferase assay), which could be blocked by treatment with an HDAC inhibitor. Realtime PCR results show that the transcriptional activity of CASZ1b mutants that abrogate NuRD or histone H3/DNA binding is significantly decreased. This indicates a model in which CASZ1b binds to chromatin and recruits NuRD complexes to orchestrate epigenetic-mediated transcriptional programs. PMID:26296975

  13. EGFR exon 19 in-frame deletion and polymorphisms of DNA repair genes in never-smoking female lung adenocarcinoma patients.

    PubMed

    Yang, Shi-Yi; Yang, Tsung-Ying; Li, Yao-Jen; Chen, Kun-Chieh; Liao, Kuo-Meng; Hsu, Kuo-Hsuan; Tsai, Chi-Ren; Chen, Chih-Yi; Hsu, Chung-Ping; Hsia, Jiun-Yi; Chuang, Cheng-Yen; Tsai, Ying-Huang; Chen, Kuan-Yu; Huang, Ming-Shyan; Su, Wu-Chou; Chen, Yuh-Min; Hsiung, Chao A; Shen, Chen-Yang; Chang, Gee-Chen; Yang, Pan-Chyr; Chen, Chien-Jen

    2013-01-15

    We explored potential associations between genetic polymorphisms in genes related to DNA repair and detoxification metabolism and epidermal growth factor receptor (EGFR) mutations in a cohort of 410 never-smoking patients with lung adenocarcinoma. Multivariate-adjusted odds ratios (aORs) and corresponding 95% confidence intervals (CI) of EGFR mutation status in association with the genotypes of DNA repair and detoxification metabolism genes were evaluated using logistic regression analysis. We found an association between in-frame deletion in EGFR exon 19 and a single nucleotide polymorphism (SNP) rs1800566C/T located in NQO1 (aOR, 2.2 with 95% CI, 1.0-4.8) in female never-smokers. The SNP rs744154C/G in ERCC4 was also associated with the EGFR exon 19 in-frame deletion both in never-smokers (aOR, 1.7 with 95% CI, 1.0-3.0) and female never-smokers (aOR, 1.9 with 95% CI, 1.0-3.6). Although the association was marginally significant in multivariate logistic regression analysis, the A/A genotype of rs1047840 in EXO1 was associated with a 7.6-fold increase in the occurrence of the EGFR exon 19 in-frame deletion in female never-smokers. Moreover, risk alleles in NQO1, ERCC4 and EXO1 were associated with an increasing aOR of the EGFR exon 19 in-frame deletion both in never-smokers (p = 0.007 for trend) and female never-smokers (p = 0.002 for trend). Our findings suggest that the in-frame deletion in EGFR exon 19 is associated with polymorphisms in DNA repair and detoxification metabolism genes in never-smoking lung adenocarcinoma patients, especially in females. PMID:22573488

  14. Decreased expression of DNA repair genes (XRCC1, ERCC1, ERCC2, and ERCC4) in squamous intraepithelial lesion and invasive squamous cell carcinoma of the cervix.

    PubMed

    Bajpai, Deepti; Banerjee, Ayan; Pathak, Sujata; Jain, Sunesh K; Singh, Neeta

    2013-05-01

    Reduced DNA repair might affect the risk of progression from infection with carcinogenic human papillomavirus (HPV), the etiologic agent for cervical cancer (CC), to persistent HPV infection, and hence to cervical pre-cancer and cancer. We assessed the variation in baseline expression of base excision repair gene XRCC1 and three nucleotide excision repair genes ERCC1, ERCC2, and ERCC4 and the risk of developing cervical cancer. A hospital-based case-control study was designed with 50 invasive cervical cancer patients, 40 squamous intraepithelial lesions (SIL) patients and 85 controls subjects. RT-qPCR and Western blotting was used to quantitate in vitro the mRNA and protein levels in fresh CC, SIL and normal cervix tissue. The levels of XRCC1, ERCC2, ERCC4, and ERCC1 transcripts and their respective proteins were lower in cervical cancer and SILs as compared to controls (p ≤ 0.001, 0.001, 0.001, and 0.025, respectively). In multivariate logistic regression analysis (adjusting for parity, age at first child birth, use of oral contraceptives, smoking status), low expression of XRCC1, ERCC2, ERCC4, and ERCC1 was associated with a significant increased risk for CC and SIL. Our results suggest that individuals whose expression of XRCC1, ERCC4, ERCC2, and ERCC1 are reduced may be at a higher risk of developing SIL which eventually leads to invasive cervical carcinoma. Moreover, independently also the reduced expression of these genes can directly lead to cervical cancer progression. PMID:23435956

  15. Association between Genetic Polymorphisms of DNA Repair Genes and Chromosomal Damage for 1,3-Butadiene-Exposed Workers in a Matched Study in China

    PubMed Central

    Xiang, Menglong; Sun, Lei; Dong, Xiaomei; Yang, Huan; Liu, Wen-bin; Zhou, Niya; Han, Xue; Zhou, Ziyuan; Cui, Zhihong; Liu, Jing-yi; Cao, Jia; Ao, Lin

    2015-01-01

    The aim of the study was to examine the association between polymorphisms of DNA repair genes and chromosomal damage of 1,3-butadiene- (BD-) exposed workers. The study was conducted in 45 pairs of occupationally exposed workers in a BD product workshop and matched control workers in an administrative office and a circulatory water workshop in China. Newly developed biomarkers (micronuclei, MNi; nucleoplasmic bridges, NPBs; nuclear buds, NBUDs) in the cytokinesis-blocked micronucleus (CBMN) cytome assay were adopted to detect chromosomal damage. PCR and PCR-restriction fragment length polymorphism (RFLP) are adopted to analyze polymorphisms of DNA repair genes, such as X-ray repair cross-complementing Group 1 (XRCC1), O6-methylguanine-DNA methyltransferase (MGMT), poly (adenosine diphosphate-ribose) polymerases (ADPRT), and apurinic/apyrimidinic endonucleases (APE1). The BD-exposed workers exhibited increased frequencies of MNi and NPBs when compared to subjects in the control group. The results also show that the BD-exposed workers carrying XRCC1 diplotypes TCGA-CCGG (4.25 ± 2.06‰) (FR = 2.10, 95% CI: 1.03–4.28) and TCGG-TCGA (5.80 ± 3.56‰) (FR = 2.75, 95% CI: 0.76–2.65) had statistically higher NBUD frequencies than those who carried diplotype TCGG-TCGG (1.89 ± 1.27‰). Our study suggests that polymorphisms of XRCC1 gene may influence chromosomal damage in BD-exposed workers. PMID:26339595

  16. Radio-adaptive response of base excision repair genes and proteins in human peripheral blood mononuclear cells exposed to gamma radiation.

    PubMed

    Toprani, Sneh M; Das, Birajalaxmi

    2015-09-01

    Radio-adaptive response is a mechanism whereby a low-dose exposure (priming dose) induces resistance to a higher dose (challenging dose) thus significantly reducing its detrimental effects. Radiation-induced DNA damage gets repaired through various DNA repair pathways in human cells depending upon the type of lesion. The base excision repair (BER) pathway repairs radiation-induced base damage, abasic sites and single-strand breaks in cellular DNA. In the present study, an attempt has been made to investigate the involvement of BER genes and proteins in the radio-adaptive response in human resting peripheral blood mononuclear cells (PBMC). Venous blood samples were collected from 20 randomly selected healthy male individuals with written informed consent. PBMC were isolated and irradiated at a priming dose of 0.1 Gy followed 4h later with a challenging dose of 2.0 Gy (primed cells). Quantitation of DNA damage was done using the alkaline comet assay immediately and expression profile of BER genes and proteins were studied 30 min after the challenging dose using real-time quantitative polymerase chain reaction and western blot, respectively. The overall result showed significant (P ≤ 0.05) reduction of DNA damage in terms of percentage of DNA in tail (%T) with a priming dose of 0.1 Gy followed by a challenging dose of 2.0 Gy after 4 h. Twelve individuals showed significant (P ≤ 0.05) reduction in %T whereas eight individuals showed marginal reduction in DNA damage that was not statistically significant. However, at the transcriptional level, BER genes such as APE1, FEN1 and LIGASE1 showed significant (P ≤ 0.05) up-regulation in both groups. Significant (P ≤ 0.05) up-regulation was also observed at the protein level for OGG1, APE1, MBD4, FEN1 and LIGASE1 in primed cells. Up-regulation of some BER genes and proteins such as APE1, FEN1 and LIGASE1 in primed cells of resting PBMC is suggestive of active involvement of the BER pathway in radio-adaptive response

  17. Oxidative stress and DNA repair and detoxification gene expression in adolescents exposed to heavy metals living in the Milazzo-Valle del Mela area (Sicily, Italy)

    PubMed Central

    Pizzino, Gabriele; Bitto, Alessandra; Interdonato, Monica; Galfo, Federica; Irrera, Natasha; Mecchio, Anna; Pallio, Giovanni; Ramistella, Vincenzo; Luca, Filippo De; Minutoli, Letteria; Squadrito, Francesco; Altavilla, Domenica

    2014-01-01

    Background The area of Milazzo-Valle del Mela (Sicily, Italy) is considered at high risk of environmental crisis by regional authorities. Objective To measure oxidative-stress, DNA repair and detoxification genes in school children living near the industrial area and in age-matched controls. Methods The parent study was a biomonitoring investigation evaluating heavy metal urine levels in 226 children aged 12–14 years, living in the high risk area, and in 29 age-matched controls living 45 km far from the industrial site. In the present study 67 exposed adolescents and 29 controls were included. Samples were analyzed for urinary 8-hydroxydeoxyguanosine (8OHdG) levels, and gene expression of OGG1 (DNA repair gene), NQO1, ST13, and MT1A (detoxifying genes). Results Urinary cadmium was higher (p = 0.0004) in exposed [geometric mean, 0.46 µg/L; 25th–75th percentile: 0.3–0.56] than in control adolescents [geometric mean, 0.26 µg/L; 25th–75th percentile: 0.2–0.3]. Chromium was also significantly elevated in exposed [geometric mean, 1.52 µg/L; 25th–75th percentile: 1.19–1.93] compared with controls [geometric mean, 1.25 µg/L; 25th–75th percentile: 1.05–1.48; p = 0.02]. Urinary 8-OHdG concentration was greater in exposed than in controls (71.49 vs 61.87 µg/L, p = 0.02), and it was correlated with cadmium levels (r = 0.46, p < 0.0001), and with the combined exposure index (r = 0.43, p < 0.0001). Moreover, cadmium levels showed a robust correlation with OGG1 and MT1A gene expression levels (r = 0.44, p < 0.0001; r = 0.39, p < 0.0001, respectively). Finally, OGG1 and MT1A were over-expressed in adolescents from Milazzo-Valle del Mela area compared with controls (p = 0.0004; p < 0.0001, respectively). Conclusions Continuous exposure at relatively low concentrations of heavy metals is associated with increased oxidative DNA damage and impaired expression of DNA repair and detoxification genes in adolescents. PMID:24936443

  18. The role of base excision repair genes OGG1, APN1 and APN2 in benzo[a]pyrene-7,8-dione induced p53 mutagenesis

    PubMed Central

    Abedin, Zahidur; Louis-Juste, Melissa; Stangl, Melissa; Field, Jeffrey

    2014-01-01

    Lung cancer is primarily caused by exposure to tobacco smoke. Tobacco smoke contains numerous carcinogens, including Polycyclic Aromatic Hydrocarbons (PAH). The most common PAH studied is benzo[a]pyrene (B[a]P). B[a]P is metabolically activated through multiple routes, one of which is catalyzed by aldo-keto reductase (AKR) to B[a]P-7,8-dione (BPQ). BPQ undergoes a futile redox cycle in the presence of NADPH to generate reactive oxygen species (ROS). ROS, in turn, damages DNA. Studies with a yeast p53 mutagenesis system found that the generation of ROS by PAH o-quinones may contribute to lung carcinogenesis because of similarities between the patterns (types of mutations) and spectra (location of mutations) and those seen in lung cancer. The patterns were dominated by G to T transversions, and the spectra in the experimental system have mutations at lung cancer hotspots. To address repair mechanisms that are responsible for BPQ induced damage we observed the effect of mutating two DNA repair genes OGG1 and APE1 (APN1 in yeast) and tested them in a yeast reporter system for p53 mutagenesis. There was an increase in both the mutant frequency and the number of G:C/T:A transversions in p53 treated with BPQ in ogg1 yeast but not in apn1 yeast. Knocking out APN2 increased mutagenesis in the apn1 cells. In addition, we did not find a strand bias on p53 treated with BPQ in ogg1 yeast. These studies suggest that Ogg1 is involved in repairing the oxidative damage caused by BPQ, Apn1 and Apn2 have redundant functions and that the stand bias seen in lung cancer may not be due to impaired repair of oxidative lesions. PMID:23117049

  19. Detecting disease-predisposing variants: The haplotype method

    SciTech Connect

    Valdes, A.M.; Thomson, G.

    1997-03-01

    For many HLA-associated diseases, multiple alleles - and, in some cases, multiple loci - have been suggested as the causative agents. The haplotype method for identifying disease-predisposing amino acids in a genetic region is a stratification analysis. We show that, for each haplotype combination containing all the amino acid sites involved in the disease process, the relative frequencies of amino acid variants at sites not involved in disease but in linkage disequilibrium with the disease-predisposing sites are expected to be the same in patients and controls. The haplotype method is robust to mode of inheritance and penetrance of the disease and can be used to determine unequivocally whether all amino acid sites involved in the disease have not been identified. Using a resampling technique, we developed a statistical test that takes account of the nonindependence of the sites sampled. Further, when multiple sites in the genetic region are involved in disease, the test statistic gives a closer fit to the null expectation when some - compared with none - of the true predisposing factors are included in the haplotype analysis. Although the haplotype method cannot distinguish between very highly correlated sites in one population, ethnic comparisons may help identify the true predisposing factors. The haplotype method was applied to insulin-dependent diabetes mellitus (IDDM) HLA class II DQA1-DQB1 data from Caucasian, African, and Japanese populations. Our results indicate that the combination DQA1 No. 52 (Arg predisposing) DQB1 No. 57 (Asp protective), which has been proposed as an important IDDM agent, does not include all the predisposing elements. With rheumatoid arthritis HLA class H DRB1 data, the results were consistent with the shared-epitope hypothesis. 35 refs., 2 figs., 6 tabs.

  20. Apigenin, a bioactive flavonoid from Lycopodium clavatum, stimulates nucleotide excision repair genes to protect skin keratinocytes from ultraviolet B-induced reactive oxygen species and DNA damage.

    PubMed

    Das, Sreemanti; Das, Jayeeta; Paul, Avijit; Samadder, Asmita; Khuda-Bukhsh, Anisur Rahman

    2013-10-01

    In this study, we examined the antioxidative and the DNA protective potentials of apigenin, a flavonoid polyphenol isolated from Lycopodium clavatum, in both in-vitro (HaCaT skin keratinocytes) and in-vivo (mice) models against UV-B radiation. We used DAPI staining in UV-B-irradiated HaCaT skin keratinocytes pre-treated with and without apigenin to assess DNA damage. We also used a flow-cytometric analysis in mice exposed to UV-B radiation with or without topical application of apigenin to assess, through a comet assay, chromosomal aberrations and quanta from reactive oxygen species (ROS) generation. Data from the stability curves for the Gibb's free energy determined from a melting-temperature profile study indicated that apigenin increased the stability of calf thymus DNA. Immunofluorescence studies revealed that apigenin caused a reduction in the number of cyclobutane pyrimidine dimers (CPDs) after 24 h, the time at which the nucleotide excision repair (NER) genes were activated. Thus, apigenin accelerated reversal of UV-B-induced CPDs through up-regulation of NER genes, removal of cyclobutane rings, inhibition of ROS generation, and down-regulation of NF-κB and MAPK, thereby revealing the precise mechanism of DNA repair. PMID:24139463

  1. The Cytosolic Iron-Sulfur Cluster Assembly Protein MMS19 Regulates Transcriptional Gene Silencing, DNA Repair, and Flowering Time in Arabidopsis.

    PubMed

    Han, Yong-Feng; Huang, Huan-Wei; Li, Lin; Cai, Tao; Chen, She; He, Xin-Jian

    2015-01-01

    MMS19 is an essential component of the cytoplasmic iron-sulfur (Fe-S) cluster assembly complex in fungi and mammals; the mms19 null mutant alleles are lethal. Our study demonstrates that MMS19/MET18 in Arabidopsis thaliana interacts with the cytoplasmic Fe-S cluster assembly complex but is not an essential component of the complex. We find that MMS19 also interacts with the catalytic subunits of DNA polymerases, which have been demonstrated to be involved in transcriptional gene silencing (TGS), DNA repair, and flowering time regulation. Our results indicate that MMS19 has a similar biological function, suggesting a functional link between MMS19 and DNA polymerases. In the mms19 null mutant, the assembly of Fe-S clusters on the catalytic subunit of DNA polymerase α is reduced but not blocked, which is consistent with the viability of the mutant. Our study suggests that MMS19 assists the assembly of Fe-S clusters on DNA polymerases in the cytosol, thereby facilitating transcriptional gene silencing, DNA repair, and flowering time control. PMID:26053632

  2. Targeting DNA repair pathways for cancer treatment: what's new?

    PubMed Central

    Kelley, Mark R; Logsdon, Derek; Fishel, Melissa L

    2014-01-01

    Disruptions in DNA repair pathways predispose cells to accumulating DNA damage. A growing body of evidence indicates that tumors accumulate progressively more mutations in DNA repair proteins as cancers progress. DNA repair mechanisms greatly affect the response to cytotoxic treatments, so understanding those mechanisms and finding ways to turn dysregulated repair processes against themselves to induce tumor death is the goal of all DNA repair inhibition efforts. Inhibition may be direct or indirect. This burgeoning field of research is replete with promise and challenge, as more intricacies of each repair pathway are discovered. In an era of increasing concern about healthcare costs, use of DNA repair inhibitors can prove to be highly effective stewardship of R&D resources and patient expenses. PMID:24947262

  3. Germline duplication of ATG2B and GSKIP predisposes to familial myeloid malignancies.

    PubMed

    Saliba, Joseph; Saint-Martin, Cécile; Di Stefano, Antonio; Lenglet, Gaëlle; Marty, Caroline; Keren, Boris; Pasquier, Florence; Valle, Véronique Della; Secardin, Lise; Leroy, Gwendoline; Mahfoudhi, Emna; Grosjean, Sarah; Droin, Nathalie; Diop, M'boyba; Dessen, Philippe; Charrier, Sabine; Palazzo, Alberta; Merlevede, Jane; Meniane, Jean-Côme; Delaunay-Darivon, Christine; Fuseau, Pascal; Isnard, Françoise; Casadevall, Nicole; Solary, Eric; Debili, Najet; Bernard, Olivier A; Raslova, Hana; Najman, Albert; Vainchenker, William; Bellanné-Chantelot, Christine; Plo, Isabelle

    2015-10-01

    No major predisposition gene for familial myeloproliferative neoplasms (MPN) has been identified. Here we demonstrate that the autosomal dominant transmission of a 700-kb duplication in four genetically related families predisposes to myeloid malignancies, including MPN, frequently progressing to leukemia. Using induced pluripotent stem cells and primary cells, we demonstrate that overexpression of ATG2B and GSKIP enhances hematopoietic progenitor differentiation, including of megakaryocytes, by increasing progenitor sensitivity to thrombopoietin (TPO). ATG2B and GSKIP cooperate with acquired JAK2, MPL and CALR mutations during MPN development. Thus, the germline duplication may change the fitness of cells harboring signaling pathway mutations and increases the probability of disease development. PMID:26280900

  4. Developmental defects and male sterility in mice lacking the ubiquitin-like DNA repair gene mHR23B.

    PubMed

    Ng, Jessica M Y; Vrieling, Harry; Sugasawa, Kaoru; Ooms, Marja P; Grootegoed, J Anton; Vreeburg, Jan T M; Visser, Pim; Beems, Rudolph B; Gorgels, Theo G M F; Hanaoka, Fumio; Hoeijmakers, Jan H J; van der Horst, Gijsbertus T J

    2002-02-01

    mHR23B encodes one of the two mammalian homologs of Saccharomyces cerevisiae RAD23, a ubiquitin-like fusion protein involved in nucleotide excision repair (NER). Part of mHR23B is complexed with the XPC protein, and this heterodimer functions as the main damage detector and initiator of global genome NER. While XPC defects exist in humans and mice, mutations for mHR23A and mHR23B are not known. Here, we present a mouse model for mHR23B. Unlike XPC-deficient cells, mHR23B(-/-) mouse embryonic fibroblasts are not UV sensitive and retain the repair characteristics of wild-type cells. In agreement with the results of in vitro repair studies, this indicates that mHR23A can functionally replace mHR23B in NER. Unexpectedly, mHR23B(-/-) mice show impaired embryonic development and a high rate (90%) of intrauterine or neonatal death. Surviving animals display a variety of abnormalities, including retarded growth, facial dysmorphology, and male sterility. Such abnormalities are not observed in XPC and other NER-deficient mouse mutants and point to a separate function of mHR23B in development. This function may involve regulation of protein stability via the ubiquitin/proteasome pathway and is not or only in part compensated for by mHR23A. PMID:11809813

  5. Inter-individual variation in nucleotide excision repair pathway is modulated by non-synonymous polymorphisms in ERCC4 and MBD4 genes.

    PubMed

    Allione, Alessandra; Guarrera, Simonetta; Russo, Alessia; Ricceri, Fulvio; Purohit, Rituraj; Pagnani, Andrea; Rosa, Fabio; Polidoro, Silvia; Voglino, Floriana; Matullo, Giuseppe

    2013-01-01

    Inter-individual differences in DNA repair capacity (DRC) may lead to genome instability and, consequently, modulate individual cancer risk. Among the different DNA repair pathways, nucleotide excision repair (NER) is one of the most versatile, as it can eliminate a wide range of helix-distorting DNA lesions caused by ultraviolet light irradiation and chemical mutagens. We performed a genotype-phenotype correlation study in 122 healthy subjects in order to assess if any associations exist between phenotypic profiles of NER and DNA repair gene single nucleotide polymorphisms (SNPs). Individuals were genotyped for 768 SNPs with a custom Illumina Golden Gate Assay, and peripheral blood mononuclear cells (PBMCs) of the same subjects were tested for a NER comet assay to measure DRC after challenging cells by benzo(a)pyrene diolepoxide (BPDE). We observed a large inter-individual variability of NER capacity, with women showing a statistically significant lower DRC (mean ± SD: 6.68 ± 4.76; p = 0.004) than men (mean ± SD: 8.89 ± 5.20). Moreover, DRC was significantly lower in individuals carrying a variant allele for the ERCC4 rs1800124 non-synonymous SNP (nsSNP) (p = 0.006) and significantly higher in subjects with the variant allele of MBD4 rs2005618 SNP (p = 0.008), in linkage disequilibrium (r(2) = 0.908) with rs10342 nsSNP. Traditional in silico docking approaches on protein-DNA and protein-protein interaction showed that Gly875 variant in ERCC4 (rs1800124) decreases the DNA-protein interaction and that Ser273 and Thr273 variants in MBD4 (rs10342) indicate complete loss of protein-DNA interactions. Our results showed that NER inter-individual capacity can be modulated by cross-talk activity involving nsSNPs in ERCC4 and MBD4 genes, and they suggested to better investigate SNP effect on cancer risk and response to chemo- and radiotherapies. PMID:24004570

  6. Nucleotide sequence of the Salmonella typhimurium mutS gene required for mismatch repair: homology of MutS and HexA of Streptococcus pneumoniae.

    PubMed Central

    Haber, L T; Pang, P P; Sobell, D I; Mankovich, J A; Walker, G C

    1988-01-01

    The mutS gene product of Escherichia coli and Salmonella typhimurium is one of at least four proteins required for methyl-directed mismatch repair in these organisms. A functionally similar repair system in Streptococcus pneumoniae requires the hex genes. We have sequenced the S. typhimurium mutS gene, showing that it encodes a 96-kilodalton protein. Amino-terminal amino acid sequencing of purified S. typhimurium MutS protein confirmed the initial portion of the deduced amino acid sequence. The S. typhimurium MutS protein is homologous to the S. pneumoniae HexA protein, suggesting that they arose from a common ancestor before the gram-negative and gram-positive bacteria diverged. Overall, approximately 36% of the amino acids of the two proteins are identical when the sequences are optimally aligned, including regions of stronger homology which are of particular interest. One such region is close to the amino terminus. Another, located closer to the carboxy terminus, includes homology to a consensus sequence thought to be diagnostic of nucleotide-binding sites. A third one, adjacent to the second, is homologous to the consensus sequence for the helix-turn-helix motif found in many DNA-binding proteins. We found that the S. typhimurium MutS protein can substitute for the E. coli MutS protein in vitro as it can in vivo, but we have not yet been able to demonstrate a similar in vitro complementation by the S. pneumoniae HexA protein. PMID:3275609

  7. Polymorphisms in base excision repair genes as colorectal cancer risk factors and modifiers of the effect of diets high in red meat

    PubMed Central

    Brevik, Asgeir; Joshi, Amit D.; Corral, Román; Onland-Moret, N. Charlotte; Siegmund, Kimberly D.; Le Marchand, Loïc; Baron, John A.; Martinez, Maria Elena; Haile, Robert W.; Ahnen, Dennis J.; Sandler, Robert S.; Lance, Peter; Stern, Mariana C.

    2010-01-01

    Background A diet high in red meat is an established colorectal cancer (CRC) risk factor. Carcinogens generated during meat cooking have been implicated as causal agents, and can induce oxidative DNA damage, which elicits repair by the base excision repair (BER) pathway. Methods Using a family-based study we investigated the role of polymorphisms in four BER genes (APEX1 Gln51His, Asp148Glu; OGG1 Ser236Cys; PARP Val742Ala; XRCC1 Arg194Trp, Arg280His, Arg399Gln) as potential CRC risk factors and modifiers of the association between high-red meat or poultry diets and CRC risk. We tested for gene-environment interactions using case-only analyses (N = 577) and compared statistically significant results to those obtained using case-unaffected sibling comparisons (N = 307 sibships). Results Carriers of the APEX1 codon 51 Gln/His genotype had a reduced CRC risk compared to carriers of the Gln/Gln genotype (OR 0.15, 95% CI 0.03-0.69, p = 0.015). The association between higher red meat intake (>3 servings/week) and CRC was modified by the PARP Val762Ala SNP (case-only interaction p = 0.026). This SNP also modified the association between higher intake of high-temperature cooked red meat (case-only interaction p = 0.0009). Conclusions We report evidence that the BER pathway PARP gene modifies the association of diets high in red meat cooked at high temperatures with risk of CRC. Impact Our findings suggest a contribution to colorectal carcinogenesis of free radical damage as one of the possible harmful effects of a high-red meat diet. PMID:21037106

  8. EGFR ligands and DNA repair genes: genomic predictors of complete response after capecitabine-based chemoradiotherapy in locally advanced rectal cancer.

    PubMed

    Sebio, A; Salazar, J; Páez, D; Berenguer-Llergo, A; Del Río, E; Tobeña, M; Martín-Richard, M; Sullivan, I; Targarona, E; Balart, J; Baiget, M; Barnadas, A

    2015-02-01

    Epidermal growth factor receptor (EGFR) activation by radiation leads to increased cell proliferation and acts as a radioresistance mechanism. Neoadjuvant chemoradiation is the standard of care for locally advanced rectal cancer, and to date, no biomarkers of response have been found. We analyzed polymorphisms in the EGFR and its ligands, DNA repair genes and the thymidylate synthase in 84 stages II and III rectal cancer patients treated with neoadjuvant capecitabine plus radiotherapy. The rs11942466 polymorphism in the amphiregulin (AREG) gene region was associated with a pathological complete response (ypCR) (odds ratio: 0.26; 95% confidence interval: 0.06-0.79; P=0.014). The rs11615 C>T polymorphism in the ERCC1 gene also correlated with the ypCR as no patients with a C/C genotype achieved ypCR; P=0.023. This is the first work to propose variants within the AREG and the ERCC1 genes as promising predictive biomarkers of ypCR in rectal cancer. PMID:25026457

  9. Epigenetic Studies Point to DNA Replication/Repair Genes as a Basis for the Heritable Nature of Long Term Complications in Diabetes.

    PubMed

    Leontovich, Alexey A; Intine, Robert V; Sarras, Michael P

    2016-01-01

    Metabolic memory (MM) is defined as the persistence of diabetic (DM) complications even after glycemic control is pharmacologically achieved. Using a zebrafish diabetic model that induces a MM state, we previously reported that, in this model, tissue dysfunction was of a heritable nature based on cell proliferation studies in limb tissue and this correlated with epigenetic DNA methylation changes that paralleled alterations in gene expression. In the current study, control, DM, and MM excised fin tissues were further analyzed by MeDIP sequencing and microarray techniques. Bioinformatics analysis of the data found that genes of the DNA replication/DNA metabolism process group (with upregulation of the apex1, mcm2, mcm4, orc3, lig1, and dnmt1 genes) were altered in the DM state and these molecular changes continued into MM. Interestingly, DNA methylation changes could be found as far as 6-13 kb upstream of the transcription start site for these genes suggesting potential higher levels of epigenetic control. In conclusion, DNA methylation changes in members of the DNA replication/repair process group best explain the heritable nature of cell proliferation impairment found in the zebrafish DM/MM model. These results are consistent with human diabetic epigenetic studies and provide one explanation for the persistence of long term tissue complications as seen in diabetes. PMID:26981540

  10. Epigenetic Studies Point to DNA Replication/Repair Genes as a Basis for the Heritable Nature of Long Term Complications in Diabetes

    PubMed Central

    Leontovich, Alexey A.; Intine, Robert V.; Sarras, Michael P.

    2016-01-01

    Metabolic memory (MM) is defined as the persistence of diabetic (DM) complications even after glycemic control is pharmacologically achieved. Using a zebrafish diabetic model that induces a MM state, we previously reported that, in this model, tissue dysfunction was of a heritable nature based on cell proliferation studies in limb tissue and this correlated with epigenetic DNA methylation changes that paralleled alterations in gene expression. In the current study, control, DM, and MM excised fin tissues were further analyzed by MeDIP sequencing and microarray techniques. Bioinformatics analysis of the data found that genes of the DNA replication/DNA metabolism process group (with upregulation of the apex1, mcm2, mcm4, orc3, lig1, and dnmt1 genes) were altered in the DM state and these molecular changes continued into MM. Interestingly, DNA methylation changes could be found as far as 6–13 kb upstream of the transcription start site for these genes suggesting potential higher levels of epigenetic control. In conclusion, DNA methylation changes in members of the DNA replication/repair process group best explain the heritable nature of cell proliferation impairment found in the zebrafish DM/MM model. These results are consistent with human diabetic epigenetic studies and provide one explanation for the persistence of long term tissue complications as seen in diabetes. PMID:26981540

  11. Incisional Hernia in Women: Predisposing Factors and Management Where Mesh is not Readily Available

    PubMed Central

    Agbakwuru, EA; Olabanji, JK; Alatise, OI; Okwerekwu, RO; Esimai, OA

    2009-01-01

    Background / Aim: Incisional hernia is still relatively common in our practice. The aim of the study was to identify risk factors associated with incisional hernia in our region. The setting is the Obafemi Awolowo University Teaching Hospitals Complex, Ile-Ife, Nigeria during a period when prosthetic mesh was not readily available. Patients and Methods: All the women who presented with incisional hernia between 1996 and 2005 were prospectively studied using a standard form to obtain information on pre-hernia (index) operations and possible predisposing factors. They all had open surgical repair and were followed up for 18–60 months. Results: Forty-four women were treated during study period. The index surgeries leading to the hernias were emergency caesarian section 26/44 (59.1%), emergency exploratory laparotomy 6/44 (13.6%), and elective surgeries 12/44 (27.3%). Major associated risk factors were the use of wrong suture materials for fascia repair, midline incisions, wound sepsis, and overweight. Conclusion: For elective surgeries, reduction of weight should be encouraged when appropriate, and transverse incisions are preferred. Absorbable sutures, especially chromic catgut, should be avoided in fascia closure. Antibiotics should be used for complicated obstetric cases. PMID:21483511

  12. The TMPRSS2-ERG gene fusion blocks XRCC4-mediated non-homologous end-joining repair and radiosensitizes prostate cancer cells to PARP inhibition

    PubMed Central

    Chatterjee, Payel; Choudhary, Gaurav S.; Alswillah, Turkeyah; Xiong, Xiahui; Heston, Warren D.; Magi-Galuzzi, Cristina; Zhang, Junran; Klein, Eric A.; Almasan, Alexandru

    2015-01-01

    Exposure to genotoxic agents, such as ionizing radiation (IR) produces DNA damage leading to DNA double-strand breaks (DSBs); IR toxicity is augmented when the DNA repair is impaired. We reported that radiosensitization by a PARP inhibitor (PARPi) was highly prominent in prostate cancer (PCa) cells expressing the TMPRSS2-ERG gene fusion protein. Here, we show that TMPRSS2-ERG blocks non-homologous end-joining (NHEJ) DNA repair by inhibiting DNA-PKcs. VCaP cells, which harbor TMPRSS2-ERG and PC3 cells that stably express it displayed γH2AX and 53BP1 foci constitutively, indicating persistent DNA damage that was absent if TMPRSS2-ERG was depleted by siRNA in VCaP cells. The extent of DNA damage was enhanced and associated with TMPRSS2-ERG’s ability to inhibit DNA-PKcs function, as indicated by its own phosphorylation (Thr2609, Ser2056) and that of its substrate, Ser1778-53BP1. DNA-PKcs deficiency caused by TMPRSS2-ERG destabilized critical NHEJ components on chromatin. Thus, XRCC4 was not recruited to chromatin, with retention of other NHEJ core factors being reduced. DNA-PKcs autophosphorylation was restored to the level of parental cells when TMPRSS2-ERG was depleted by siRNA. Following IR, TMPRSS2-ERG-expressing PC3 cells had elevated Rad51 foci and homologous recombination (HR) activity, indicating that HR compensated for defective NHEJ in these cells, hence addressing why TMPRSS2-ERG alone did not lead to radiosensitization. However, the presence of TMPRSS2-ERG, by inhibiting NHEJ DNA repair, enhanced PARPi-mediated radiosensitization. IR in combination with PARPi resulted in enhanced DNA damage in TMPRSS2-ERG-expressing cells. Thus, by inhibiting NHEJ, TMPRSS2-ERG provides a synthetic lethal interaction with PARPi in PCa patients expressing TMPRSS2-ERG. PMID:26026052

  13. The TMPRSS2-ERG Gene Fusion Blocks XRCC4-Mediated Nonhomologous End-Joining Repair and Radiosensitizes Prostate Cancer Cells to PARP Inhibition.

    PubMed

    Chatterjee, Payel; Choudhary, Gaurav S; Alswillah, Turkeyah; Xiong, Xiahui; Heston, Warren D; Magi-Galluzzi, Cristina; Zhang, Junran; Klein, Eric A; Almasan, Alexandru

    2015-08-01

    Exposure to genotoxic agents, such as ionizing radiation (IR), produces DNA damage, leading to DNA double-strand breaks (DSB); IR toxicity is augmented when the DNA repair is impaired. We reported that radiosensitization by a PARP inhibitor (PARPi) was highly prominent in prostate cancer cells expressing the TMPRSS2-ERG gene fusion protein. Here, we show that TMPRSS2-ERG blocks nonhomologous end-joining (NHEJ) DNA repair by inhibiting DNA-PKcs. VCaP cells, which harbor TMPRSS2-ERG and PC3 cells that stably express it, displayed γH2AX and 53BP1 foci constitutively, indicating persistent DNA damage that was absent if TMPRSS2-ERG was depleted by siRNA in VCaP cells. The extent of DNA damage was enhanced and associated with TMPRSS2-ERG's ability to inhibit DNA-PKcs function, as indicated by its own phosphorylation (Thr2609, Ser2056) and that of its substrate, Ser1778-53BP1. DNA-PKcs deficiency caused by TMPRSS2-ERG destabilized critical NHEJ components on chromatin. Thus, XRCC4 was not recruited to chromatin, with retention of other NHEJ core factors being reduced. DNA-PKcs autophosphorylation was restored to the level of parental cells when TMPRSS2-ERG was depleted by siRNA. Following IR, TMPRSS2-ERG-expressing PC3 cells had elevated Rad51 foci and homologous recombination (HR) activity, indicating that HR compensated for defective NHEJ in these cells, hence addressing why TMPRSS2-ERG alone did not lead to radiosensitization. However, the presence of TMPRSS2-ERG, by inhibiting NHEJ DNA repair, enhanced PARPi-mediated radiosensitization. IR in combination with PARPi resulted in enhanced DNA damage in TMPRSS2-ERG-expressing cells. Therefore, by inhibiting NHEJ, TMPRSS2-ERG provides a synthetic lethal interaction with PARPi in prostate cancer patients expressing TMPRSS2-ERG. PMID:26026052

  14. Incompatibilities Involving Yeast Mismatch Repair Genes: A Role for Genetic Modifiers and Implications for Disease Penetrance and Variation in Genomic Mutation Rates

    PubMed Central

    Demogines, Ann; Wong, Alex; Aquadro, Charles; Alani, Eric

    2008-01-01

    Genetic background effects underlie the penetrance of most genetically determined phenotypes, including human diseases. To explore how such effects can modify a mutant phenotype in a genetically tractable system, we examined an incompatibility involving the MLH1 and PMS1 mismatch repair genes using a large population sample of geographically and ecologically diverse Saccharomyces cerevisiae strains. The mismatch repair incompatibility segregates into naturally occurring yeast strains, with no strain bearing the deleterious combination. In assays measuring the mutator phenotype conferred by different combinations of MLH1 and PMS1 from these strains, we observed a mutator phenotype only in combinations predicted to be incompatible. Surprisingly, intragenic modifiers could be mapped that specifically altered the strength of the incompatibility over a 20-fold range. Together, these observations provide a powerful model in which to understand the basis of disease penetrance and how such genetic variation, created through mating, could result in new mutations that could be the raw material of adaptive evolution in yeast populations. PMID:18566663

  15. Downregulation of SWI5 and CTC1 genes: hepatitis B virus DNA polymerase transactivated protein 1-mediated inhibition of DNA repair.

    PubMed

    Yao, X-K; Pan, Z-P; Li, Y; Lun, Y-Z; Chi, Q; Jiang, S-J; Wang, F; Sui, W

    2016-06-01

    Hepatitis B virus (HBV) DNA polymerase transactivated protein 1 (HBVDNAPTP1) is a novel protein upregulated by HBV DNA polymerase, which has been screened by suppression subtractive hybridization technique (SSH) (GenBank Acc. No. AY450389). A vector pcDNA3.1 (-)/myc-His A-HBVDNAPTP1 was constructed and used to transfect acute monocytic leukemia cell line THP-1. HBVDNAPTP1 expression was detected by Western blot analysis in the cells. A cDNA library of genes downregulated by HBVDNAPTP1 in THP-1 cells was made in pGEM-T Easy using SSH. The cDNAs were sequenced and analyzed with BLAST search against the sequences in GenBank. Some sequences, such as DNA repair protein SWI5 homolog (SWI5) and CTS telomere maintenance complex component 1 (CTC1), might be involved in DNA repair. Protein expression of SWI5 and CTC1 was identified by Western blot in THP-1 cells. HBVDNAPTP1 could downregulate the expression of SWI5 and CTC1 at translation level. PMID:27265469

  16. Knock-in of large reporter genes in human cells via CRISPR/Cas9-induced homology-dependent and independent DNA repair

    PubMed Central

    He, Xiangjun; Tan, Chunlai; Wang, Feng; Wang, Yaofeng; Zhou, Rui; Cui, Dexuan; You, Wenxing; Zhao, Hui; Ren, Jianwei; Feng, Bo

    2016-01-01

    CRISPR/Cas9-induced site-specific DNA double-strand breaks (DSBs) can be repaired by homology-directed repair (HDR) or non-homologous end joining (NHEJ) pathways. Extensive efforts have been made to knock-in exogenous DNA to a selected genomic locus in human cells; which, however, has focused on HDR-based strategies and was proven inefficient. Here, we report that NHEJ pathway mediates efficient rejoining of genome and plasmids following CRISPR/Cas9-induced DNA DSBs, and promotes high-efficiency DNA integration in various human cell types. With this homology-independent knock-in strategy, integration of a 4.6 kb promoterless ires-eGFP fragment into the GAPDH locus yielded up to 20% GFP+ cells in somatic LO2 cells, and 1.70% GFP+ cells in human embryonic stem cells (ESCs). Quantitative comparison further demonstrated that the NHEJ-based knock-in is more efficient than HDR-mediated gene targeting in all human cell types examined. These data support that CRISPR/Cas9-induced NHEJ provides a valuable new path for efficient genome editing in human ESCs and somatic cells. PMID:26850641

  17. Knock-in of large reporter genes in human cells via CRISPR/Cas9-induced homology-dependent and independent DNA repair.

    PubMed

    He, Xiangjun; Tan, Chunlai; Wang, Feng; Wang, Yaofeng; Zhou, Rui; Cui, Dexuan; You, Wenxing; Zhao, Hui; Ren, Jianwei; Feng, Bo

    2016-05-19

    CRISPR/Cas9-induced site-specific DNA double-strand breaks (DSBs) can be repaired by homology-directed repair (HDR) or non-homologous end joining (NHEJ) pathways. Extensive efforts have been made to knock-in exogenous DNA to a selected genomic locus in human cells; which, however, has focused on HDR-based strategies and was proven inefficient. Here, we report that NHEJ pathway mediates efficient rejoining of genome and plasmids following CRISPR/Cas9-induced DNA DSBs, and promotes high-efficiency DNA integration in various human cell types. With this homology-independent knock-in strategy, integration of a 4.6 kb promoterless ires-eGFP fragment into the GAPDH locus yielded up to 20% GFP+ cells in somatic LO2 cells, and 1.70% GFP+ cells in human embryonic stem cells (ESCs). Quantitative comparison further demonstrated that the NHEJ-based knock-in is more efficient than HDR-mediated gene targeting in all human cell types examined. These data support that CRISPR/Cas9-induced NHEJ provides a valuable new path for efficient genome editing in human ESCs and somatic cells. PMID:26850641

  18. Meta-analysis of estrogen response in MCF-7 distinguishes early target genes involved in signaling and cell proliferation from later target genes involved in cell cycle and DNA repair

    PubMed Central

    2011-01-01

    Background Many studies have been published outlining the global effects of 17β-estradiol (E2) on gene expression in human epithelial breast cancer derived MCF-7 cells. These studies show large variation in results, reporting between ~100 and ~1500 genes regulated by E2, with poor overlap. Results We performed a meta-analysis of these expression studies, using the Rank product method to obtain a more accurate and stable list of the differentially expressed genes, and of pathways regulated by E2. We analyzed 9 time-series data sets, concentrating on response at 3-4 hrs (early) and at 24 hrs (late). We found >1000 statistically significant probe sets after correction for multiple testing at 3-4 hrs, and >2000 significant probe sets at 24 hrs. Differentially expressed genes were examined by pathway analysis. This revealed 15 early response pathways, mostly related to cell signaling and proliferation, and 20 late response pathways, mostly related to breast cancer, cell division, DNA repair and recombination. Conclusions Our results confirm that meta-analysis identified more differentially expressed genes than the individual studies, and that these genes act together in networks. These results provide new insight into E2 regulated mechanisms, especially in the context of breast cancer. PMID:21878096

  19. Adenovirus-mediated HIF-1α gene transfer promotes repair of mouse airway allograft microvasculature and attenuates chronic rejection.

    PubMed

    Jiang, Xinguo; Khan, Mohammad A; Tian, Wen; Beilke, Joshua; Natarajan, Ramesh; Kosek, Jon; Yoder, Mervin C; Semenza, Gregg L; Nicolls, Mark R

    2011-06-01

    Chronic rejection, manifested as small airway fibrosis (obliterative bronchiolitis [OB]), is the main obstacle to long-term survival in lung transplantation. Recent studies demonstrate that the airways involved in a lung transplant are relatively hypoxic at baseline and that OB pathogenesis may be linked to ischemia induced by a transient loss of airway microvasculature. Here, we show that HIF-1α mediates airway microvascular repair in a model of orthotopic tracheal transplantation. Grafts with a conditional knockout of Hif1a demonstrated diminished recruitment of recipient-derived Tie2⁺ angiogenic cells to the allograft, impaired repair of damaged microvasculature, accelerated loss of microvascular perfusion, and hastened denudation of epithelial cells. In contrast, graft HIF-1α overexpression induced via an adenoviral vector prolonged airway microvascular perfusion, preserved epithelial integrity, extended the time window for the graft to be rescued from chronic rejection, and attenuated airway fibrotic remodeling. HIF-1α overexpression induced the expression of proangiogenic factors such as Sdf1, Plgf, and Vegf, and promoted the recruitment of vasoreparative Tie2⁺ cells. This study demonstrates that a therapy that enhances vascular integrity during acute rejection may promote graft health and prevent chronic rejection. PMID:21606594

  20. Tumor Mismatch Repair Immunohistochemistry and DNA MLH1 Methylation Testing of Patients With Endometrial Cancer Diagnosed at Age Younger Than 60 Years Optimizes Triage for Population-Level Germline Mismatch Repair Gene Mutation Testing

    PubMed Central

    Buchanan, Daniel D.; Tan, Yen Y.; Walsh, Michael D.; Clendenning, Mark; Metcalf, Alexander M.; Ferguson, Kaltin; Arnold, Sven T.; Thompson, Bryony A.; Lose, Felicity A.; Parsons, Michael T.; Walters, Rhiannon J.; Pearson, Sally-Ann; Cummings, Margaret; Oehler, Martin K.; Blomfield, Penelope B.; Quinn, Michael A.; Kirk, Judy A.; Stewart, Colin J.; Obermair, Andreas; Young, Joanne P.; Webb, Penelope M.; Spurdle, Amanda B.

    2014-01-01

    Purpose Clinicopathologic data from a population-based endometrial cancer cohort, unselected for age or family history, were analyzed to determine the optimal scheme for identification of patients with germline mismatch repair (MMR) gene mutations. Patients and Methods Endometrial cancers from 702 patients recruited into the Australian National Endometrial Cancer Study (ANECS) were tested for MMR protein expression using immunohistochemistry (IHC) and for MLH1 gene promoter methylation in MLH1-deficient cases. MMR mutation testing was performed on germline DNA of patients with MMR-protein deficient tumors. Prediction of germline mutation status was compared for combinations of tumor characteristics, age at diagnosis, and various clinical criteria (Amsterdam, Bethesda, Society of Gynecologic Oncology, ANECS). Results Tumor MMR-protein deficiency was detected in 170 (24%) of 702 cases. Germline testing of 158 MMR-deficient cases identified 22 truncating mutations (3% of all cases) and four unclassified variants. Tumor MLH1 methylation was detected in 99 (89%) of 111 cases demonstrating MLH1/PMS2 IHC loss; all were germline MLH1 mutation negative. A combination of MMR IHC plus MLH1 methylation testing in women younger than 60 years of age at diagnosis provided the highest positive predictive value for the identification of mutation carriers at 46% versus ≤ 41% for any other criteria considered. Conclusion Population-level identification of patients with MMR mutation-positive endometrial cancer is optimized by stepwise testing for tumor MMR IHC loss in patients younger than 60 years, tumor MLH1 methylation in individuals with MLH1 IHC loss, and germline mutations in patients exhibiting loss of MSH6, MSH2, or PMS2 or loss of MLH1/PMS2 with absence of MLH1 methylation. PMID:24323032

  1. DNA repair gene XPD Asp312Asn and XRCC4 G-1394T polymorphisms and the risk of autism spectrum disorder.

    PubMed

    Dasdemir, S; Guven, M; Pekkoc, K C; Ulucan, H; Dogangun, B; Kirtas, E; Kadak, M T; Kucur, M; Seven, M

    2016-01-01

    Autism spectrum disorder (ASD) is a complex disorder, and its extreme heterogeneity further complicates our understanding of its biology. Epidemiological evidence from family and twin studies supports a strong genetic component in ASD etiology. Oxidative stress and abnormal DNA methylation have been implicated in the pathophysiology of ASD. Brain tissues from ASD cases showed higher levels of oxidative stress biomarkers than healthy controls in postmortem analysis. Association between oxidative stress and DNA damage has been well-known. Thus, we sought to investigate a potential link between DNA repair genes and ASD and analyze the role of XPD Asp312Asn and XRCC4 G-1394T gene polymorphisms for ASD in the Turkish population. Genotyping was conducted by PCR-RFLP based on 100 patients and 96 unrelated healthy controls. We, for the first time, demonstrated a positive association between XRCC4 gene variants and ASD risk. Frequencies of XRCC4-1394 T/G+G/G genotypes were higher in patients (%34) than the controls (%18.7). The statistical analysis revealed that the individuals who had XRCC4-1394 T/G+G/G genotype had an increased risk for ASD (OR = 2.23, 95% CI = 1.10-4.55). However, no significant association was found for XPD Asp312Asn polymorphism with the risk of ASD. Our findings suggest that XRCC4 G-1394T polymorphism might be associated with ASD pathogenesis. PMID:27064873

  2. Effects of the eukaryotic initiation factor 6 gene on expression levels of inflammatory mediators in M2 macrophages during scar repair.

    PubMed

    Bai, Yong-Qiang; Feng, Jian-Ke; Zhang, Qing-Fu; Wu, Hong-Zhi; Du, Ya-Ru; Wei, Wei

    2016-07-01

    The aim of the present study was to evaluate the effects of the eukaryotic initiation factor 6 (eIF6) gene on the secretion of M2 macrophage fibrosis‑associated factors and the expression levels of key proteases during scar repair. Male eIF6 wild‑type (eIF6+/+) and knockout (eIF6+/‑) C57BL/6 mice were intraperitoneally lavaged to obtain macrophages, which were induced to the M2 type using interleukin‑4. Differences between the gene expression profiles of these macrophages were compared with gene microarrays, and the results were validated using reverse transcription-quantitative polymerase chain reaction analysis and ELISA. Compared with the eIF6+/‑ mice, the mRNA and protein expression levels of vascular endothelial growth factor (VEGF) and tissue inhibitor of metalloproteinase‑2 (TIMP‑2) in the M2 macrophages of the eIF6+/+ mice were significantly downregulated (P<0.05), whereas the mRNA and protein expression levels of matrix metalloproteinase‑2 (MMP‑2) were significantly upregulated (P<0.05). Therefore, the results indicated that eIF6 alleviated cicatrization, possibly by inhibiting the generation of VEGF, in order to prevent overgrowth of blood vessels and granulation tissues, and to regulate the MMP-2/TIMP-2 ratio to balance the degradation and deposition of the extracellular matrix. PMID:27220600

  3. CITED2 silencing sensitizes cancer cells to cisplatin by inhibiting p53 trans-activation and chromatin relaxation on the ERCC1 DNA repair gene

    PubMed Central

    Liu, Yu-Chin; Chang, Pu-Yuan; Chao, Chuck C.-K.

    2015-01-01

    In this study, we show that silencing of CITED2 using small-hairpin RNA (shCITED2) induced DNA damage and reduction of ERCC1 gene expression in HEK293, HeLa and H1299 cells, even in the absence of cisplatin. In contrast, ectopic expression of ERCC1 significantly reduced intrinsic and induced DNA damage levels, and rescued the effects of CITED2 silencing on cell viability. The effects of CITED2 silencing on DNA repair and cell death were associated with p53 activity. Furthermore, CITED2 silencing caused severe elimination of the p300 protein and markers of relaxed chromatin (acetylated H3 and H4, i.e. H3K9Ac and H3K14Ac) in HEK293 cells. Chromatin immunoprecipitation assays further revealed that DNA damage induced binding of p53 along with H3K9Ac or H3K14Ac at the ERCC1 promoter, an effect which was almost entirely abrogated by silencing of CITED2 or p300. Moreover, lentivirus-based CITED2 silencing sensitized HeLa cell line-derived tumor xenografts to cisplatin in immune-deficient mice. These results demonstrate that CITED2/p300 can be recruited by p53 at the promoter of the repair gene ERCC1 in response to cisplatin-induced DNA damage. The CITED2/p300/p53/ERCC1 pathway is thus involved in the cell response to cisplatin and represents a potential target for cancer therapy. PMID:26384430

  4. CITED2 silencing sensitizes cancer cells to cisplatin by inhibiting p53 trans-activation and chromatin relaxation on the ERCC1 DNA repair gene.

    PubMed

    Liu, Yu-Chin; Chang, Pu-Yuan; Chao, Chuck C-K

    2015-12-15

    In this study, we show that silencing of CITED2 using small-hairpin RNA (shCITED2) induced DNA damage and reduction of ERCC1 gene expression in HEK293, HeLa and H1299 cells, even in the absence of cisplatin. In contrast, ectopic expression of ERCC1 significantly reduced intrinsic and induced DNA damage levels, and rescued the effects of CITED2 silencing on cell viability. The effects of CITED2 silencing on DNA repair and cell death were associated with p53 activity. Furthermore, CITED2 silencing caused severe elimination of the p300 protein and markers of relaxed chromatin (acetylated H3 and H4, i.e. H3K9Ac and H3K14Ac) in HEK293 cells. Chromatin immunoprecipitation assays further revealed that DNA damage induced binding of p53 along with H3K9Ac or H3K14Ac at the ERCC1 promoter, an effect which was almost entirely abrogated by silencing of CITED2 or p300. Moreover, lentivirus-based CITED2 silencing sensitized HeLa cell line-derived tumor xenografts to cisplatin in immune-deficient mice. These results demonstrate that CITED2/p300 can be recruited by p53 at the promoter of the repair gene ERCC1 in response to cisplatin-induced DNA damage. The CITED2/p300/p53/ERCC1 pathway is thus involved in the cell response to cisplatin and represents a potential target for cancer therapy. PMID:26384430

  5. Mouse BAZ1A (ACF1) Is Dispensable for Double-Strand Break Repair but Is Essential for Averting Improper Gene Expression during Spermatogenesis

    PubMed Central

    Dowdle, James A.; Mehta, Monika; Kass, Elizabeth M.; Vuong, Bao Q.; Inagaki, Akiko; Egli, Dieter; Jasin, Maria; Keeney, Scott

    2013-01-01

    ATP-dependent chromatin remodelers control DNA access for transcription, recombination, and other processes. Acf1 (also known as BAZ1A in mammals) is a defining subunit of the conserved ISWI-family chromatin remodelers ACF and CHRAC, first purified over 15 years ago from Drosophila melanogaster embryos. Much is known about biochemical properties of ACF and CHRAC, which move nucleosomes in vitro and in vivo to establish ordered chromatin arrays. Genetic studies in yeast, flies and cultured human cells clearly implicate these complexes in transcriptional repression via control of chromatin structures. RNAi experiments in transformed mammalian cells in culture also implicate ACF and CHRAC in DNA damage checkpoints and double-strand break repair. However, their essential in vivo roles in mammals are unknown. Here, we show that Baz1a-knockout mice are viable and able to repair developmentally programmed DNA double-strand breaks in the immune system and germ line, I-SceI endonuclease-induced breaks in primary fibroblasts via homologous recombination, and DNA damage from mitomycin C exposure in vivo. However, Baz1a deficiency causes male-specific sterility in accord with its high expression in male germ cells, where it displays dynamic, stage-specific patterns of chromosomal localization. Sterility is caused by pronounced defects in sperm development, most likely a consequence of massively perturbed gene expression in spermatocytes and round spermatids in the absence of BAZ1A: the normal spermiogenic transcription program is largely intact but more than 900 other genes are mis-regulated, primarily reflecting inappropriate up-regulation. We propose that large-scale changes in chromatin composition that occur during spermatogenesis create a window of vulnerability to promiscuous transcription changes, with an essential function of ACF and/or CHRAC chromatin remodeling activities being to safeguard against these alterations. PMID:24244200

  6. The association of six polymorphisms of five genes involved in three steps of nucleotide excision repair pathways with hepatocellular cancer risk

    PubMed Central

    Yang, Huai-wei; Sun, Li-ping; Yuan, Yuan

    2016-01-01

    Background Hundreds of single nucleotide polymorphisms (SNPs) of the genes encoding nucleotide excision repair (NER) proteins are involved in every step of the DNA recognition–unwinding–incision process, which may affect cancer risk. However, only a limited number of studies have examined the association of NER SNPs with hepatocellular cancer (HCC) risk. Results In screening stage, single-locus analysis showed that six SNPs in five genes were associated with HCC risk, including three risk SNPs (XPA rs10817938, XPC rs1870134 and ERCC2 rs238417) and three protective SNPs (ERCC1 rs2298881 and rs3212961, and ERCC5 rs873601). In verification stage, only XPC rs1870134 was verified to be associated with HCC risk (P = 4.7 × 10−4). Furthermore, multivariate logistic regression and MDR analysis consistently revealed a gene–gene interaction among ERCC1 rs2298881 and XPC rs1870134 SNPs associated with HCC risk (Pinteraction = 0.023). When analyzing the effect of the positive SNP on the mRNA expression, we found XPC rs1870134 GG genotype which was associated with an increased HCC risk showed lower XPC mRNA expression. Methods This study designed as “screening-verification” experiments and included a total of 1472 participants (570 HCC patients vs. 902 controls). We explored 39 SNPs in eight genes involved in NER Pathways, including XPA, XPC, DDB2, ERCC3, ERCC2, ERCC1, ERCC4 and ERCC5, using Sequenom MassARRAY and KASPar platform. Eighty-six cases of HCC and the neighboring noncancerous tissues were subjected to the measurement of mRNA expression level of the promising gene. Conclusions XPC promoter rs1870134 SNP and SNP-SNP interaction were associated with HCC risk. PMID:26967386

  7. Factors predisposing to a complicated initial febrile convulsion.

    PubMed Central

    Wallace, S J

    1975-01-01

    131 consecutive admissions to hospital for a first febrile convulsion were studied to find which factors predisposed to a complicated fit--defined as one lasting more than 30 minutes, unilateral, or repeated within the same illness. A significant excess of complicated attacks occurred where the age of onset was less than 16 months, where both family history of convulsive disorder and perinatal abnormality were present, and, in females only, where it was suspected that neurological disorder preceded the first fit. PMID:1220608

  8. DIMETHYLARSINIC ACID ALTERS EXPRESSION OF OXIDATIVE STRESS AND DNA REPAIR GENES IN A DOSE DEPENDENT MANNER IN THE TRANSITIONAL EPITHELIUM OF THE URINARY BLADDER FROM FEMALE F344 RATS.

    EPA Science Inventory

    Dose-dependent alteration of oxidative stress and DNA repair gene expression by Dimethylarsinic acid [DMA(V)] in transitional epithelium of urinary bladder from female F344 rats.
    Arsenic (As) is a major concern as millions of people are at risk from drinking arsenic contaminat...

  9. Gene-Silencing Screen for Mammalian Axon Regeneration Identifies Inpp5f (Sac2) as an Endogenous Suppressor of Repair after Spinal Cord Injury

    PubMed Central

    Zou, Yixiao; Stagi, Massimiliano; Wang, Xingxing; Yigitkanli, Kazim; Siegel, Chad S.; Nakatsu, Fubito; Cafferty, William B. J.

    2015-01-01

    Axonal growth and neuronal rewiring facilitate functional recovery after spinal cord injury. Known interventions that promote neural repair remain limited in their functional efficacy. To understand genetic determinants of mammalian CNS axon regeneration, we completed an unbiased RNAi gene-silencing screen across most phosphatases in the genome. We identified one known and 17 previously unknown phosphatase suppressors of injury-induced CNS axon growth. Silencing Inpp5f (Sac2) leads to robust enhancement of axon regeneration and growth cone reformation. Results from cultured Inpp5f−/− neurons confirm lentiviral shRNA results from the screen. Consistent with the nonoverlapping substrate specificity between Inpp5f and PTEN, rapamycin does not block enhanced regeneration in Inpp5f−/− neurons, implicating mechanisms independent of the PI3K/AKT/mTOR pathway. Inpp5f−/− mice develop normally, but show enhanced anatomical and functional recovery after mid-thoracic dorsal hemisection injury. More serotonergic axons sprout and/or regenerate caudal to the lesion level, and greater numbers of corticospinal tract axons sprout rostral to the lesion. Functionally, Inpp5f-null mice exhibit enhanced recovery of motor functions in both open-field and rotarod tests. This study demonstrates the potential of an unbiased high-throughput functional screen to identify endogenous suppressors of CNS axon growth after injury, and reveals Inpp5f (Sac2) as a novel suppressor of CNS axon repair after spinal cord injury. SIGNIFICANCE STATEMENT The extent of axon regeneration is a critical determinant of neurological recovery from injury, and is extremely limited in the adult mammalian CNS. We describe an unbiased gene-silencing screen that uncovered novel molecules suppressing axonal regeneration. Inpp5f (Sac2) gene deletion promoted recovery from spinal cord injury with no side effects. The mechanism of action is distinct from another lipid phosphatase implicated in regeneration

  10. Gastroschisis repair

    MedlinePlus

    ... and surgery in general are: Allergic reactions to medicines Breathing problems Bleeding Infection Risks for gastroschisis repair are: Breathing problems if the baby's belly area (abdominal space) is smaller than normal. The baby may need ...

  11. Hydrocele repair

    MedlinePlus

    ... is excellent. However, another hydrocele may form over time, or if there was also a hernia present. Alternative Names Hydrocelectomy Images Hydrocele repair - series References Aiken JJ, Oldham KT. Inguinal hernias. In: ...

  12. Silencing of end-joining repair for efficient site-specific gene insertion after TALEN/CRISPR mutagenesis in Aedes aegypti

    PubMed Central

    Basu, Sanjay; Aryan, Azadeh; Overcash, Justin M.; Samuel, Glady Hazitha; Anderson, Michelle A. E.; Dahlem, Timothy J.; Myles, Kevin M.; Adelman, Zach N.

    2015-01-01

    Conventional control strategies for mosquito-borne pathogens such as malaria and dengue are now being complemented by the development of transgenic mosquito strains reprogrammed to generate beneficial phenotypes such as conditional sterility or pathogen resistance. The widespread success of site-specific nucleases such as transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 in model organisms also suggests that reprogrammable gene drive systems based on these nucleases may be capable of spreading such beneficial phenotypes in wild mosquito populations. Using the mosquito Aedes aegypti, we determined that mutations in the FokI domain used in TALENs to generate obligate heterodimeric complexes substantially and significantly reduce gene editing rates. We found that CRISPR/Cas9-based editing in the mosquito Ae. aegypti is also highly variable, with the majority of guide RNAs unable to generate detectable editing. By first evaluating candidate guide RNAs using a transient embryo assay, we were able to rapidly identify highly effective guide RNAs; focusing germ line-based experiments only on this cohort resulted in consistently high editing rates of 24–90%. Microinjection of double-stranded RNAs targeting ku70 or lig4, both essential components of the end-joining response, increased recombination-based repair in early embryos as determined by plasmid-based reporters. RNAi-based suppression of Ku70 concurrent with embryonic microinjection of site-specific nucleases yielded consistent gene insertion frequencies of 2–3%, similar to traditional transposon- or ΦC31-based integration methods but without the requirement for an initial docking step. These studies should greatly accelerate investigations into mosquito biology, streamline development of transgenic strains for field releases, and simplify the evaluation of novel Cas9-based gene drive systems. PMID:25775608

  13. Silencing of end-joining repair for efficient site-specific gene insertion after TALEN/CRISPR mutagenesis in Aedes aegypti.

    PubMed

    Basu, Sanjay; Aryan, Azadeh; Overcash, Justin M; Samuel, Glady Hazitha; Anderson, Michelle A E; Dahlem, Timothy J; Myles, Kevin M; Adelman, Zach N

    2015-03-31

    Conventional control strategies for mosquito-borne pathogens such as malaria and dengue are now being complemented by the development of transgenic mosquito strains reprogrammed to generate beneficial phenotypes such as conditional sterility or pathogen resistance. The widespread success of site-specific nucleases such as transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 in model organisms also suggests that reprogrammable gene drive systems based on these nucleases may be capable of spreading such beneficial phenotypes in wild mosquito populations. Using the mosquito Aedes aegypti, we determined that mutations in the FokI domain used in TALENs to generate obligate heterodimeric complexes substantially and significantly reduce gene editing rates. We found that CRISPR/Cas9-based editing in the mosquito Ae. aegypti is also highly variable, with the majority of guide RNAs unable to generate detectable editing. By first evaluating candidate guide RNAs using a transient embryo assay, we were able to rapidly identify highly effective guide RNAs; focusing germ line-based experiments only on this cohort resulted in consistently high editing rates of 24-90%. Microinjection of double-stranded RNAs targeting ku70 or lig4, both essential components of the end-joining response, increased recombination-based repair in early embryos as determined by plasmid-based reporters. RNAi-based suppression of Ku70 concurrent with embryonic microinjection of site-specific nucleases yielded consistent gene insertion frequencies of 2-3%, similar to traditional transposon- or ΦC31-based integration methods but without the requirement for an initial docking step. These studies should greatly accelerate investigations into mosquito biology, streamline development of transgenic strains for field releases, and simplify the evaluation of novel Cas9-based gene drive systems. PMID:25775608

  14. The contribution of deleterious germline mutations in BRCA1, BRCA2 and the mismatch repair genes to ovarian cancer in the population

    PubMed Central

    Song, Honglin; Cicek, Mine S.; Dicks, Ed; Harrington, Patricia; Ramus, Susan J.; Cunningham, Julie M.; Fridley, Brooke L.; Tyrer, Jonathan P.; Alsop, Jennifer; Jimenez-Linan, Mercedes; Gayther, Simon A.; Goode, Ellen L.; Pharoah, Paul D.P.

    2014-01-01

    The aim of this study was to estimate the contribution of deleterious mutations in BRCA1, BRCA2, MLH1, MSH2, MSH6 and PMS2 to invasive epithelial ovarian cancer (EOC) in the population. The coding sequence and splice site boundaries of all six genes were amplified in germline DNA from 2240 invasive EOC cases and 1535 controls. Barcoded fragment libraries were sequenced using the Illumina GAII or HiSeq and sequence data for each subject de-multiplexed prior to interpretation. GATK and Annovar were used for variant detection and annotation. After quality control 2222 cases (99.2%) and 1528 controls (99.5%) were included in the final analysis. We identified 193 EOC cases (8.7%) carrying a deleterious mutation in at least one gene compared with 10 controls (0.65%). Mutations were most frequent in BRCA1 and BRCA2, with 84 EOC cases (3.8%) carrying a BRCA1 mutation and 94 EOC cases (4.2%) carrying a BRCA2 mutation. The combined BRCA1 and BRCA2 mutation prevalence was 11% in high-grade serous disease. Seventeen EOC cases carried a mutation in a mismatch repair gene, including 10 MSH6 mutation carriers (0.45%) and 4 MSH2 mutation carriers (0.18%). At least 1 in 10 women with high-grade serous EOC has a BRCA1 or BRCA2 mutation. The development of next generation sequencing technologies enables rapid mutation screening for multiple susceptibility genes at once, suggesting that routine clinical testing of all incidence cases should be considered. PMID:24728189

  15. Evaluating the performance of clinical criteria for predicting mismatch repair gene mutations in Lynch syndrome: a comprehensive analysis of 3,671 families.

    PubMed

    Steinke, Verena; Holzapfel, Stefanie; Loeffler, Markus; Holinski-Feder, Elke; Morak, Monika; Schackert, Hans K; Görgens, Heike; Pox, Christian; Royer-Pokora, Brigitte; von Knebel-Doeberitz, Magnus; Büttner, Reinhard; Propping, Peter; Engel, Christoph

    2014-07-01

    Carriers of mismatch repair (MMR) gene mutations have a high lifetime risk for colorectal and endometrial cancers, as well as other malignancies. As mutation analysis to detect these patients is expensive and time-consuming, clinical criteria and tumor-tissue analysis are widely used as pre-screening methods. The aim of our study was to evaluate the performance of commonly applied clinical criteria (the Amsterdam I and II Criteria, and the original and revised Bethesda Guidelines) and the results of tumor-tissue analysis in predicting MMR gene mutations. We analyzed 3,671 families from the German HNPCC Registry and divided them into nine mutually exclusive groups with different clinical criteria. A total of 680 families (18.5%) were found to have a pathogenic MMR gene mutation. Among all 1,284 families with microsatellite instability-high (MSI-H) colorectal cancer, the overall mutation detection rate was 53.0%. Mutation frequencies and their distribution between the four MMR genes differed significantly between clinical groups (p < 0.001). The highest frequencies were found in families fulfilling the Amsterdam Criteria (46.4%). Families with loss of MSH2 expression had higher mutation detection rates (69.5%) than families with loss of MLH1 expression (43.1%). MMR mutations were found significantly more often in families with at least one MSI-H small-bowel cancer (p < 0.001). No MMR mutations were found among patients under 40-years-old with only colorectal adenoma. Familial clustering of Lynch syndrome-related tumors, early age of onset, and familial occurrence of small-bowel cancer were clinically relevant predictors for Lynch syndrome. PMID:24493211

  16. The use of SHP-2 gene transduced bone marrow mesenchymal stem cells to promote osteogenic differentiation and bone defect repair in rat.

    PubMed

    Fan, Dapeng; Liu, Shen; Jiang, Shichao; Li, Zhiwei; Mo, Xiumei; Ruan, Hongjiang; Zou, Gang-Ming; Fan, Cunyi

    2016-08-01

    Bone tissue engineering is a promising approach for bone regeneration, in which growth factors play an important role. The tyrosine phosphatase Src-homology region 2-containing protein tyrosine phosphatase 2 (SHP2), encoded by the PTPN11 gene, is essential for the differentiation, proliferation and metabolism of osteoblasts. However, SHP-2 has never been systematically studied for its effect in osteogenesis. We predicted that overexpression of SHP-2 could promote bone marrow-derived mesenchymal stem cell (BMSC)osteogenic differentiation and SHP-2 transduced BMSCs could enhance new bone formation, determined using the following study groups: (1) BMSCs transduced with SHP-2 and induced with osteoblast-inducing liquid (BMSCs/SHP-2/OL); (2) BMSCs transduced with SHP-2 (BMSCs/-SHP-2); (3) BMSCs induced with osteoblast-inducing liquid (BMSCs/OL) and (4) pure BMSCs. Cells were assessed for osteogenic differentiation by quantitative real-time polymerase chain reaction analysis, western blot analysis, alkaline phosphatase activity and alizarin red S staining. For in vivo assessment, cells were combined with beta-tricalcium phosphate scaffolds and transplanted into rat calvarial defects for 8 weeks. Following euthanasia, skull samples were explanted for osteogenic evaluation, including micro-computed tomography measurement, histology and immunohistochemistry staining. SHP-2 and upregulation of its gene promoted BMSC osteogenic differentiation and therefore represents a potential new therapeutic approach to bone repair. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1871-1881, 2016. PMID:26999642

  17. Differential expression of genes involved in immunity and biomineralization during Brown Ring Disease development and shell repair in the Manila clam, Ruditapes philippinarum.

    PubMed

    Jeffroy, Fanny; Brulle, Franck; Paillard, Christine

    2013-06-01

    Severe drop in Manila clams production in French aquacultured fields since the end of the 1980's is associated to Brown Ring Disease (BRD). This disease, caused by the bacteria Vibrio tapetis, is characterized by specific symptoms on the inner face of the shell. Diseased animals develop conchiolin deposit to enrobe bacteria and form new calcified layers on the shell. Suppression subtractive hybridization was performed to identify genes differentially expressed during the early interaction of V. tapetis and Ruditapes philippinarum. Results revealed 301 unique genes differentially expressed during V. tapetis challenge. Several candidates involved in immune and biomineralization processes were selected from libraries. Transcriptional expression of selected candidates was determined in hemolymph and mantle tissues and revealed spatial and temporal variations. At 56 days after infection, when clams were in phase of shell repair, transcripts of galectin and ferritin in hemocytes showed higher expression. Ca-like and serpin transcripts were specifically expressed in mantle and could contribute to defense against BRD. PMID:23500956

  18. Epigenetic suppression of Fli1, a potential predisposing factor in the pathogenesis of systemic sclerosis.

    PubMed

    Asano, Yoshihide

    2015-10-01

    Systemic sclerosis (SSc) is a multisystem connective tissue disease featured by immune abnormalities, vasculopathy and tissue fibrosis with unknown etiology. A series of studies on disease-susceptibility genes and twins have demonstrated the association of genetic factors with autoimmunity and disease severity and the contribution of environmental factors to the induction of clinical features in this disease. Friend leukemia virus integration 1 (Fli1), a member of Ets transcription factor family, is epigenetically suppressed in the lesional skin of SSc patients, suggesting that Fli1 is a potential predisposing factor of SSc reflecting the influence of environmental factors. Consistent with this idea, Fli1 deficiency induces SSc-like phenotypes in dermal fibroblasts and dermal microvascular endothelial cells in vivo and in vitro at molecular levels. Furthermore, Fli1 haploinsufficiency recapitulates tissue fibrosis, vascular activation and inflammation characteristic of SSc to a greater extent in bleomycin-treated mice. Importantly, bosentan, a dual endothelin receptor antagonist with a potential disease-modifying effect on SSc vasculopathy, reverses the expression of Fli1 protein by increasing its protein stability. Therefore, Fli1 may serve as a predisposing factor of SSc and can be a promising therapeutic target of this incurable and devastating disease. This article is part of a Directed Issue entitled: Epigenetics dynamics in development and disease. PMID:26055516

  19. Posttranscriptional regulation of the RAD5 DNA repair gene by the Dun1 kinase and the Pan2-Pan3 poly(A)-nuclease complex contributes to survival of replication blocks.

    PubMed

    Hammet, Andrew; Pike, Brietta L; Heierhorst, Jörg

    2002-06-21

    The yeast Dun1 kinase has complex checkpoint functions including DNA damage-dependent cell cycle arrest in G(2)/M, transcriptional induction of repair genes, and regulation of postreplicative DNA repair pathways. Here we report that the Dun1 forkhead-associated domain interacts with the Pan3 subunit of the poly(A)-nuclease complex and that dun1pan2 and dun1pan3 double mutants are dramatically hypersensitive to replicational stress. This phenotype was independent of the function of Dun1 in regulating deoxyribonucleotide levels as it was also observed in strains lacking the ribonucleotide reductase inhibitor Sml1. dun1pan2 mutants initially arrested normally in response to replication blocks but died in the presence of persistent replication blocks with considerably delayed kinetics compared with mutants lacking the Rad53 kinase, indicating that the double mutation does not compromise the intra-S phase checkpoint. Interestingly, the RAD5 gene involved in error-free postreplication repair pathways was specifically up-regulated in dun1pan2 double mutants. Moreover, inducible overexpression of RAD5 mimicked the double mutant phenotype by hypersensitizing dun1 mutants to replication blocks. The data indicate that Dun1 and Pan2-Pan3 cooperate to regulate the stoichiometry and thereby the activity of postreplication repair complexes, suggesting that posttranscriptional mechanisms complement the transcriptional response in the regulation of gene expression by checkpoint signaling pathways in Saccharomyces cerevisiae. PMID:11953437

  20. The Saccharomyces Cerevisiae Rad30 Gene, a Homologue of Escherichia Coli Dinb and Umuc, Is DNA Damage Inducible and Functions in a Novel Error-Free Postreplication Repair Mechanism

    PubMed Central

    McDonald, J. P.; Levine, A. S.; Woodgate, R.

    1997-01-01

    Damage-inducible mutagenesis in prokaryotes is largely dependent upon the activity of the UmuD'C-like proteins. Since many DNA repair processes are structurally and/or functionally conserved between prokaryotes and eukaryotes, we investigated the role of RAD30, a previously uncharacterized Saccharomyces cerevisiae DNA repair gene related to the Escherichia coli dinB, umuC and S. cerevisiae REV1 genes, in UV resistance and UV-induced mutagenesis. Similar to its prokaryotic homologues, RAD30 was found to be damage inducible. Like many S. cerevisiae genes involved in error-prone DNA repair, epistasis analysis clearly places RAD30 in the RAD6 group and rad30 mutants display moderate UV sensitivity reminiscent of rev mutants. However, unlike rev mutants, no defect in UV-induced reversion was seen in rad30 strains. While rad6 and rad18 are both epistatic to rad30, no epistasis was observed with rev1, rev3, rev7 or rad5, all of which are members of the RAD6 epistasis group. These findings suggest that RAD30 participates in a novel error-free repair pathway dependent on RAD6 and RAD18, but independent of REV1, REV3, REV7 and RAD5. PMID:9409821

  1. Pharmacogenetic Study in Rectal Cancer Patients Treated With Preoperative Chemoradiotherapy: Polymorphisms in Thymidylate Synthase, Epidermal Growth Factor Receptor, GSTP1, and DNA Repair Genes

    SciTech Connect

    Paez, David; Salazar, Juliana; Pare, Laia; Pertriz, Lourdes; Targarona, Eduardo; Rio, Elisabeth del; Barnadas, Agusti; Marcuello, Eugenio; Baiget, Montserrat

    2011-12-01

    Purpose: Several studies have been performed to evaluate the usefulness of neoadjuvant treatment using oxaliplatin and fluoropyrimidines for locally advanced rectal cancer. However, preoperative biomarkers of outcome are lacking. We studied the polymorphisms in thymidylate synthase, epidermal growth factor receptor, glutathione S-transferase pi 1 (GSTP1), and several DNA repair genes to evaluate their usefulness as pharmacogenetic markers in a cohort of 128 rectal cancer patients treated with preoperative chemoradiotherapy. Methods and Materials: Blood samples were obtained from 128 patients with Stage II-III rectal cancer. DNA was extracted from the peripheral blood nucleated cells, and the genotypes were analyzed by polymerase chain reaction amplification and automated sequencing techniques or using a 48.48 dynamic array on the BioMark system. The germline polymorphisms studied were thymidylate synthase, (VNTR/5 Prime UTR, 2R G>C single nucleotide polymorphism [SNP], 3R G>C SNP), epidermal growth factor receptor (Arg497Lys), GSTP1 (Ile105val), excision repair cross-complementing 1 (Asn118Asn, 8092C>A, 19716G>C), X-ray repair cross-complementing group 1 (XRCC1) (Arg194Trp, Arg280His, Arg399Gln), and xeroderma pigmentosum group D (Lys751Gln). The pathologic response, pathologic regression, progression-free survival, and overall survival were evaluated according to each genotype. Results: The Asterisk-Operator 3/ Asterisk-Operator 3 thymidylate synthase genotype was associated with a greater response rate (pathologic complete remission and microfoci residual tumor, 59% in Asterisk-Operator 3/ Asterisk-Operator 3 vs. 35% in Asterisk-Operator 2/ Asterisk-Operator 2 and Asterisk-Operator 2/ Asterisk-Operator 3; p = .013). For the thymidylate synthase genotype, the median progression-free survival was 103 months for the Asterisk-Operator 3/ Asterisk-Operator 3 patients and 84 months for the Asterisk-Operator 2/ Asterisk-Operator 2 and Asterisk-Operator 2/ Asterisk

  2. In vitro osteogenic induction of bone marrow stromal cells with encapsulated gene-modified bone marrow stromal cells and in vivo implantation for orbital bone repair.

    PubMed

    Deng, Yuan; Zhou, Huifang; Yan, Chenxi; Wang, Yefei; Xiao, Caiwen; Gu, Ping; Fan, Xianqun

    2014-07-01

    Osteogenic induction with either growth factors or genetic modification has limitations due to the short half-life and cost of the former, or safety concerns regarding the latter. The objective of this study was to employ a microcapsulation technique to separate genetically modified and nonmodified bone marrow stromal cells (BMSCs) to establish a cost-effective and biosafe osteogenic induction methodology with functional evaluation in vitro and in vivo in a canine model. Autologous BMSCs were isolated and transduced with adenoviral vectors containing either BMP-2 or vascular endothelial growth factor (VEGF) or were dual transduced followed by encapsulation in alginate microcapsules using an electrostatic bead generator. After cocultured with encapsulated cells, normal autologous BMSCs were analyzed for osteogenic differentiation and seeded onto tricalcium phosphate (TCP) scaffolds for in vivo implantation to repair orbital wall bone defects (12 mm in diameter) in a canine model. In vitro assays showed that the expression of the transduced genes was significantly upregulated, with significantly more transduced proteins released from the transduced cells compared with control cells. Importantly, examination of the BMSCs induced by soluble factors released from the encapsulated cells revealed a significant upregulation of expression of osteogenic markers Runx2, BSP, OPN, and OCN in dual-transduction or induction groups. In addition, dual transduction and induction resulted in the highest increase of alkaline phosphatase activity and mineralization compared with other experimental groups. In vivo assays using CT, micro-CT, and histology further supported the qPCR and western blot findings. In conclusion, encapsulation of genetically modified BMSCs was able to release a sufficient amount of BMP-2 and VEGF, which effectively induced osteogenic differentiation of normal-cultured BMSCs and demonstrated bone repair of the orbital wall defect after implantation with

  3. Activated RecA protein may induce expression of a gene that is not controlled by the LexA repressor and whose function is required for mutagenesis and repair of UV-irradiated bacteriophage lambda

    SciTech Connect

    Calsou, P.; Villaverde, A.; Defais, M.

    1987-10-01

    The activated form of the RecA protein (RecA) is known to be involved in the reactivation and mutagenesis of UV-irradiated bacteriophage lambda and in the expression of the SOS response in Escherichia coli K-12. The expression of the SOS response requires cleavage of the LexA repressor by RecA and the subsequent expression of LexA-controlled genes. The evidence presented here suggests that RecA induces the expression of a gene(s) that is not under LexA control and that is also necessary for maximal repair and mutagenesis of damaged phage. This conclusion is based on the chloramphenicol sensitivity of RecA -dependent repair and mutagenesis of damaged bacteriophage lambda in lexA(Def) hosts.

  4. DNA repair in Mycoplasma gallisepticum

    PubMed Central

    2013-01-01

    Background DNA repair is essential for the maintenance of genome stability in all living beings. Genome size as well as the repertoire and abundance of DNA repair components may vary among prokaryotic species. The bacteria of the Mollicutes class feature a small genome size, absence of a cell wall, and a parasitic lifestyle. A small number of genes make Mollicutes a good model for a “minimal cell” concept. Results In this work we studied the DNA repair system of Mycoplasma gallisepticum on genomic, transcriptional, and proteomic levels. We detected 18 out of 22 members of the DNA repair system on a protein level. We found that abundance of the respective mRNAs is less than one per cell. We studied transcriptional response of DNA repair genes of M. gallisepticum at stress conditions including heat, osmotic, peroxide stresses, tetracycline and ciprofloxacin treatment, stationary phase and heat stress in stationary phase. Conclusions Based on comparative genomic study, we determined that the DNA repair system M. gallisepticum includes a sufficient set of proteins to provide a cell with functional nucleotide and base excision repair and mismatch repair. We identified SOS-response in M. gallisepticum on ciprofloxacin, which is a known SOS-inducer, tetracycline and heat stress in the absence of established regulators. Heat stress was found to be the strongest SOS-inducer. We found that upon transition to stationary phase of culture growth transcription of DNA repair genes decreases dramatically. Heat stress does not induce SOS-response in a stationary phase. PMID:24148612

  5. Excision repair cross complementation group 1 is a chemotherapy-tolerating gene in cisplatin-based treatment for non-small cell lung cancer.

    PubMed

    Wang, Shoufeng; Pan, Hong; Liu, Desen; Mao, Naiquan; Zuo, Chuantian; Li, Li; Xie, Tong; Huang, Dingming; Huang, Yaoyuan; Pan, Qi; Yang, Li; Wu, Junwei

    2015-02-01

    This study aimed to evaluate the biological functions of excision repair cross complementation goup 1 (ERCC1) in cell proliferation, cell cycle, invasion and cisplatin response of non-small cell lung cancer (NSCLC) cells. Firstly, ERCC1 gene was successfully transfected into H1299 cells by gene cloning and transfection techniques. Then, cell proliferation was determined with the cell growth curve and colony-forming assays. Flow cytometry (FCM) was employed to investigate the cell cycle distribution. The ability of cell invasion was estimated by means of Matrigel invasion assays. Response of NSCLC cells to cisplatin was detected utilizing MTT assays, and the intracellular drug concentrations were determined by the high performance liquid chromatography (HPLC) analysis. Expression of the two cell membrane proteins, P-glycoprotein (P-gp) and multidrug resistance-associated protein (MRP), was also evaluated utilizing FCM technique. By contrast, ERCC1 expression in the NSCLC A549 cells was silenced by small interfering RNA (siRNA) through RNAi technique. In addition, the cytotoxic effect of cisplatin on A549 cells was detected by MTT assays. In the present study, the results demonstrated that ERCC1 had no effect on cell proliferation, cell cycle and the ability of invasion, but showed significant impact on cisplatin response of the NSCLC H1299 cells. Furthermore, siRNA-induced suppression of ERCC1 evidently enhanced sensitivity to cisplatin of NSCLC A549 cells. Therefore, it is confirmed that ERCC1 is a chemotherapy-tolerating gene and a promising predictor in tailoring chemotherapy of NSCLC. PMID:25434755

  6. Genomic alterations in DNA repair and chromatin remodeling genes in estrogen receptor-positive metastatic breast cancer patients with exceptional responses to capecitabine.

    PubMed

    Levin, Maren K; Wang, Kai; Yelensky, Roman; Cao, Ying; Ramos, Corinne; Hoke, Nicholas; Pippen, John; Blum, Joanne L; Brooks, Barry; Palmer, Gary; Palma, Norma; Balasubramanian, Sohail; Ross, Jeffrey S; O'Shaughnessy, Joyce

    2015-08-01

    We analyzed the genomic and phosphoproteomic profiles of breast cancer tissue obtained from six patients with estrogen receptor (ER)-positive, HER2-negative metastatic breast cancer who had highly durable (≥ 5 years) and, in some cases, ongoing clinical responses with capecitabine. Formalin-fixed, paraffin-embedded tissue samples from patients' primary (n = 4) or metastatic (n = 2) breast cancers were utilized for targeted next-generation sequencing and reversed phase protein microarray. Two patients received capecitabine monotherapy. Four patients received capecitabine in combination with paclitaxel; three of these continued single-agent capecitabine after stopping paclitaxel. Capecitabine was discontinued for progressive disease after a mean of 66 months in four patients (range 54-86 months), and two patients remain on therapy, having received capecitabine for >91 months and >122 months, respectively. Three patients' cancers (50%) had likely functional alterations in DNA repair and chromatin remodeling genes, while three other patients' cancers had variants of unknown significance in these pathways. Mutations in PIK3CA, amplifications of FGFR1 or ZNF703, or phosphorylation of HER family receptors and their downstream proteins did not preclude exceptional responses to capecitabine. None of the patients' tumors harbored TP53 or PTEN mutations. Four of the patients had breast cancer tissue available for PTEN immunohistochemistry, and all four patients' cancers were positive for PTEN. These surprising findings in a group of phenotypically similar patients with ER-positive, endocrine therapy-pretreated, HER2-negative metastases, are supported by preclinical data showing that sensitivity to 5-fluorouracil is enhanced by deficiencies in chromatin remodeling and homologous recombination genes. Our findings suggest that mutations that inactivate homologous recombination and/or chromatin remodeling genes within ER-positive, HER2-negative breast cancers may predict for

  7. “Same Day” Ex-vivo Regional Gene Therapy: A Novel Strategy to Enhance Bone Repair

    PubMed Central

    Virk, Mandeep S; Sugiyama, Osamu; Park, Sang H; Gambhir, Sanjiv S; Adams, Douglas J; Drissi, Hicham; Lieberman, Jay R

    2011-01-01

    Ex-vivo regional gene therapy with bone marrow cells (BMCs) overexpressing bone morphogenetic protein-2 (BMP-2) has demonstrated efficacy in healing critical sized bone defects in preclinical studies. The purpose of this preclinical study was to compare the osteoinductive potential of a novel “same day” ex-vivo regional gene therapy versus a traditional two-step approach, which involves culture expansion of the donor cells before implantation. In the “same day” strategy buffy coat cells were harvested from the rat bone marrow, transduced with a lentiviral vector-expressing BMP-2 for 1 hour and implanted into a rat femoral defect in the same sitting. There was no significant difference (P = 0.22) with respect to the radiographic healing rates between the femoral defects treated with the “same day” strategy (13/13; 100%) versus the traditional two-step approach (11/14; 78%). However, the femoral defects treated with the “same day” strategy induced earlier radiographic bone healing (P = 0.004) and higher bone volume (BV) [micro-computed tomography (micro-CT); P < 0.001]. The “same day” strategy represents a significant advance in the field of ex-vivo regional gene therapy because it offers a solution to limitations associated with the culture expansion process required in the traditional ex vivo approach. This strategy should be cost-effective when adapted for human use. PMID:21343916

  8. Tissue repair

    PubMed Central

    2010-01-01

    As living beings that encounter every kind of traumatic event from paper cut to myocardial infarction, we must possess ways to heal damaged tissues. While some animals are able to regrow complete body parts following injury (such as the earthworm who grows a new head following bisection), humans are sadly incapable of such feats. Our means of recovery following tissue damage consists largely of repair rather than pure regeneration. Thousands of times in our lives, a meticulously scripted but unseen wound healing drama plays, with cells serving as actors, extracellular matrix as the setting and growth factors as the means of communication. This article briefly reviews the cells involved in tissue repair, their signaling and proliferation mechanisms and the function of the extracellular matrix, then presents the actors and script for the three acts of the tissue repair drama. PMID:21220961

  9. Stem cells with FGF4-bFGF fused gene enhances the expression of bFGF and improves myocardial repair in rats

    SciTech Connect

    Chen, Xiang-Qi; Chen, Liang-Long Fan, Lin; Fang, Jun; Chen, Zhao-Yang; Li, Wei-Wei

    2014-04-25

    Highlights: • BFGF exists only in the cytoplasm of live cells. • BFGF cannot be secreted into the extracellular space to promote cell growth. • We combine the secretion-promoting signal peptide of FGF4. • We successfully modified BMSCs with the fused genes of FGF4-bFGF. • We promoted the therapeutic effects of transplanted BMSCs in myocardial infarction. - Abstract: The aim of this study was to investigate whether the modification of bone marrow-derived mesenchymal stem cells (BMSCs) with the fused FGF4 (fibroblast growth factor 4)-bFGF (basic fibroblast growth factor) gene could improve the expression and secretion of BFGF, and increase the efficacies in repairing infarcted myocardium. We used In-Fusion technique to construct recombinant lentiviral vectors containing the individual gene of bFGF, enhanced green fluorescent protein (EGFP), or genes of FGF4-bFGF and EGFP, and then transfected these lentiviruses into rat BMSCs. We conducted an in vitro experiment to compare the secretion of bFGF in BMSCs infected by these lentiviruses and also examined their therapeutic effects in the treatment of myocardial infraction in a rodent study. Sixty rats were tested in the following five conditions: Group-SHAM received only sham operation as controls; Group-AMI received only injection of placebo PBS buffer; Group-BMSC, Group-bFGF and Group-FGF4-bFGF received implantation of BMSCs with empty lentivirus, bFGF lentivirus, and FGF4-bFGF lentivirus, respectively. Our results found out that the transplanted FGF4-bFGF BMSCs had the highest survival rate, and also the highest myocardial expression of bFGF and microvascular density as evidenced by Western blotting and immunohistochemistry, respectively. As compared to other groups, the Group-FGF4-BFGF rats had the lowest myocardial fibrotic fraction, and the highest left ventricular ejection fraction. These results suggest that the modification of BMSCs with the FGF4-bFGF fused gene can not only increase the expression of

  10. Hereditary breast and ovarian cancer: new genes in confined pathways.

    PubMed

    Nielsen, Finn Cilius; van Overeem Hansen, Thomas; Sørensen, Claus Storgaard

    2016-09-01

    Genetic abnormalities in the DNA repair genes BRCA1 and BRCA2 predispose to hereditary breast and ovarian cancer (HBOC). However, only approximately 25% of cases of HBOC can be ascribed to BRCA1 and BRCA2 mutations. Recently, exome sequencing has uncovered substantial locus heterogeneity among affected families without BRCA1 or BRCA2 mutations. The new pathogenic variants are rare, posing challenges to estimation of risk attribution through patient cohorts. In this Review article, we examine HBOC genes, focusing on their role in genome maintenance, the possibilities for functional testing of putative causal variants and the clinical application of new HBOC genes in cancer risk management and treatment decision-making. PMID:27515922

  11. Timing of predisposing factors is important in necrotic enteritis models.

    PubMed

    Van Waeyenberghe, Lieven; De Gussem, Maarten; Verbeke, Joren; Dewaele, Isabelle; De Gussem, Jeroen

    2016-06-01

    Since the ban of antimicrobial growth promotors, the importance of necrotic enteritis in broilers increases. Reliable and reproducible infection models are required for pathogenesis studies and product screening. Two major predisposing factors in necrotic enteritis models are fishmeal supplementation to feed and Eimeria infection. However, many unsolved issues regarding these predisposing factors still exist. Therefore, the influence of timepoint of fishmeal administration (onset on day 8 or day 18), timing of coccidiosis challenge (day 15 or day 19) and strain of coccidiosis challenge (field strain vs. commercial vaccine) on the induction of necrotic enteritis lesions was investigated. The birds were inoculated with Clostridium perfringens three times per day for four consecutive days (day 17 until day 20) and were scored for the presence of necrotic enteritis on days 22, 23, 24, 25 and 26. Supplementation of the diet with fishmeal from day 8 onwards increased the likelihood of necrotic enteritis compared to supplementation from day 18 onwards. Birds challenged on day 19 with coccidiosis were more likely to have necrotic enteritis on scoring days 23 and 24 compared to birds challenged on day 15. Differences on other scoring days were less pronounced. Finally, the strain of coccidiosis challenge had little influence on the induction of necrotic enteritis. Findings of this study can help researchers to set up successful necrotic enteritis infection models. PMID:26927291

  12. Association of single nucleotide polymorphisms of DNA repair genes in NER pathway and susceptibility to pancreatic cancer

    PubMed Central

    Zhao, Fuli; Shang, Yuhong; Zeng, Chen; Gao, Dongdong; Li, Ke

    2015-01-01

    In our study, we conducted a case-control study to investigate the association of ERCC1, ERCC2, ERCC3, ERCC4, ERCC5, XPA, XPC and DDB2 gene polymorphisms in the risk of pancreatic cancer. Between May 2012 and May 2014, a total of 246 patients with who were newly diagnosed with histopathologically confirmed primary pancreatic cancer and 246 controls were selected into our study. Genotyping of ERCC1 rs3212986 and rs11615, ERCC2 rs13181, ERCC3 rs4150441, ERCC4 rs6498486, ERCC5 rs873601, XPA rs2808668, XPC rs2228000, XPC rs2228001 and DDB2 rs2029298 were analyzed using polymerase chain reaction (PCR) coupled with restriction fragment length polymorphism (RFLP). By conditional logistic regression analysis, individuals carrying with TT genotype of ERCC1 rs3212986 and GG genotype of ERCC2 rs13181 were associated with increased risk of pancreatic cancer when compared with wide-type genotype, and the adjusted ORs (95% CI) were 2.40 (1.29-4.52) and 2.27 (1.26-4.15), respectively. We found that individuals carrying with GT+TT genotype of ERCC1 rs3212986 and TG+GG genotype of ERCC2 rs1318 gene polymorphisms were correlated with higher risk of pancreatic cancer in smokers when compared with non-smokers, and the adjusted ORs (95% CI) were 1.89 (1.05-3.40) and 1.88 (1.06-3.34), respectively. In conclusion, our study suggests that ERCC1 rs3212986 and ERCC2 rs1318 gene polymorphisms contribute to the development of pancreatic cancer, especially in smokers. PMID:26617894

  13. Association of single nucleotide polymorphisms of DNA repair genes in NER pathway and susceptibility to pancreatic cancer.

    PubMed

    Zhao, Fuli; Shang, Yuhong; Zeng, Chen; Gao, Dongdong; Li, Ke

    2015-01-01

    In our study, we conducted a case-control study to investigate the association of ERCC1, ERCC2, ERCC3, ERCC4, ERCC5, XPA, XPC and DDB2 gene polymorphisms in the risk of pancreatic cancer. Between May 2012 and May 2014, a total of 246 patients with who were newly diagnosed with histopathologically confirmed primary pancreatic cancer and 246 controls were selected into our study. Genotyping of ERCC1 rs3212986 and rs11615, ERCC2 rs13181, ERCC3 rs4150441, ERCC4 rs6498486, ERCC5 rs873601, XPA rs2808668, XPC rs2228000, XPC rs2228001 and DDB2 rs2029298 were analyzed using polymerase chain reaction (PCR) coupled with restriction fragment length polymorphism (RFLP). By conditional logistic regression analysis, individuals carrying with TT genotype of ERCC1 rs3212986 and GG genotype of ERCC2 rs13181 were associated with increased risk of pancreatic cancer when compared with wide-type genotype, and the adjusted ORs (95% CI) were 2.40 (1.29-4.52) and 2.27 (1.26-4.15), respectively. We found that individuals carrying with GT+TT genotype of ERCC1 rs3212986 and TG+GG genotype of ERCC2 rs1318 gene polymorphisms were correlated with higher risk of pancreatic cancer in smokers when compared with non-smokers, and the adjusted ORs (95% CI) were 1.89 (1.05-3.40) and 1.88 (1.06-3.34), respectively. In conclusion, our study suggests that ERCC1 rs3212986 and ERCC2 rs1318 gene polymorphisms contribute to the development of pancreatic cancer, especially in smokers. PMID:26617894

  14. Genetic variation in GPBAR1 predisposes to quantitative changes in colonic transit and bile acid excretion

    PubMed Central

    Shin, Andrea; Busciglio, Irene; Carlson, Paula; Acosta, Andres; Bharucha, Adil E.; Burton, Duane; Lamsam, Jesse; Lueke, Alan; Donato, Leslie J.; Zinsmeister, Alan R.

    2014-01-01

    The pathobiology of irritable bowel syndrome (IBS) is multifaceted. We aimed to identify candidate genes predisposing to quantitative traits in IBS. In 30 healthy volunteers, 30 IBS-constipation, and 64 IBS-diarrhea patients, we measured bowel symptoms, bile acid (BA) synthesis (serum 7α-hydroxy-4-cholesten-3-one and FGF19), fecal BA and fat, colonic transit (CT by scintigraphy), and intestinal permeability (IP by 2-sugar excretion). We assessed associations of candidate genes controlling BA metabolism (KLB rs17618244 and FGFR4 rs351855), BA receptor (GPBAR1 rs11554825), serotonin (5-HT) reuptake (SLC6A4 through rs4795541 which encodes for the 44-bp insert in 5HTTLPR), or immune activation (TNFSF15 rs4263839) with three primary quantitative traits of interest: colonic transit, BA synthesis, and fecal BA excretion. There were significant associations between fecal BA and CT at 48 h (r = 0.43; P < 0.001) and IP (r = 0.23; P = 0.015). GPBAR1 genotype was associated with CT48 (P = 0.003) and total fecal BA [P = 0.030, false detection rate (FDR) P = 0.033]. Faster CT48 observed with both CC and TT GPBAR1 genotypes was due to significant interaction with G allele of KLB, which increases BA synthesis and excretion. Other univariate associations (P < 0.05, without FDR correction) observed between GPBAR1 and symptom phenotype and gas sensation ratings support the role of GPBAR1 receptor. Associations between SLC6A4 and stool consistency, ease of passage, postprandial colonic tone, and total fecal BA excretion provide data in support of future hypothesis-testing studies. Genetic control of GPBAR1 receptor predisposing to pathobiological mechanisms in IBS provides evidence from humans in support of the importance of GPBAR1 to colonic motor and secretory functions demonstrated in animal studies. PMID:25012842

  15. Subnormal albumin gene expression is associated with weight loss in immunodeficient/DNA-repair-deficient wasted mice

    SciTech Connect

    Libertin, C.R.; Weaver, P.; Woloschak, G.E. |; Mobarhan, S.

    1993-09-01

    Mice bearing the autosomal recessive mutation wst express a disease syndrome of immunodeficiency, neurologic dysfunction, and increased sensitivity to the killing effects of ionizing radiation. The mice were originally characterized as ``wasted`` because of their dramatic weight loss that begins at 21 days of age and progresses until death at 28-32 days of age. Because of the reported association between abnormal liver status and weight loss, we examined expression of a variety of liver-specific genes in wst/wst 10 mice relative to littermate (wst/{center_dot}) and parental strain (BCF{sub 1}) controls. Interestingly, the results revealed a greater than 67% reduction in albumin mRNA expression in livers derived from wst/wst mice relative to both controls. Expression of alpha-fetoprotein as well as a variety of other liver-specific genes (secretory component, metallothionein, cytochrome P{sub 1}450, transferrin receptor, tumor necrosis factor, and Ia antigen) was unaffected. These results suggest a relationship between low albumin expression and wasting syndromes in mice. In addition, we believe that our data suggest the wasted mouse as a unique model for subnormal albumin expression in humans.

  16. Outboard Repair.

    ERIC Educational Resources Information Center

    Hardway, Jack

    This consortium-developed instructor's manual for small engine repair (with focus on outboard motors) consists of the following nine instructional units: electrical remote control assembly, mechanical remote control assembly, tilt assemblies, exhaust housing, propeller and trim tabs, cooling system, mechanical gearcase, electrical gearcase, and…

  17. Snowmobile Repair.

    ERIC Educational Resources Information Center

    Helbling, Wayne

    This guide is designed to provide and/or improve instruction for occupational training in the area of snowmobile repair, and includes eight areas. Each area consists of one or more units of instruction, with each instructional unit including some or all of the following basic components: Performance objectives, suggested activities for teacher and…

  18. Motorcycle Repair.

    ERIC Educational Resources Information Center

    Hein, Jim; Bundy, Mike

    This motorcycle repair curriculum guide contains the following ten areas of study: brake systems, clutches, constant mesh transmissions, final drives, suspension, mechanical starting mechanisms, electrical systems, fuel systems, lubrication systems, and overhead camshafts. Each area consists of one or more units of instruction. Each instructional…

  19. Hydrocele repair

    MedlinePlus

    ... small surgical cut in the fold of the groin, and then drains the fluid. The sac (hydrocele) holding the fluid may be removed. The surgeon strengthens the muscle wall with stitches. This is called a hernia repair. Sometimes the surgeon uses a laparoscope to do ...

  20. Bladder exstrophy repair

    MedlinePlus

    Bladder birth defect repair; Everted bladder repair; Exposed bladder repair; Repair of bladder exstrophy ... in boys and is often linked to other birth defects. Surgery is necessary to: Allow the child to ...

  1. DNA Repair Gene (XRCC1) Polymorphism (Arg399Gln) Associated with Schizophrenia in South Indian Population: A Genotypic and Molecular Dynamics Study

    PubMed Central

    Sujitha, S. P.; Kumar, D. Thirumal; Doss, C. George Priya; Aavula, K.; Ramesh, R.; Lakshmanan, S.; Gunasekaran, S.; Anilkumar, G.

    2016-01-01

    This paper depicts the first report from an Indian population on the association between the variant Arg399Gln of XRCC1 locus in the DNA repair system and schizophrenia, the debilitating disease that affects 1% of the world population. Genotypic analysis of a total of 523 subjects (260 patients and 263 controls) revealed an overwhelming presence of Gln399Gln in the case subjects against the controls (P < 0.0068), indicating significant level of association of this nsSNP with schizophrenia; the Gln399 allele frequency was also perceptibly more in cases than in controls (p < 0.003; OR = 1.448). The results of the genotypic studies were further validated using pathogenicity and stability prediction analysis employing computational tools [I-Mutant Suite, iStable, PolyPhen2, SNAP, and PROVEAN], with a view toassess the magnitude of deleteriousness of the mutation. The pathogenicity analysis reveals that the nsSNP could be deleterious inasmuch as it could affect the functionality of the gene, and interfere with protein function. Molecular dynamics simulation of 60ns was performed using GROMACS to analyse structural change due to a mutation (Arg399Gln) that was never examined before. RMSD, RMSF, hydrogen bonds, radius of gyration and SASA analysis showedthe existence of asignificant difference between the native and the mutant protein. The present study gives astrong indication that the XRCC1 locus deserves serious attention, as it could be a potential candidatecontributing to the etio-pathogenesis of the disease. PMID:26824244

  2. Polymorphisms of DNA repair genes OGG1 and XPD and the risk of age-related cataract in Egyptians

    PubMed Central

    Gharib, Amal F.; Dabour, Sherif A.; Fouad, Rania A.

    2014-01-01

    Purpose To analyze the association of the polymorphisms of xeroderma pigmentosum complementation group D (XPD) and 8-oxoguanine glycosylase-1 (OGG1) genes with the risk of age-related cataract (ARC) in an Egyptian population. Methods This case-control study included 150 patients with ARC and 50 controls. Genotyping of XPD Asp312Asn was performed by amplification refractory mutation system PCR assay and genotyping of OGG1 Ser326Cys was carried out by PCR including confronting two-pair primers. Results The Asn/Asn genotype of XPD gene was significantly associated with increased risk of ARC (odds ratio [OR] = 2.74, 95% confidence interval [CI] = 1.01–7.43, p = 0.04) and cortical cataract (OR = 5.06, 95% CI = 1.70–15.05, p = 0.002). The Asn312 allele was significantly associated with an increased risk of ARC (OR = 1.75, 95% CI 1.06–2.89, p = 0.03) and cortical cataract (OR = 2.81, 95% CI = 1.56–5.08, p<0.001). The OGG1 Cys/Cys genotype frequency was significantly higher in ARC (OR = 4.13, 95% CI = 0.93–18.21, p = 0.04) and the Cys326 allele (OR = 1.85, 95% CI = 1.07–3.20, p = 0.03). Moreover, the Cys/Cys genotype of the OGG1 gene was significantly higher in cortical cataract (OR = 6.00, 95% CI = 1.24–28.99, p = 0.01) and the Cys326 allele was also significantly associated with cortical cataract (OR = 2.45, 95% CI = 1.30–4.63, p = 0.005). Conclusions The results suggest that the Asn/Asn genotype and Asn312 allele of XPD polymorphism, as well as the Cys/Cys genotype and Cys326 allele of the OGG1 polymorphism, may be associated with increased risk of the development of ARC, particularly the cortical type, in the Egyptian population. PMID:24868140

  3. Body mass index in early adulthood and colorectal cancer risk for carriers and non-carriers of germline mutations in DNA mismatch repair genes

    PubMed Central

    Win, A K; Dowty, J G; English, D R; Campbell, P T; Young, J P; Winship, I; Macrae, F A; Lipton, L; Parry, S; Young, G P; Buchanan, D D; Martínez, M E; Jacobs, E T; Ahnen, D J; Haile, R W; Casey, G; Baron, J A; Lindor, N M; Thibodeau, S N; Newcomb, P A; Potter, J D; Le Marchand, L; Gallinger, S; Hopper, J L; Jenkins, M A

    2011-01-01

    Background: Carriers of germline mutations in DNA mismatch repair (MMR) genes have a high risk of colorectal cancer (CRC), but the modifiers of this risk are not well established. We estimated an association between body mass index (BMI) in early adulthood and subsequent risk of CRC for carriers and, as a comparison, estimated the association for non-carriers. Methods: A weighted Cox regression was used to analyse height and weight at 20 years reported by 1324 carriers of MMR gene mutations (500 MLH1, 648 MSH2, 117 MSH6 and 59 PMS2) and 1219 non-carriers from the Colon Cancer Family Registry. Results: During 122 304 person-years of observation, we observed diagnoses of CRC for 659 carriers (50%) and 36 non-carriers (3%). For carriers, the risk of CRC increased by 30% for each 5 kg m–2 increment in BMI in early adulthood (hazard ratio, HR: 1.30; 95% confidence interval, CI: 1.08–1.58; P=0.01), and increased by 64% for non-carriers (HR: 1.64; 95% CI: 1.02–2.64; P=0.04) after adjusting for sex, country, cigarette smoking and alcohol drinking (and the MMR gene that was mutated in carriers). The difference in HRs for carriers and non-carriers was not statistically significant (P=0.50). For MLH1 and PMS2 (MutLα heterodimer) mutation carriers combined, the corresponding increase was 36% (HR: 1.36; 95% CI: 1.05–1.76; P=0.02). For MSH2 and MSH6 (MutSα heterodimer) mutation carriers combined, the HR was 1.26 (95% CI: 0.96–1.65; P=0.09). There was no significant difference between the HRs for MutLα and MutSα heterodimer carriers (P=0.56). Conclusion: Body mass index in early adulthood is positively associated with risk of CRC for MMR gene mutation carriers and non-carriers. PMID:21559014

  4. Turbine repair process, repaired coating, and repaired turbine component

    DOEpatents

    Das, Rupak; Delvaux, John McConnell; Garcia-Crespo, Andres Jose

    2015-11-03

    A turbine repair process, a repaired coating, and a repaired turbine component are disclosed. The turbine repair process includes providing a turbine component having a higher-pressure region and a lower-pressure region, introducing particles into the higher-pressure region, and at least partially repairing an opening between the higher-pressure region and the lower-pressure region with at least one of the particles to form a repaired turbine component. The repaired coating includes a silicon material, a ceramic matrix composite material, and a repaired region having the silicon material deposited on and surrounded by the ceramic matrix composite material. The repaired turbine component a ceramic matrix composite layer and a repaired region having silicon material deposited on and surrounded by the ceramic matrix composite material.

  5. Proteoglycans and cartilage repair.

    PubMed

    Ouzzine, Mohamed; Venkatesan, Narayanan; Fournel-Gigleux, Sylvie

    2012-01-01

    Repair of damaged articular cartilage in osteoarthritis (OA) is a clinical challenge. Because cartilage is an avascular and aneural tissue, normal mechanisms of tissue repair through recruitment of cells to the site of tissue destruction are not feasible. Proteoglycan (PG) depletion induced by the proinflammatory cytokine interleukin-1β, a principal mediator in OA, is a major factor in the onset and progression of joint destruction. Current symptomatic treatments of OA by anti-inflammatory drugs do not alter the progression of the disease. Various therapeutic strategies have been developed to antagonize the effect of proinflammatory cytokines. However, relatively few studies were conducted to stimulate anabolic activity, in an attempt to enhance cartilage repair. To this aim, a nonviral gene transfer strategy of glycosyltransferases responsible for PG synthesis has been developed and tested for its capacity to promote cartilage PG synthesis and deposition. Transfection of chondrocytes or cartilage explants by the expression vector for the glycosyltransferase β-1,3-glucuronosyltransferase-I (GlcAT-I) enhanced PG synthesis and deposition in the ECM by promoting the synthesis of chondroitin sulfate GAG chains of the cartilage matrix. This indicates that therapy mediated through GT gene delivery may constitute a new strategy for the treatment of OA. PMID:22252645

  6. Role of single nucleotide polymorphisms of DNA repair genes in susceptibility to pancreatic cancer in Chinese population.

    PubMed

    Ying, M F; Zhao, R

    2016-01-01

    We conducted a case-control study to investigate the role of ERCC1-ERCC5 gene polymorphisms in the risk of pancreatic cancer. This study included 195 patients who were newly diagnosed with histopathologically confirmed primary pancreatic cancer, and 254 controls were recruited from Sir Run Run Shaw Hospital, between January 2012 and December 2014. Genotyping of ERCC1 rs3212986 and rs11615, ERCC2 rs13181, ERCC3 rs4150441, ERCC4 rs6498486, and ERCC5 rs2094258 polymorphisms was carried out using polymerase chain reaction coupled with restriction fragment length polymorphism. Unconditional logistic regression analyses showed that the TT genotype of ERCC1 rs3212986 was associated with an increased risk of pancreatic cancer, and the OR (95%CI) was 2.26 (1.21-4.22). However, we did not find a significant association between ERCC1 rs11615, ERCC2 rs13181, ERCC3 rs4150441, ERCC4 rs6498486, and ERCC5 rs2094258 polymorphisms and risk of pancreatic cancer. In summary, we found that the presence of the ERCC1 rs3212986 polymorphism correlated with an increased risk of pancreatic cancer. PMID:27050953

  7. A Gene Gun-mediated Nonviral RNA trans-splicing Strategy for Col7a1 Repair.

    PubMed

    Peking, Patricia; Koller, Ulrich; Hainzl, Stefan; Kitzmueller, Sophie; Kocher, Thomas; Mayr, Elisabeth; Nyström, Alexander; Lener, Thomas; Reichelt, Julia; Bauer, Johann W; Murauer, Eva M

    2016-01-01

    RNA trans-splicing represents an auspicious option for the correction of genetic mutations at RNA level. Mutations within COL7A1 causing strong reduction or absence of type VII collagen are associated with the severe skin blistering disease dystrophic epidermolysis bullosa. The human COL7A1 mRNA constitutes a suitable target for this RNA therapy approach, as only a portion of the almost 9 kb transcript has to be delivered into the target cells. Here, we have proven the feasibility of 5' trans-splicing into the Col7a1 mRNA in vitro and in vivo. We designed a 5' RNA trans-splicing molecule, capable of replacing Col7a1 exons 1-15 and verified it in a fluorescence-based trans-splicing model system. Specific and efficient Col7a1 trans-splicing was confirmed in murine keratinocytes. To analyze trans-splicing in vivo, we used gene gun delivery of a minicircle expressing a FLAG-tagged 5' RNA trans-splicing molecule into the skin of wild-type mice. Histological and immunofluorescence analysis of bombarded skin sections revealed vector delivery and expression within dermis and epidermis. Furthermore, we have detected trans-spliced type VII collagen protein using FLAG-tag antibodies. In conclusion, we describe a novel in vivo nonviral RNA therapy approach to restore type VII collagen expression for causative treatment of dystrophic epidermolysis bullosa. PMID:26928235

  8. Subclinical Mastitis in Dairy Animals: Incidence, Economics, and Predisposing Factors

    PubMed Central

    Sinha, Mukesh Kr.; Thombare, N. N.; Mondal, Biswajit

    2014-01-01

    A study was conducted to assess the incidence and economics of subclinical form of bovine mastitis in Central Region of India. Daily milk records of 187 animals during three seasons were collected and subjected to analysis. The economic loss due to reduction in yield, clinical expenses, and additional resources used were quantified and aggregated. The losses due to mastitis in monetary terms were estimated to be INR1390 per lactation, among which around 49% was owing to loss of value from milk and 37% on account of veterinary expenses. Higher losses were observed in crossbred cows due to their high production potential that was affected during mastitis period. The cost of treating an animal was estimated to be INR509 which includes cost of medicine (31.10%) and services (5.47%). Inadequate sanitation, hygiene, and veterinary services were the main predisposing factors for incidence and spread of mastitis as perceived by the respondents. PMID:25093203

  9. Rumen conditions that predispose cattle to pasture bloat.

    PubMed

    Majak, W; Howarth, R E; Cheng, K J; Hall, J W

    1983-08-01

    Rumen contents from the dorsal sac were examined before alfalfa ingestion to determine factors that predispose cattle to pasture bloat. Chlorophyll concentration, buoyancy of particulate matter, and rates of gas production were significantly higher in cattle that subsequently bloated than in those that did not. Higher chlorophyll in bloat cases indicated accumulation of suspended chloroplast particles in the dorsal sac, perhaps due to increased buoyancy of the particulate matter. The higher fermentation rates (in the presence of glucose) suggested that the latent capacity for gas production was due to microbial colonization of suspended feed particles. Chlorophyll 4 h after feeding was also higher in bloated as compared to unbloated animals. In short, the microbial colonization and retention of particulate matter provided active inocula for promoting rapid legume digestion. Consequently, gas production was enhanced when feeding commenced, but the fermentation gases were trapped by the buoyant, frothy ingesta, resulting in the condition of pasture bloat. PMID:6619348

  10. Genes related to growth regulation, DNA repair and apoptosis in an oestrogen receptor-negative (MDA-231) versus an oestrogen receptor-positive (MCF-7) breast tumour cell line.

    PubMed

    Skog, Sven; He, Qimin; Khoshnoud, Reza; Fornander, Tommy; Rutqvist, Lars-Erik

    2004-01-01

    The molecular mechanism(s) behind the development of endocrine resistance in breast cancer remains controversial. Here, we compare the capability of oestrogen receptor (ER)-negative cells (MDA-231) versus ER-positive tamoxifen-sensitive cells (MCF-7) to handle DNA repair, transmit signals from damaged DNA, initiate cell death via apoptosis, and then to control transmitted signals from the cell cycle and to synthesize growth factors and receptors. Genes related to these events were studied by cDNA micro-array. Normal human breast cells (H2F) and human lymphoblastoid tumour cells (CEM) were used as controls. Of the 18 genes investigated, 10 genes showed differences in their expression between the cell types. The ER-negative cells showed higher expressions of BRCA1, BRCA2, cdc2, cyclin B1, cyclin D1, cyclin E, IGFBP-3, TGF-alpha, TGF beta 2 and a lower expression of TGF beta R1. No differences in the expressions of bax, bcl-2, p53, p21 and GADD45 were found between the two cell lines. We found that the ER-negative cells were characterized by: (1) a stimulated expression of growth factors and cell cycle regulation compounds, (2) improved DNA repair capacity, but (3) no change in DNA damage signals and apoptotic pathways. Improved DNA repair capacity of ER-negative cells would have a growth advantage over ER-positive tumours when receiving antitumour therapy. PMID:15192311

  11. Suicide gene approach using a dual-expression lentiviral vector to enhance the safety of ex vivo gene therapy for bone repair

    PubMed Central

    Alaee, F; Sugiyama, O; Virk, MS; Tang, H; Drissi, H; Lichtler, AC; Lieberman, JR

    2014-01-01

    ‘Ex vivo’ gene therapy using viral vectors to overexpress BMP-2 is shown to heal critical-sized bone defects in experimental animals. To increase its safety, we constructed a dual-expression lentiviral vector to overexpress BMP-2 or luciferase and an HSV1-tk analog, Δtk (LV-Δtk-T2A-BMP-2/Luc). We hypothesized that administering ganciclovir (GCV) will eliminate the transduced cells at the site of implantation. The vector-induced expression of BMP-2 and luciferase in a mouse stromal cell line (W-20-17 cells) and mouse bone marrow cells (MBMCs) was reduced by 50% compared with the single-gene vector. W-20-17 cells were more sensitive to GCV compared with MBMCs (90–95% cell death at 12 days with GCV at 1 µg ml−1 in MBMCs vs 90–95% cell death at 5 days by 0.1 µg ml−1 of GCV in W-20-17 cells). Implantation of LV-Δtk-T2A-BMP-2 transduced MBMCs healed a 2 mm femoral defect at 4 weeks. Early GCV treatment (days 0–14) postoperatively blocked bone formation confirming a biologic response. Delayed GCV treatment starting at day 14 for 2 or 4 weeks reduced the luciferase signal from LV-Δtk-T2A-Luc-transduced MBMCs, but the signal was not completely eliminated. These data suggest that this suicide gene strategy has potential for clinical use in the future, but will need to be optimized for increased efficiency. PMID:24285218

  12. Predisposing factors and surgical outcome of complicated liver hydatid cysts

    PubMed Central

    Akcan, Alper; Sozuer, Erdogan; Akyildiz, Hizir; Ozturk, Ahmet; Atalay, Altay; Yilmaz, Zeki

    2010-01-01

    AIM: To evaluate the predisposing factors for peritoneal perforation and intrabiliary rupture and the effects of these complications on surgical outcome in liver hydatid disease. METHODS: A total of 372 patients with liver hydatid cysts who had undergone surgical treatment were evaluated retrospectively. Twenty eight patients with peritoneal perforation, 93 patients with spontaneous intrabiliary perforation, and 251 patients with noncomplicated hydatid cysts were treated in our clinics. RESULTS: When the predisposing factors for complications were evaluated, younger age, superficial position, and larger cyst dimensions (P < 0.05; range, 0.001-0.017) increased peritoneal perforation rates. It was shown that older age increased cyst dimensions, and presence of multiple and bilobar cysts increased intrabiliary rupture rates (P < 0.05; range, 0.001-0.028). Partial pericystectomy and drainage was the most frequent surgical procedure in all groups (71.6%). The incidence of post-operative complications in the peritoneal perforated group, in the intrabiliary ruptured group, and in the noncomplicated group was 25%, 16.1% and 5.5%, respectively. When compared, complication rates were significantly different (P = 0.002). When length of hospital stay was compared, there was no significant difference between the groups (P > 0.05). The overall recurrence rate was 3.8% (14 patients), but there was not any statistical difference among the patient groups (P = 0.13). The early postoperative mortality rate was 1.1%. CONCLUSION: In peritoneally perforated and intrabiliary ruptured cases, the most important steps are irrigation of the peritoneal cavity and clearance of the cystic material from the biliary tree. PMID:20572308

  13. Varus gonarthrosis predisposes to varus malalignment in TKA.

    PubMed

    Heyse, Thomas J; Decking, Ralf; Davis, Jack; Boettner, Friedrich; Laskin, Richard S

    2009-09-01

    Postoperative alignment is a predictor for long-term survival of total knee arthroplasty (TKA). The purpose of this study was to evaluate whether or not preoperative deformities predispose to intraoperative malposition of TKA components. A retrospective radiographic analysis of 53 primary TKA cases was performed. Preoperative AP hip to ankle and lateral knee radiographs were compared with postoperative views to evaluate component positioning. The following angles were measured: the hip-knee-ankle (HKA) angle expressing the mechanical axis of the leg, the mechanical lateral distal femur angle (mLDFA), the medial proximal tibia angle (MPTA), the posterior distal femur angle (PDFA), and the posterior proximal tibia angle (PPTA). Postoperative measurement of the HKA revealed 34.0% of the cases had a deviation of >+/-3 degrees from neutral alignment. Sixteen knees (30.2%) were in varus and, with one exception, all presented with severe varus gonarthrosis prior to surgery with a mean tibiofemoral angle of 12.4 degrees compared with 1.0 degrees of valgus in the optimally aligned group. Patients (93.3%) with preoperative valgus malalignment showed optimal postoperative HKA. Odds ratios for malalignment of TKA for varus knees in comparison with valgus knees were 7.1 for HKA, 2.4 for MPTA, 4.9 for PDFA, and 1.7 for PPTA. The overall number of outliers in the presented data corresponds well with reports from other authors using different implants and guide systems. The presented data indicate that patients with preoperative varus alignment have a higher risk of postoperative implant malposition than patients with valgus alignment. The data supports that preoperative varus deformity predisposes to varus malposition of TKA. The risk for intraoperative malposition is significantly lower in valgus knees. PMID:19455367

  14. Meniscal Repair

    PubMed Central

    Yoon, Kyoung Ho

    2014-01-01

    The meniscus has several important roles, such as transmission of the load, absorption of the shock in the knee joint, acting as a secondary anteroposterior stabilizer of the knee joint, and contributing to proprioception of the knee joint. Degenerative changes of the knee joint develop in the long-term follow-up even after partial meniscectomy. Thus, there has been growing interest in meniscal repair. In addition, with increased understanding of the important roles of the meniscal root and advancement of diagnostic methods, efforts have been made to ensure preservation of the meniscal roots. In this review article, we will discuss operative techniques and clinical outcomes of arthroscopic repair of the meniscus and the meniscal root and postoperative rehabilitation and complications as well. PMID:24944971

  15. AB153. Down-regulated expression of excision of repair cross-complementing gene 1 reduces resistance to hydroxycamptothecine in bladder cancer

    PubMed Central

    Liu, Yili

    2016-01-01

    Objective The purpose of the study was to explore the potential mechanisms that interference of excision of repair cross-complementing gene 1 (ERCC1) mediated by lentiviral vector in bladder cancer T24 Cells. Methods the expression of ERCC1 was observed by immunohistochemical method in 25 cases of primary bladder cancer and recurrent bladder cancer tissues respectively from 25 patients. T24 cells were silenced targeting ERCC1 by lentiviruses .The transfection efficiency for ERCC1 was observed by fluorescence microscope and the interference efficiency was detected by real-time polymerase clain reaction and western blot assay. CCK-8 assay was used to assess the cell proliferation. Effects of cell apoptosis were detected by flow cytometry. Finally, the pathway of apoptosis was studied by using western blot method. Results As a result, we discovered that the expression level of ERCC1 in recurrent bladder cancer tissues (52%) was significantly higher than that in primary bladder cancer tissues (20%) (P<0.05). Compared with the T24 cells that did not silence the ERCC1 gene (control group) at different time periods (29.45%, 36.48%, 38.45%, 40.35%), the proliferation of T24 cells that silenced ERCC1 gene (experimental group) (27.25%, 37.45%, 32.5%, 42.05%) was not significantly changed (P>0.05). Hydroxycamptothecine (HCPT) inhibited the proliferation of T24 cells in dosage and time dependent manner. The inhibitory effect of HCPT on the experimental group was significantly higher than that of the control group (P<0.05).With the HCPT concentration increased, the apoptosis rate of the experimental group was significantly higher than that of the control group (P<0.05). After silencing of ERCC1, the sensitivity of T24 cells was increased to HCPT which could inhibit cell proliferation and induce cell apoptosis. Conclusions Therefore, ERCC1 may be a potential target protein used to guide the postoperative chemotherapy of bladder cancer.

  16. CDKN2A and BAP1 germline mutations predispose to melanoma and mesothelioma.

    PubMed

    Betti, M; Aspesi, A; Biasi, A; Casalone, E; Ferrante, D; Ogliara, P; Gironi, L C; Giorgione, R; Farinelli, P; Grosso, F; Libener, R; Rosato, S; Turchetti, D; Maffè, A; Casadio, C; Ascoli, V; Dianzani, C; Colombo, E; Piccolini, E; Pavesi, M; Miccoli, S; Mirabelli, D; Bracco, C; Righi, L; Boldorini, R; Papotti, M; Matullo, G; Magnani, C; Pasini, B; Dianzani, I

    2016-08-10

    BAP1 germline mutations predispose to a cancer predisposition syndrome that includes mesothelioma, cutaneous melanoma, uveal melanoma and other cancers. This co-occurrence suggests that these tumors share a common carcinogenic pathway. To evaluate this hypothesis, we studied 40 Italian families with mesothelioma and/or melanoma. The probands were sequenced for BAP1 and for the most common melanoma predisposition genes (i.e. CDKN2A, CDK4, TERT, MITF and POT1) to investigate if these genes may also confer susceptibility to mesothelioma. In two out of six families with both mesothelioma and melanoma we identified either a germline nonsense mutation (c.1153C > T, p.Arg385*) in BAP1 or a recurrent pathogenic germline mutation (c.301G > T, p.Gly101Trp) in CDKN2A. Our study suggests that CDKN2A, in addition to BAP1, could be involved in the melanoma and mesothelioma susceptibility, leading to the rare familial cancer syndromes. It also suggests that these tumors share key steps that drive carcinogenesis and that other genes may be involved in inherited predisposition to malignant mesothelioma and melanoma. PMID:27181379

  17. Haploinsufficiency of the ESCRT Component HD-PTP Predisposes to Cancer.

    PubMed

    Manteghi, Sanaz; Gingras, Marie-Claude; Kharitidi, Dmitri; Galarneau, Luc; Marques, Maud; Yan, Ming; Cencic, Regina; Robert, Francis; Paquet, Marilène; Witcher, Michael; Pelletier, Jerry; Pause, Arnim

    2016-05-31

    Endosomal sorting complexes required for transport (ESCRT) drive cell surface receptor degradation resulting in attenuation of oncogenic signaling and pointing to a tumor suppressor function. Here, we show that loss of function of an ESCRT protein (HD-PTP encoded by the PTPN23 gene, located on the tumor suppressor gene cluster 3p21.3) drives tumorigenesis in vivo. Indeed, Ptpn23(+/-) loss predisposes mice to sporadic lung adenoma, B cell lymphoma, and promotes Myc-driven lymphoma onset, dissemination, and aggressiveness. Ptpn23(+/-)-derived tumors exhibit an unaltered remaining allele and maintain 50% of HD-PTP expression. Consistent with the role of HD-PTP in attenuation of integrin recycling, cell migration, and invasion, hemizygous Ptpn23(+/-) loss increases integrin β1-dependent B cell lymphoma survival and dissemination. Finally, we reveal frequent PTPN23 deletion and downregulation in human tumors that correlates with poor survival. Altogether, we establish HD-PTP/PTPN23 as a prominent haploinsufficient tumor suppressor gene preventing tumor progression through control of integrin trafficking. PMID:27210750

  18. Germline BAP1 mutation predisposes to uveal melanoma, lung adenocarcinoma, meningioma, and other cancers

    PubMed Central

    Abdel-Rahman, Mohamed H; Pilarski, Robert; Cebulla, Colleen M; Massengill, James B; Christopher, Benjamin N; Boru, Getachew; Hovland, Peter; Davidorf, Frederick H

    2013-01-01

    Objective To investigate the potential contribution of germline sequence alterations in the BAP1 gene in uveal melanoma (UM) patients with possible predisposition to hereditary cancer. Design A total of 53 unrelated UM patients with high risk for hereditary cancer and five additional family members of one proband were studied. Mutational screening was carried out by direct sequencing. Results Of the 53 UM patients studied, a single patient was identified with a germline BAP1 truncating mutation, c. 799 C→T (p.Q267X), which segregated in several family members and was associated with UM and other cancers. Biallelic inactivation of BAP1 and decreased BAP1 expression were identified in the UM, lung adenocarcinoma and meningioma tumours from three family members with this germline BAP1 mutation. Germline BAP1 variants of uncertain significance, likely non-pathogenic, were also identified in two additional UM patients. Conclusion This study reports a novel hereditary cancer syndrome caused by a germline BAP1 mutation that predisposes patients to UM, lung carcinoma, meningioma, and possibly other cancers. The results indicate that BAP1 is the candidate gene in only a small subset of hereditary UM, suggesting the contribution of other candidate genes. PMID:21941004

  19. Nucleotide excision repair in humans.

    PubMed

    Spivak, Graciela

    2015-12-01

    The demonstration of DNA damage excision and repair replication by Setlow, Howard-Flanders, Hanawalt and their colleagues in the early 1960s, constituted the discovery of the ubiquitous pathway of nucleotide excision repair (NER). The serial steps in NER are similar in organisms from unicellular bacteria to complex mammals and plants, and involve recognition of lesions, adducts or structures that disrupt the DNA double helix, removal of a short oligonucleotide containing the offending lesion, synthesis of a repair patch copying the opposite undamaged strand, and ligation, to restore the DNA to its original form. The transcription-coupled repair (TCR) subpathway of NER, discovered nearly two decades later, is dedicated to the removal of lesions from the template DNA strands of actively transcribed genes. In this review I will outline the essential factors and complexes involved in NER in humans, and will comment on additional factors and metabolic processes that affect the efficiency of this important process. PMID:26388429

  20. DNA repair in mammalian embryos.

    PubMed

    Jaroudi, Souraya; SenGupta, Sioban

    2007-01-01

    Mammalian cells have developed complex mechanisms to identify DNA damage and activate the required response to maintain genome integrity. Those mechanisms include DNA damage detection, DNA repair, cell cycle arrest and apoptosis which operate together to protect the conceptus from DNA damage originating either in parental gametes or in the embryo's somatic cells. DNA repair in the newly fertilized preimplantation embryo is believed to rely entirely on the oocyte's machinery (mRNAs and proteins deposited and stored prior to ovulation). DNA repair genes have been shown to be expressed in the early stages of mammalian development. The survival of the embryo necessitates that the oocyte be sufficiently equipped with maternal stored products and that embryonic gene expression commences at the correct time. A Medline based literature search was performed using the keywords 'DNA repair' and 'embryo development' or 'gametogenesis' (publication dates between 1995 and 2006). Mammalian studies which investigated gene expression were selected. Further articles were acquired from the citations in the articles obtained from the preliminary Medline search. This paper reviews mammalian DNA repair from gametogenesis to preimplantation embryos to late gestational stages. PMID:17141556

  1. Association between codon 399 polymorphism in the X-ray repair cross-complementing group 1 gene and risk of prostate cancer in Asians: A study of 4,479 cases and 4,281 controls

    PubMed Central

    Yuanyuan, Mi; Xiaoming, You; Lijie, Zhu; Ninghan, Feng

    2015-01-01

    Objective: The polymorphism in codon 399 of the X-ray repair cross-complementing group 1 (XRCC1) gene may subtly alter structure of DNA repair enzymes and modulate the repair capacity. Impaired DNA repair can lead to the development of cancers such as prostate cancer (PCA). Although the association between the XRCC1 codon 399 polymorphism and PCA risk has been extensively reported, the results have been ambiguous. Methods: We conducted an updated analysis of 18 case–control studies to determine the association between the XRCC1 codon 399 polymorphism and PCA risk. We performed a literature search of the PubMed database to identify all eligible articles that reported this association. Odds ratios (ORs) with 95% confidence intervals (CI) were evaluated to assess the association. Results: Significant associations between PCA risk and XRCC1 codon 399 polymorphism were found (such as A-allele vs. G-allele: OR = 1.11, 95% CI = 1.01–1.23). Moreover, subgroup analysis based on ethnicity revealed similar significant associations in Asians (such as AA vs. GG: OR = 1.53, 95% CI = 1.19–1.97). Egger’s test did not reveal the presence of a publication bias. Conclusions: Our updated analysis provides evidence for significant association between XRCC1 codon 399 polymorphism and PCA risk. Further carefully designed studies should be performed. PMID:26649000

  2. Mutations in Haemophilus influenzae mismatch repair genes increase mutation rates of dinucleotide repeat tracts but not dinucleotide repeat-driven pilin phase variation rates.

    PubMed

    Bayliss, Christopher D; Sweetman, Wendy A; Moxon, E Richard

    2004-05-01

    High-frequency, reversible switches in expression of surface antigens, referred to as phase variation (PV), are characteristic of Haemophilus influenzae. PV enables this bacterial species, an obligate commensal and pathogen of the human upper respiratory tract, to adapt to changes in the host environment. Phase-variable hemagglutinating pili are expressed by many H. influenzae isolates. PV involves alterations in the number of 5' TA repeats located between the -10 and -35 promoter elements of the overlapping, divergently orientated promoters of hifA and hifBCDE, whose products mediate biosynthesis and assembly of pili. Dinucleotide repeat tracts are destabilized by mismatch repair (MMR) mutations in Escherichia coli. The influence of mutations in MMR genes of H. influenzae strain Rd on dinucleotide repeat-mediated PV rates was investigated by using reporter constructs containing 20 5' AT repeats. Mutations in mutS, mutL, and mutH elevated rates approximately 30-fold, while rates in dam and uvrD mutants were increased 14- and 3-fold, respectively. PV rates of constructs containing 10 to 12 5' AT repeats were significantly elevated in mutS mutants of H. influenzae strains Rd and Eagan. An intact hif locus was found in 14 and 12% of representative nontypeable H. influenzae isolates associated with either otitis media or carriage, respectively. Nine or more tandem 5' TA repeats were present in the promoter region. Surprisingly, inactivation of mutS in two serotype b H. influenzae strains did not alter pilin PV rates. Thus, although functionally analogous to the E. coli MMR pathway and active on dinucleotide repeat tracts, defects in H. influenzae MMR do not affect 5' TA-mediated pilin PV. PMID:15126452

  3. Influence of allelic Variations of hypoxia-related and DNA repair genes on patient outcome and toxicity in head and neck cancer treated with radiotherapy plus cetuximab.

    PubMed

    Muñoz, Carmen; Caballero, Miguel; Hakim, Sofia; Verger, Eugenia; Grau, Juan Jose

    2016-08-01

    Although cetuximab plus radiotherapy is a standard treatment for patients with inoperable head and neck squamous cell carcinoma (HNSCC), its efficacy varies greatly among individuals. To identify predictive markers of efficacy, we examined the effects of single nucleotide polymorphisms (SNPs) in hypoxia-related and DNA repair genes on the clinical outcome and occurrence of skin toxicity. We analyzed 61 consecutive patients with HNSCC for the presence of specific SNPs (HIF-1α, HIF-2α, HIF-1β, VHL, FIH-1, XRCC1, and XRCC5). The results were then correlated with time to progression (TTP), overall survival (OS), and toxicity (epithelitis, mucositis, and folliculitis). The median TTP and OS were better in patients with severe vs mild mucositis (17 vs 7 months, p = 0.03; and 26 vs 12 months, p = 0.016, respectively) and folliculitis (10 vs 7 months, p = 0.01, and 26 vs 10 months, p < 0.001, respectively). Patients with the HIF-1α CT/TT genotype had better OS than those with the wild-type HIF-1α CC genotype (28 vs 13 months, p = 0.035). Patients with the XRCC5 GG/AA genotype had longer TTP than patients with the XRCC5 AG genotype (11 vs 7 months, p = 0.035). Severe skin toxicity and SNPs of HIF-1α and XRCC5 were associated with different outcomes among patients treated with radiotherapy plus cetuximab. PMID:26245169

  4. Epigenetic reduction of DNA repair in progression to gastrointestinal cancer

    PubMed Central

    Bernstein, Carol; Bernstein, Harris

    2015-01-01

    Deficiencies in DNA repair due to inherited germ-line mutations in DNA repair genes cause increased risk of gastrointestinal (GI) cancer. In sporadic GI cancers, mutations in DNA repair genes are relatively rare. However, epigenetic alterations that reduce expression of DNA repair genes are frequent in sporadic GI cancers. These epigenetic reductions are also found in field defects that give rise to cancers. Reduced DNA repair likely allows excessive DNA damages to accumulate in somatic cells. Then either inaccurate translesion synthesis past the un-repaired DNA damages or error-prone DNA repair can cause mutations. Erroneous DNA repair can also cause epigenetic alterations (i.e., epimutations, transmitted through multiple replication cycles). Some of these mutations and epimutations may cause progression to cancer. Thus, deficient or absent DNA repair is likely an important underlying cause of cancer. Whole genome sequencing of GI cancers show that between thousands to hundreds of thousands of mutations occur in these cancers. Epimutations that reduce DNA repair gene expression and occur early in progression to GI cancers are a likely source of this high genomic instability. Cancer cells deficient in DNA repair are more vulnerable than normal cells to inactivation by DNA damaging agents. Thus, some of the most clinically effective chemotherapeutic agents in cancer treatment are DNA damaging agents, and their effectiveness often depends on deficient DNA repair in cancer cells. Recently, at least 18 DNA repair proteins, each active in one of six DNA repair pathways, were found to be subject to epigenetic reduction of expression in GI cancers. Different DNA repair pathways repair different types of DNA damage. Evaluation of which DNA repair pathway(s) are deficient in particular types of GI cancer and/or particular patients may prove useful in guiding choice of therapeutic agents in cancer therapy. PMID:25987950

  5. Predisposing factors of restless legs syndrome in pregnancy.

    PubMed

    Tunç, Tuğba; Karadağ, Yeşim Sücüllü; Doğulu, Funda; Inan, Levent E

    2007-04-15

    The occurrence of restless legs syndrome in pregnancy is well known. However, the mechanism of this association is unclear. In this study, we aimed to identify the factors that predispose women to have restless legs syndrome during pregnancy. A total of 146 pregnant women were included in the study. Patients were asked questions regarding demographic characteristics, complications of pregnancy, medical therapy (vitamin and iron intake), sleep disorders, muscle cramps, and excessive daytime sleepiness. Electroneurography, routine blood biochemistry tests, complete blood count, and thyroid function tests were performed and vitamin B12, folic acid, serum iron, iron-binding capacity, ferritin, iron saturation, prolactin, estradiol, and progesterone were measured. Of the participants, 38 were diagnosed as having restless legs syndrome. In women with restless legs syndrome, additional medical problems, night cramps, and excessive daytime sleepiness were more frequent. In women without restless legs syndrome, serum hemoglobin levels were significantly higher and the use of supplemental iron or vitamins was greater. Among the women with restless legs syndrome, progesterone levels were slightly higher but this difference was not statistically significant. In summary, in this study, lower hemoglobin levels and supplementation deficits of iron and vitamins were found be the risk factors for restless legs syndrome in pregnancy. PMID:17285614

  6. Nontuberculous mycobacteria: opportunistic environmental pathogens for predisposed hosts.

    PubMed

    Cook, James L

    2010-01-01

    Nontuberculous mycobacterial (NTM) infections are caused by environmental mycobacteria. Patients with pulmonary NTM disease usually have predisposing lung abnormalities. Diagnostic methods are evolving. Treatment is largely empiric. Data were extracted from peer reviewed publications, guidelines, and case series. Progressive NTM lung disease should be treated. Multidrug regimens are mostly macrolide based and are occasionally complemented by lung resection. Disease persistence and relapse are not uncommon and are a greater problem with so-called rapid-grower NTM infections. Some of the issues considered in this review are: the role of antibiotic susceptibility testing in predicting treatment effectiveness, optimal drug combinations, daily vs. intermittent dosing intervals for different NTM infections and disease severity, when the goal of cure should be replaced with observation or palliation, and patient selection for surgery. Future needs for development and research include improved epidemiology, definition of genetic and other risk factors, definition of predictors of treatment outcome, multicenter treatment studies, new drug discovery and animal models of disease and treatment. PMID:20977990

  7. FOXE3 mutations predispose to thoracic aortic aneurysms and dissections

    PubMed Central

    Kuang, Shao-Qing; Medina-Martinez, Olga; Guo, Dong-chuan; Gong, Limin; Regalado, Ellen S.; Reynolds, Corey L.; Boileau, Catherine; Jondeau, Guillaume; Prakash, Siddharth K.; Kwartler, Callie S.; Zhu, Lawrence Yang; Peters, Andrew M.; Duan, Xue-Yan; Bamshad, Michael J.; Shendure, Jay; Nickerson, Debbie A.; Santos-Cortez, Regie L.; Dong, Xiurong; Leal, Suzanne M.; Majesky, Mark W.; Swindell, Eric C.; Jamrich, Milan; Milewicz, Dianna M.

    2016-01-01

    The ascending thoracic aorta is designed to withstand biomechanical forces from pulsatile blood. Thoracic aortic aneurysms and acute aortic dissections (TAADs) occur as a result of genetically triggered defects in aortic structure and a dysfunctional response to these forces. Here, we describe mutations in the forkhead transcription factor FOXE3 that predispose mutation-bearing individuals to TAAD. We performed exome sequencing of a large family with multiple members with TAADs and identified a rare variant in FOXE3 with an altered amino acid in the DNA-binding domain (p.Asp153His) that segregated with disease in this family. Additional pathogenic FOXE3 variants were identified in unrelated TAAD families. In mice, Foxe3 deficiency reduced smooth muscle cell (SMC) density and impaired SMC differentiation in the ascending aorta. Foxe3 expression was induced in aortic SMCs after transverse aortic constriction, and Foxe3 deficiency increased SMC apoptosis and ascending aortic rupture with increased aortic pressure. These phenotypes were rescued by inhibiting p53 activity, either by administration of a p53 inhibitor (pifithrin-α), or by crossing Foxe3–/– mice with p53–/– mice. Our data demonstrate that FOXE3 mutations lead to a reduced number of aortic SMCs during development and increased SMC apoptosis in the ascending aorta in response to increased biomechanical forces, thus defining an additional molecular pathway that leads to familial thoracic aortic disease. PMID:26854927

  8. Ankle sprain: pathophysiology, predisposing factors, and management strategies

    PubMed Central

    Hubbard, Tricia J; Wikstrom, Erik A

    2010-01-01

    With the high percentage (up to 75%) of initial lateral ankle sprains (LAS) leading to repetitive sprains and chronic symptoms, it is imperative to better understand how best to treat and rehabilitate LAS events. The purpose of this paper is to review LAS pathophysiology, predisposing factors, and the current evidence regarding therapeutic modalities and exercises used in the treatment of LAS. Functional rehabilitation, early mobilization with support, is the current standard of care for LAS. However, the high percentage of reinjury occurrence and development of chronic symptoms (up to 75%) after a LAS, suggests the current standard of care may not be effective. Recent evidence has shown the need for more stringent immobilization to facilitate ligament healing and restoration of joint stability and function after a LAS. Additionally, the importance of adding adjunctive therapies, specifically joint mobilizations and balance training have been shown to improve function and decrease the incidence of reinjury after a LAS. Modifying current rehabilitation protocols to include protecting the ankle joint with stringent immobilization, and including joint mobilizations and balance training may be the first step to decreasing the incidence of short and long term ankle joint dysfunction. PMID:24198549

  9. Ankle sprain: pathophysiology, predisposing factors, and management strategies.

    PubMed

    Hubbard, Tricia J; Wikstrom, Erik A

    2010-01-01

    With the high percentage (up to 75%) of initial lateral ankle sprains (LAS) leading to repetitive sprains and chronic symptoms, it is imperative to better understand how best to treat and rehabilitate LAS events. The purpose of this paper is to review LAS pathophysiology, predisposing factors, and the current evidence regarding therapeutic modalities and exercises used in the treatment of LAS. Functional rehabilitation, early mobilization with support, is the current standard of care for LAS. However, the high percentage of reinjury occurrence and development of chronic symptoms (up to 75%) after a LAS, suggests the current standard of care may not be effective. Recent evidence has shown the need for more stringent immobilization to facilitate ligament healing and restoration of joint stability and function after a LAS. Additionally, the importance of adding adjunctive therapies, specifically joint mobilizations and balance training have been shown to improve function and decrease the incidence of reinjury after a LAS. Modifying current rehabilitation protocols to include protecting the ankle joint with stringent immobilization, and including joint mobilizations and balance training may be the first step to decreasing the incidence of short and long term ankle joint dysfunction. PMID:24198549

  10. Molecular regulation of UV-induced DNA repair.

    PubMed

    Shah, Palak; He, Yu-Ying

    2015-01-01

    Ultraviolet (UV) radiation from sunlight is a major etiologic factor for skin cancer, the most prevalent cancer in the United States, as well as premature skin aging. In particular, UVB radiation causes formation of specific DNA damage photoproducts between pyrimidine bases. These DNA damage photoproducts are repaired by a process called nucleotide excision repair, also known as UV-induced DNA repair. When left unrepaired, UVB-induced DNA damage leads to accumulation of mutations, predisposing people to carcinogenesis as well as to premature aging. Genetic loss of nucleotide excision repair leads to severe disorders, namely, xeroderma pigmentosum (XP), trichothiodystrophy (TTD) and Cockayne syndrome (CS), which are associated with predisposition to skin carcinogenesis at a young age as well as developmental and neurological conditions. Regulation of nucleotide excision repair is an attractive avenue to preventing or reversing these detrimental consequences of impaired nucleotide excision repair. Here, we review recent studies on molecular mechanisms regulating nucleotide excision repair by extracellular cues and intracellular signaling pathways, with a special focus on the molecular regulation of individual repair factors. PMID:25534312

  11. Molecular Regulation of UV-Induced DNA Repair

    PubMed Central

    Shah, Palak; He, Yu-Ying

    2014-01-01

    Ultraviolet (UV) radiation from sunlight is a major etiologic factor for skin cancer, the most prevalent cancer in the U.S., as well as premature skin aging. In particular, UVB radiation causes formation of specific DNA damage photoproducts between pyrimidine bases. These DNA damage photoproducts are repaired by a process called nucleotide excision repair, also known as UV-induced DNA repair. When left unrepaired, UVB-induced DNA damage leads to accumulation of mutations, predisposing people to carcinogenesis as well as to premature aging. Genetic loss of nucleotide excision repair leads to severe disorders, namely, xeroderma pigmentosum (XP), trichothiodystrophy (TTD) and Cockayne syndrome (CS), which are associated with predisposition to skin carcinogenesis at a young age as well as developmental and neurological conditions. Regulation of nucleotide excision repair is an attractive avenue to preventing or reversing these detrimental consequences of impaired nucleotide excision repair. Here we review recent studies on molecular mechanisms regulating nucleotide excision repair by extracellular cues and intracellular signaling pathways, with a special focus on the molecular regulation of individual repair factors. PMID:25534312

  12. Aortic aneurysm repair - endovascular

    MedlinePlus

    ... Endovascular aneurysm repair - aorta; AAA repair - endovascular; Repair - aortic aneurysm - endovascular ... leaking or bleeding. You may have an abdominal aortic aneurysm that is not causing any symptoms or problems. ...

  13. Eye muscle repair - discharge

    MedlinePlus

    ... Lazy eye repair - discharge; Strabismus repair - discharge; Extraocular muscle surgery - discharge ... You or your child had eye muscle repair surgery to correct eye muscle ... term for crossed eyes is strabismus. Children most often ...

  14. Brain aneurysm repair

    MedlinePlus

    ... aneurysm repair; Dissecting aneurysm repair; Endovascular aneurysm repair - brain; Subarachnoid hemorrhage - aneurysm ... Your scalp, skull, and the coverings of the brain are opened. A metal clip is placed at ...

  15. Aortic aneurysm repair - endovascular

    MedlinePlus

    EVAR; Endovascular aneurysm repair - aorta; AAA repair - endovascular; Repair - aortic aneurysm - endovascular ... leaking or bleeding. You may have an abdominal aortic aneurysm that is not causing any symptoms or problems. ...

  16. Impact of Polymorphic Variations of Gemcitabine Metabolism, DNA Damage Repair, and Drug-Resistance Genes on the Effect of High-Dose Chemotherapy for Relapsed or Refractory Lymphoid Malignancies.

    PubMed

    Shinozuka, Keiji; Tang, Hongwei; Jones, Roy B; Li, Donghui; Nieto, Yago

    2016-05-01

    The goal of this study was to determine whether single nucleotide polymorphisms (SNPs) in genes involved in gemcitabine metabolism, DNA damage repair, multidrug resistance, and alkylator detoxification influence the clinical outcome of patients with refractory/relapsed lymphoid malignancies receiving high-dose gemcitabine/busulfan/melphalan (Gem/Bu/Mel) with autologous stem cell support. We evaluated 21 germline SNPs of the gemcitabine metabolism genes CDA, deoxycytidine kinase, and hCNT3; DNA damage repair genes RECQL, X-ray repair complementing 1, RAD54L, ATM, ATR, MLH1, MSH2, MSH3, TREX1, EXO1, and TP73; and multidrug-resistance genes MRP2 and MRP5; as well as glutathione-S-transferase GSTP1 in 153 patients with relapsed or refractory lymphoma or myeloma receiving Gem/Bu/Mel. We studied the association of genotypes with overall survival (OS), progression-free survival (PFS), and nonhematological grade 3 or 4 toxicity. CDA C111T and TREX1 Ex14-460C>T genotypes had a significant effect on OS (P = .007 and P = .005, respectively), and CDA C111T, ATR C340T, and EXO1 P757L genotypes were significant predictors for severe toxicity (P = .037, P = .024, and P = .025, respectively) in multivariable models that adjusted for clinical variables. The multi-SNP risk score analysis identified the combined genotypes of TREX1 Ex14-460 TT and hCNT3 Ex5 +25A>G AA as significant predictors for OS and the combination of MRP2 Ex10 + 40GG/GA and MLH1 IVS12-169 TT as significant predictor for PFS. Polymorphic variants of certain genes involved in gemcitabine metabolism and DNA damage repair pathways may be potential biomarkers for clinical outcome in patients with refractory/relapsed lymphoid tumors receiving Gem/Bu/Mel. PMID:26743341

  17. DNA double-strand break repair inhibitors as cancer therapeutics.

    PubMed

    Srivastava, Mrinal; Raghavan, Sathees C

    2015-01-22

    Among DNA damages, double-strand breaks (DSBs) are one of the most harmful lesions to a cell. Failure in DSB repair could lead to genomic instability and cancer. Homologous recombination (HR) and nonhomologous end joining (NHEJ) are major DSB repair pathways in higher eukaryotes. It is known that expression of DSB repair genes is altered in various cancers. Activation of DSB repair genes is one of the reasons for chemo- and radioresistance. Therefore, targeting DSB repair is an attractive strategy to eliminate cancer. Besides, therapeutic agents introduce breaks in the genome as an intermediate. Therefore, blocking the residual repair using inhibitors can potentiate the efficacy of cancer treatment. In this review, we discuss the importance of targeting DSB repair pathways for the treatment of cancer. Recent advances in the development of DSB repair inhibitors and their clinical relevance are also addressed. PMID:25579208

  18. INTERNAL REPAIR OF PIPELINES

    SciTech Connect

    Robin Gordon; Bill Bruce; Nancy Porter; Mike Sullivan; Chris Neary

    2003-05-01

    The two broad categories of deposited weld metal repair and fiber-reinforced composite repair technologies were reviewed for potential application for internal repair of gas transmission pipelines. Both are used to some extent for other applications and could be further developed for internal, local, structural repair of gas transmission pipelines. Preliminary test programs were developed for both deposited weld metal repairs and for fiber-reinforced composite repair. To date, all of the experimental work pertaining to the evaluation of potential repair methods has focused on fiber-reinforced composite repairs. Hydrostatic testing was also conducted on four pipeline sections with simulated corrosion damage: two with composite liners and two without.

  19. Stimulating endogenous cardiac repair

    PubMed Central

    Finan, Amanda; Richard, Sylvain

    2015-01-01

    The healthy adult heart has a low turnover of cardiac myocytes. The renewal capacity, however, is augmented after cardiac injury. Participants in cardiac regeneration include cardiac myocytes themselves, cardiac progenitor cells, and peripheral stem cells, particularly from the bone marrow compartment. Cardiac progenitor cells and bone marrow stem cells are augmented after cardiac injury, migrate to the myocardium, and support regeneration. Depletion studies of these populations have demonstrated their necessary role in cardiac repair. However, the potential of these cells to completely regenerate the heart is limited. Efforts are now being focused on ways to augment these natural pathways to improve cardiac healing, primarily after ischemic injury but in other cardiac pathologies as well. Cell and gene therapy or pharmacological interventions are proposed mechanisms. Cell therapy has demonstrated modest results and has passed into clinical trials. However, the beneficial effects of cell therapy have primarily been their ability to produce paracrine effects on the cardiac tissue and recruit endogenous stem cell populations as opposed to direct cardiac regeneration. Gene therapy efforts have focused on prolonging or reactivating natural signaling pathways. Positive results have been demonstrated to activate the endogenous stem cell populations and are currently being tested in clinical trials. A potential new avenue may be to refine pharmacological treatments that are currently in place in the clinic. Evidence is mounting that drugs such as statins or beta blockers may alter endogenous stem cell activity. Understanding the effects of these drugs on stem cell repair while keeping in mind their primary function may strike a balance in myocardial healing. To maximize endogenous cardiac regeneration, a combination of these approaches could ameliorate the overall repair process to incorporate the participation of multiple cellular players. PMID:26484341

  20. Acute Normal Tissue Reactions in Head-and-Neck Cancer Patients Treated With IMRT: Influence of Dose and Association With Genetic Polymorphisms in DNA DSB Repair Genes

    SciTech Connect

    Werbrouck, Joke Ruyck, Kim de; Duprez, Frederic; Veldeman, Liv; Claes, Kathleen; Eijkeren, Marc van; Boterberg, Tom; Willems, Petra; Vral, Anne; Neve, Wilfried de; Thierens, Hubert

    2009-03-15

    Purpose: To investigate the association between dose-related parameters and polymorphisms in DNA DSB repair genes XRCC3 (c.-1843A>G, c.562-14A>G, c.722C>T), Rad51 (c.-3429G>C, c.-3392G>T), Lig4 (c.26C>T, c.1704T>C), Ku70 (c.-1310C>G), and Ku80 (c.2110-2408G>A) and the occurrence of acute reactions after radiotherapy. Materials and Methods: The study population consisted of 88 intensity-modulated radiation therapy (IMRT)-treated head-and-neck cancer patients. Mucositis, dermatitis, and dysphagia were scored using the Common Terminology Criteria (CTC) for Adverse Events v.3.0 scale. The population was divided into a CTC0-2 and CTC3+ group for the analysis of each acute effect. The influence of the dose on critical structures was analyzed using dose-volume histograms. Genotypes were determined by polymerase chain reaction (PCR) combined with restriction fragment length polymorphism or PCR-single base extension assays. Results: The mean dose (D{sub mean}) to the oral cavity and constrictor pharyngeus (PC) muscles was significantly associated with the development of mucositis and dysphagia, respectively. These parameters were considered confounding factors in the radiogenomics analyses. The XRCC3c.722CT/TT and Ku70c.-1310CG/GG genotypes were significantly associated with the development of severe dysphagia (CTC3+). No association was found between the investigated polymorphisms and the development of mucositis or dermatitis. A risk analysis model for severe dysphagia, which was developed based on the XRCC3c.722CT/TT and Ku70c.-1310CG/GG genotypes and the PC dose, showed a sensitivity of 78.6% and a specificity of 77.6%. Conclusions: The XRCC3c.722C>T and Ku70c.-1310C>G polymorphisms as well as the D{sub mean} to the PC muscles were highly associated with the development of severe dysphagia after IMRT. The prediction model developed using these parameters showed a high sensitivity and specificity.

  1. Somatic Overgrowth Predisposes to Seizures in Autism Spectrum Disorders

    PubMed Central

    Brachini, Francesca; Apicella, Fabio; Cosenza, Angela; Ferrari, Anna Rita; Guerrini, Renzo; Muratori, Filippo; Romano, Maria Francesca; Santorelli, Filippo M.; Tancredi, Raffaella; Sicca, Federico

    2013-01-01

    Background Comorbidity of Autism Spectrum Disorders with seizures or abnormal EEG (Autism-Epilepsy Phenotype) suggests shared pathomechanisms, and might be a starting point to identify distinct populations within the clinical complexity of the autistic spectrum. In this study, we tried to assess whether distinct subgroups, having distinctive clinical hallmarks, emerge from this comorbid condition. Methods Two-hundred and six individuals with idiopathic Autism Spectrum Disorders were subgrouped into three experimental classes depending on the presence of seizures and EEG abnormalities. Neurobehavioral, electroclinical and auxological parameters were investigated to identify differences among groups and features which increase the risk of seizures. Our statistical analyses used ANOVA, post-hoc multiple comparisons, and the Chi-squared test to analyze continuous and categorical variables. A correspondence analysis was also used to decompose significant Chi-squared and reduce variables dimensions. Results The high percentage of children with seizures (28.2% of our whole cohort) and EEG abnormalities (64.1%) confirmed that the prevalence of epilepsy in Autism Spectrum Disorders exceeds that of the general population. Seizures were associated with severe intellectual disability, and not with autism severity. Interestingly, tall stature (without macrocephaly) was significantly associated with EEG abnormalities or later onset seizures. However, isolated macrocephaly was equally distributed among groups or associated with early onset seizures when accompanied by tall stature. Conclusions Tall stature seems to be a phenotypic “biomarker” of susceptibility to EEG abnormalities or late epilepsy in Autism Spectrum Disorders and, when concurring with macrocephaly, predisposes to early onset seizures. Growth pattern might act as an endophenotypic marker in Autism-Epilepsy comorbidity, delineating distinct pathophysiological subtypes and addressing personalized diagnostic work

  2. DNA Repair Dysfunction and Neurodegeneration: Lessons From Rare Pediatric Disorders.

    PubMed

    Shabbir, Syed H

    2016-03-01

    Nucleotide excision repair disorders display a wide range of clinical syndromes and presentations, all associated at the molecular level by dysfunction of genes participating in the nucleotide excision repair pathway. Genotype-phenotype relationships are remarkably complex and not well understood. This article outlines neurodegenerative symptoms seen in nucleotide excision repair disorders and explores the role that nucleotide excision repair dysfunction can play in the pathogenesis of chronic neurodegenerative diseases. PMID:26116382

  3. [Studies of the repair of radiation-induced genetic damage in Drosophila]. Annual progress report, September 1, 1990--July 1, 1991

    SciTech Connect

    1991-12-31

    Research is focused on the following areas: characterization of DNA double-strand break repair; using injected oligonucleotides as templates to repair double-strand DNA breaks; analysis of a gene required for postreplication repair; cloning of a gene required for resistance to DNA cross-linking agents; cloning of a gene required for excision repair; cloning of a gene required for X-ray resistance; and transposon tagging DNA repair genes.

  4. Laryngeal cancer risk associated with smoking and alcohol consumption is modified by genetic polymorphisms in ERCC5, ERCC6 and RAD23B but not by polymorphisms in five other nucleotide excision repair genes.

    PubMed

    Abbasi, Rashda; Ramroth, Heribert; Becher, Heiko; Dietz, Andreas; Schmezer, Peter; Popanda, Odilia

    2009-09-15

    Laryngeal cancer is known to be associated with smoking and high alcohol consumption. Nucleotide excision repair (NER) plays a key role in repairing DNA damage induced by these exposures and might affect laryngeal cancer susceptibility. In a population-based case-control study including 248 cases and 647 controls, the association of laryngeal cancer with 14 single nucleotide polymorphisms (SNPs) in 8 NER genes (XPC, XPA, ERCC1, ERCC2, ERCC4, ERCC5, ERCC6 and RAD23B) was analyzed with respect to smoking and alcohol exposure. For genotyping, sequence specific hybridization probes were used. Data were evaluated by conditional logistic regression analysis, stratified for age and gender, and adjusted for smoking, alcohol consumption and education. Pro-carriers of ERCC6 Arg1230Pro showed a decreased risk for laryngeal cancer (OR = 0.53, 95% CI 0.34-0.85), strongest in heavy smokers and high alcohol consumers. ERCC5 Asp1104His was associated with risk in heavy smokers (OR = 1.70, 95% CI 1.1-2.5). Val-carriers of RAD23B Ala249Val had an increased cancer risk in heavy smokers (OR = 1.6, 95% CI 1.1-2.5) and high alcohol consumers (OR = 2.0, 95% CI 1.1-3.4). The combined effect of smoking and alcohol intake affected risk, at high exposure level, for ERCC6 1230Pro carriers (OR = 0.47, 95% CI 0.22-0.98) and RAD23B 249Val carriers (OR = 2.6, 95% CI 1.3-4.9). When tested for gene-gene interaction, presence of 3 risk alleles in the XPC-RAD23B complex increased the risk 2.1-fold. SNPs in the other genes did not show a significant association with laryngeal cancer risk. We conclude that common genetic variations in NER genes can significantly modify laryngeal cancer risk. PMID:19444904

  5. An autosomal locus predisposing to multiple deletions of mtDNA on chromosome 3p

    SciTech Connect

    Kaukonen, J.A.; Suomalainen, A.; Peltonen, L.; Amati, P.; Zeviani, M.

    1996-04-01

    Autosomal dominant progressive external ophthalmoplegia (adPEO) is a disorder characterized by ptosis, progressive weakness of the external eye muscles, and general muscle weakness. The patients have multiple deletions of mtDNA on Southern blots or in PCR analysis of muscle DNA and a mild deficiency of one or more respiratory-chain enzymes carrying mtDNA-encoded subunits. The pattern of inheritance indicates a nuclear gene defect predisposing to secondary mtDNA deletions. Recently, in one Finnish family, we assigned an adPEO locus to chromosome 10q23.3-24.3 but also excluded linkage to this same locus in two Italian adPEO families with a phenotype closely resembling the Finnish one. We applied a random mapping approach to informative non-10q-linked Italian families to assign the second locus for adPEO and found strong evidence for linkage on chromosome 3p14.1-21.2 in three Italian families, with a maximum two-point lod score of 4.62 at a recombination fraction of .0. However, in three additional families, linkage to the same chromosomal region was clearly absent, indicating further genetic complexity of the adPEO trait. 19 refs., 3 figs., 2 tabs.

  6. Relationship of the FKBP5 C/T polymorphism with dysfunctional attitudes predisposing to depression.

    PubMed

    Suzuki, Akihito; Matsumoto, Yoshihiko; Sadahiro, Ryoichi; Enokido, Masanori; Goto, Kaoru; Otani, Koichi

    2014-08-01

    FK506-binding protein 51 (FKBP5) is a co-chaperone of the glucocorticoid receptor, and plays an important role in the negative feedback regulation of the hypothalamic-pituitary-adrenal axis. The C/T single nucleotide polymorphism in the intron 2 of the FKBP5 gene affects cortisol secretion, and has been implicated in the pathophysiology of depression. In this study, the relationship of the FKBP5 C/T polymorphism with dysfunctional attitudes predisposing to depression was examined. The subjects were 300 healthy Japanese. The FKBP5 genotypes were determined by a real-time PCR and cycling probe technology for SNP typing. Dysfunctional attitudes were assessed by the 24-item version of the Dysfunctional Attitude Scale (DAS-24), which has the Achievement, Self-control, and Dependency subscales. DAS-24 total scores were significantly higher in the group with the T allele than in that without this allele (p=0.001). Regarding the subscales, scores of the Achievement (p=0.003) and Self-control (p=0.009) subscales, but not those of the Dependency subscale, were significantly higher in the former group than in the latter group. The present study suggests that the FKBP5 C/T polymorphism is implicated in formation of dysfunctional attitudes, especially those about achievement and self-control. PMID:24889341

  7. Genome-wide association study identifies novel loci predisposing to cutaneous melanoma.

    PubMed

    Amos, Christopher I; Wang, Li-E; Lee, Jeffrey E; Gershenwald, Jeffrey E; Chen, Wei V; Fang, Shenying; Kosoy, Roman; Zhang, Mingfeng; Qureshi, Abrar A; Vattathil, Selina; Schacherer, Christopher W; Gardner, Julie M; Wang, Yuling; Bishop, D Tim; Barrett, Jennifer H; MacGregor, Stuart; Hayward, Nicholas K; Martin, Nicholas G; Duffy, David L; Mann, Graham J; Cust, Anne; Hopper, John; Brown, Kevin M; Grimm, Elizabeth A; Xu, Yaji; Han, Younghun; Jing, Kaiyan; McHugh, Caitlin; Laurie, Cathy C; Doheny, Kim F; Pugh, Elizabeth W; Seldin, Michael F; Han, Jiali; Wei, Qingyi

    2011-12-15

    We performed a multistage genome-wide association study of melanoma. In a discovery cohort of 1804 melanoma