Reduced levels of prostaglandin I2 synthase: a distinctive feature of the cancer-free trichothiodystrophy

The cancer-free photosensitive trichothiodystrophy (PS-TTD) and the cancer-prone xeroderma pigmentosum (XP) are rare monogenic disorders that can arise from mutations in the same genes, namely ERCC2/XPD or ERCC3/XPB. Both XPD and XPB proteins belong to the 10-subunit complex transcription factor IIH (TFIIH) that plays a key role in transcription and nucleotide excision repair, the DNA repair pathway devoted to the removal of ultraviolet-induced DNA lesions. Compelling evidence suggests that mutations affecting the DNA repair activity of TFIIH are responsible for the pathological features of XP, whereas those also impairing transcription give rise to TTD. By adopting a relatives-based whole transcriptome sequencing approach followed by specific gene expression profiling in primary fibroblasts from a large cohort of TTD or XP cases with mutations in ERCC2/XPD gene, we identify the expression alterations specific for TTD primary dermal fibroblasts. While most of these transcription deregulations do not impact on the protein level, very low amounts of prostaglandin I2 synthase (PTGIS) are found in TTD cells. PTGIS catalyzes the last step of prostaglandin I2 synthesis, a potent vasodilator and inhibitor of platelet aggregation. Its reduction characterizes all TTD cases so far investigated, both the PS-TTD with mutations in TFIIH coding genes as well as the nonphotosensitive (NPS)-TTD. A severe impairment of TFIIH and RNA polymerase II recruitment on the PTGIS promoter is found in TTD but not in XP cells. Thus, PTGIS represents a biomarker that combines all PS- and NPS-TTD cases and distinguishes them from XP.

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Mutational signatures are jointly shaped by DNA damage and repair

10.1101/686295 ◽

2019 ◽

Cited By ~ 2

Author(s):

Nadezda V Volkova ◽

Bettina Meier ◽

Víctor González-Huici ◽

Simone Bertolini ◽

Santiago Gonzalez ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Excision Repair ◽

Dna Lesions ◽

Single Nucleotide Variants ◽

Mutational Signatures ◽

Experimental Conditions ◽

Repair Deficiency ◽

Dna Repair Deficiency ◽

Mutation Spectra

AbstractMutations arise when DNA lesions escape DNA repair. To delineate the contributions of DNA damage and DNA repair deficiency to mutagenesis we sequenced 2,717 genomes of wild-type and 53 DNA repair defective C. elegans strains propagated through several generations or exposed to 11 genotoxins at multiple doses. Combining genotoxin exposure and DNA repair deficiency alters mutation rates or leads to unexpected mutation spectra in nearly 40% of all experimental conditions involving 9/11 of genotoxins tested and 32/53 genotypes. For 8/11 genotoxins, signatures change in response to more than one DNA repair deficiency, indicating that multiple genes and pathways are involved in repairing DNA lesions induced by one genotoxin. For many genotoxins, the majority of observed single nucleotide variants results from error-prone translesion synthesis, rather than primary mutagenicity of altered nucleotides. Nucleotide excision repair mends the vast majority of genotoxic lesions, preventing up to 99% of mutations. Analogous mutagenic DNA damage-repair interactions can also be found in cancers, but, except for rare cases, effects are weak owing to the unknown histories of genotoxic exposures and DNA repair status. Overall, our data underscore that mutation spectra are joint products of DNA damage and DNA repair and imply that mutational signatures computationally derived from cancer genomes are more variable than currently anticipated.

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Structural basis of TFIIH activation for nucleotide excision repair

10.1101/628032 ◽

2019 ◽

Author(s):

Goran Kokic ◽

Aleksandar Chernev ◽

Dimitry Tegunov ◽

Christian Dienemann ◽

Henning Urlaub ◽

...

Keyword(s):

Dna Repair ◽

Transcription Factor ◽

Nucleotide Excision Repair ◽

Excision Repair ◽

Dna Lesions ◽

Structural Basis ◽

Disease Mutations ◽

Mechanistic Analysis ◽

Nucleotide Excision ◽

Dna Complex

AbstractGenomes are constantly threatened by DNA damage, but cells can remove a large variety of DNA lesions by nucleotide excision repair (NER)1. Mutations in NER factors compromise cellular fitness and cause human diseases such as Xeroderma pigmentosum (XP), Cockayne syndrome and trichothiodystrophy2,3. The NER machinery is built around the multisubunit transcription factor IIH (TFIIH), which opens the DNA repair bubble, scans for the lesion, and coordinates excision of the damaged DNA single strand fragment1,4. TFIIH consists of a kinase module and a core module that contains the ATPases XPB and XPD5. Here we prepare recombinant human TFIIH and show that XPB and XPD are stimulated by the additional NER factors XPA and XPG, respectively. We then determine the cryo-electron microscopy structure of the human core TFIIH-XPA-DNA complex at 3.6 Å resolution. The structure represents the lesion-scanning intermediate on the NER pathway and rationalizes the distinct phenotypes of disease mutations. It reveals that XPB and XPD bind double- and single-stranded DNA, respectively, consistent with their translocase and helicase activities. XPA forms a bridge between XPB and XPD, and retains the DNA at the 5’-edge of the repair bubble. Biochemical data and comparisons with prior structures6,7 explain how XPA and XPG can switch TFIIH from a transcription factor to a DNA repair factor. During transcription, the kinase module inhibits the repair helicase XPD8. For DNA repair, XPA dramatically rearranges the core TFIIH structure, which reorients the ATPases, releases the kinase module and displaces a ‘plug’ element from the DNA-binding pore in XPD. This enables XPD to move by ~80 Å, engage with DNA, and scan for the lesion in a XPG-stimulated manner. Our results provide the basis for a detailed mechanistic analysis of the NER mechanism.

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Increased Sensitivity of PBMCs Isolated from Patients with Rheumatoid Arthritis to DNA Damaging Agents Is Connected with Inefficient DNA Repair

Journal of Clinical Medicine ◽

10.3390/jcm9040988 ◽

2020 ◽

Vol 9 (4) ◽

pp. 988

Author(s):

Grzegorz Galita ◽

Olga Brzezińska ◽

Izabela Gulbas ◽

Joanna Sarnik ◽

Marta Poplawska ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Dna Repair ◽

Mononuclear Cells ◽

Excision Repair ◽

Dna Lesions ◽

Dna Breaks ◽

Peripheral Blood Mononuclear ◽

Dna Damaging Agents ◽

Repair Efficiency ◽

Systemic Inflammatory Disease

Rheumatoid arthritis (RA) is a systemic, inflammatory disease of the joints and surrounding tissues. RA manifests itself with severe joint pain, articular inflammation, and oxidative stress. RA is associated with certain types of cancer. We have assumed that RA patients’ increased susceptibility to cancer may be linked with genomic instability induced by impaired DNA repair and sensitivity to DNA damaging agents. The aim of this work was to analyze the sensitivity of peripheral blood mononuclear cells (PBMCs) isolated from RA patients to DNA damaging agents: tert-butyl hydroperoxide (TBH), bleomycin, ultraviolet (UV) radiation, and methyl methanesulfonate (MMS) and calculate the repair efficiency. TBH induce oxidative DNA lesions repaired mainly by base excision repair (BER). Bleomycin induced mainly DNA double-strand breaks repaired by non-homologous end joining (NHEJ) and homologous recombination repair (HRR). We included 20 rheumatoid arthritis patients and 20 healthy controls and used an alkaline version of the comet assay with modification to measure sensitivity to DNA damaging agents and DNA repair efficiency. We found an increased number of DNA breaks and alkali-labile sites in the RA patients compared to those in the controls. Exposure to DNA damaging agents evoked the same increased damage in both groups, but we observed statistically higher PMBC sensitivity to TBH, MMS, bleomycin as well as UV. Examination of the repair kinetics of both groups revealed that the DNA lesions induced by TBH and bleomycin were more efficiently repaired in the controls than in the patients. These data suggest impaired DNA repair in RA patients, which may accelerate PBMC aging and/or lead to higher cancer incidence among RA patients.

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Formation of cytosine glycol and 5,6-dihydroxycytosine in deoxyribonucleic acid on treatment with osmium tetroxide

Biochemical Journal ◽

10.1042/bj2350531 ◽

1986 ◽

Vol 235 (2) ◽

pp. 531-536 ◽

Cited By ~ 76

Author(s):

M Dizdaroglu ◽

E Holwitt ◽

M P Hagan ◽

W F Blakely

Keyword(s):

Dna Repair ◽

Formic Acid ◽

Excision Repair ◽

Dna Lesions ◽

Gas Chromatography Mass Spectrometry ◽

Dna Glycosylases ◽

Thymine Glycol ◽

Repair Enzymes ◽

Base Excision

OsO4 selectively forms thymine glycol lesions in DNA. In the past, OsO4-treated DNA has been used as a substrate in studies of DNA repair utilizing base-excision repair enzymes such as DNA glycosylases. There is, however, no information available on the chemical identity of other OsO4-induced base lesions in DNA. A complete knowledge of such DNA lesions may be of importance for repair studies. Using a methodology developed recently for characterization of oxidative base damage in DNA, we provide evidence for the formation of cytosine glycol and 5,6-dihydroxycytosine moieties, in addition to thymine glycol, in DNA on treatment with OsO4. For this purpose, samples of OsO4-treated DNA were hydrolysed with formic acid, then trimethylsilylated and analysed by capillary gas chromatography-mass spectrometry. In addition to thymine glycol, 5-hydroxyuracil (isobarbituric acid), 5-hydroxycytosine and 5,6-dihydroxyuracil (isodialuric acid or dialuric acid) were identified in OsO4-treated DNA. It is suggested that 5-hydroxyuracil was formed by formic acid-induced deamination and dehydration of cytosine glycol, which was the actual oxidation product of the cytosine moiety in DNA. 5-Hydroxycytosine obviously resulted from dehydration of cytosine glycol, and 5,6-dihydroxyuracil from deamination of 5,6-dihydroxycytosine. This scheme was supported by the presence of 5-hydroxyuracil, uracil glycol and 5,6-dihydroxyuracil in OsO4-treated cytosine. Treatment of OsO4-treated cytosine with formic acid caused the complete conversion of uracil glycol into 5-hydroxyuracil. The implications of these findings relative to studies of DNA repair are discussed.

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Databases and Bioinformatics Tools for the Study of DNA Repair

Molecular Biology International ◽

10.4061/2011/475718 ◽

2011 ◽

Vol 2011 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Kaja Milanowska ◽

Kristian Rother ◽

Janusz M. Bujnicki

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Excision Repair ◽

Uv Light ◽

Dna Lesions ◽

Environmental Chemicals ◽

Nonhomologous End Joining ◽

End Joining ◽

Repair Mechanisms ◽

Base Excision

DNA is continuously exposed to many different damaging agents such as environmental chemicals, UV light, ionizing radiation, and reactive cellular metabolites. DNA lesions can result in different phenotypical consequences ranging from a number of diseases, including cancer, to cellular malfunction, cell death, or aging. To counteract the deleterious effects of DNA damage, cells have developed various repair systems, including biochemical pathways responsible for the removal of single-strand lesions such as base excision repair (BER) and nucleotide excision repair (NER) or specialized polymerases temporarily taking over lesion-arrested DNA polymerases during the S phase in translesion synthesis (TLS). There are also other mechanisms of DNA repair such as homologous recombination repair (HRR), nonhomologous end-joining repair (NHEJ), or DNA damage response system (DDR). This paper reviews bioinformatics resources specialized in disseminating information about DNA repair pathways, proteins involved in repair mechanisms, damaging agents, and DNA lesions.

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Genome instability is a consequence of transcription deficiency in patients with bone marrow failure harboring biallelic ERCC6L2 variants

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1803275115 ◽

2018 ◽

Vol 115 (30) ◽

pp. 7777-7782 ◽

Cited By ~ 9

Author(s):

Hemanth Tummala ◽

Arran D. Dokal ◽

Amanda Walne ◽

Alicia Ellison ◽

Shirleny Cardoso ◽

...

Keyword(s):

Bone Marrow ◽

Dna Repair ◽

Rna Polymerase Ii ◽

Nuclear Dna ◽

Genome Instability ◽

Excision Repair ◽

Bone Marrow Failure ◽

Rna Pol Ii ◽

Pol Ii ◽

Repair Defect

Biallelic variants in the ERCC excision repair 6 like 2 gene (ERCC6L2) are known to cause bone marrow failure (BMF) due to defects in DNA repair and mitochondrial function. Here, we report on eight cases of BMF from five families harboring biallelic variants in ERCC6L2, two of whom present with myelodysplasia. We confirm that ERCC6L2 patients’ lymphoblastoid cell lines (LCLs) are hypersensitive to DNA-damaging agents that specifically activate the transcription coupled nucleotide excision repair (TCNER) pathway. Interestingly, patients’ LCLs are also hypersensitive to transcription inhibitors that interfere with RNA polymerase II (RNA Pol II) and display an abnormal delay in transcription recovery. Using affinity-based mass spectrometry we found that ERCC6L2 interacts with DNA-dependent protein kinase (DNA-PK), a regulatory component of the RNA Pol II transcription complex. Chromatin immunoprecipitation PCR studies revealed ERCC6L2 occupancy on gene bodies along with RNA Pol II and DNA-PK. Patients’ LCLs fail to terminate transcript elongation accurately upon DNA damage and display a significant increase in nuclear DNA–RNA hybrids (R loops). Collectively, we conclude that ERCC6L2 is involved in regulating RNA Pol II-mediated transcription via its interaction with DNA-PK to resolve R loops and minimize transcription-associated genome instability. The inherited BMF syndrome caused by biallelic variants in ERCC6L2 can be considered as a primary transcription deficiency rather than a DNA repair defect.

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New perspectives in cancer biology from a study of canonical and non-canonical functions of base excision repair proteins with a focus on early steps

Mutagenesis ◽

10.1093/mutage/gez051 ◽

2019 ◽

Vol 35 (1) ◽

pp. 129-149 ◽

Cited By ~ 5

Author(s):

Matilde Clarissa Malfatti ◽

Giulia Antoniali ◽

Marta Codrich ◽

Silvia Burra ◽

Giovanna Mangiapane ◽

...

Keyword(s):

Dna Repair ◽

Base Excision Repair ◽

Cancer Biology ◽

Molecular Mechanisms ◽

Excision Repair ◽

Dna Lesions ◽

Cancer Development ◽

Dna Repair Enzymes ◽

Repair Enzymes ◽

Base Excision

Abstract Alterations of DNA repair enzymes and consequential triggering of aberrant DNA damage response (DDR) pathways are thought to play a pivotal role in genomic instabilities associated with cancer development, and are further thought to be important predictive biomarkers for therapy using the synthetic lethality paradigm. However, novel unpredicted perspectives are emerging from the identification of several non-canonical roles of DNA repair enzymes, particularly in gene expression regulation, by different molecular mechanisms, such as (i) non-coding RNA regulation of tumour suppressors, (ii) epigenetic and transcriptional regulation of genes involved in genotoxic responses and (iii) paracrine effects of secreted DNA repair enzymes triggering the cell senescence phenotype. The base excision repair (BER) pathway, canonically involved in the repair of non-distorting DNA lesions generated by oxidative stress, ionising radiation, alkylation damage and spontaneous or enzymatic deamination of nucleotide bases, represents a paradigm for the multifaceted roles of complex DDR in human cells. This review will focus on what is known about the canonical and non-canonical functions of BER enzymes related to cancer development, highlighting novel opportunities to understand the biology of cancer and representing future perspectives for designing new anticancer strategies. We will specifically focus on APE1 as an example of a pleiotropic and multifunctional BER protein.

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Bacterial DNA repair genes and their eukaryotic homologues: 4. The role of nucleotide excision DNA repair (NER) system in mammalian cells.

Acta Biochimica Polonica ◽

10.18388/abp.2007_3222 ◽

2007 ◽

Vol 54 (3) ◽

pp. 469-482 ◽

Cited By ~ 12

Author(s):

Leena Maddukuri ◽

Dominika Dudzińska ◽

Barbara Tudek

Keyword(s):

Dna Repair ◽

Mammalian Cells ◽

Excision Repair ◽

Wide Spectrum ◽

Genetic Disorders ◽

Dna Lesions ◽

Entire Genome ◽

Sequence Of Events ◽

Nucleotide Excision

The eukaryotic cell encounters more than one million various kinds of DNA lesions per day. The nucleotide excision repair (NER) pathway is one of the most important repair mechanisms that removes a wide spectrum of different DNA lesions. NER operates through two sub pathways: global genome repair (GGR) and transcription-coupled repair (TCR). GGR repairs the DNA damage throughout the entire genome and is initiated by the HR23B/XPC complex, while the CSB protein-governed TCR process removes DNA lesions from the actively transcribed strand. The sequence of events and the role of particular NER proteins are currently being extensively discussed. NER proteins also participate in other cellular processes like replication, transcription, chromatin maintenance and protein turnover. Defects in NER underlay severe genetic disorders: xeroderma pigmentosum (XP), Cockayne syndrome (CS) and trichothiodystrophy (TTD).

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An alternative eukaryotic DNA excision repair pathway.

Molecular and Cellular Biology ◽

10.1128/mcb.15.8.4572 ◽

1995 ◽

Vol 15 (8) ◽

pp. 4572-4577 ◽

Cited By ~ 42

Author(s):

G A Freyer ◽

S Davey ◽

J V Ferrer ◽

A M Martin ◽

D Beach ◽

...

Keyword(s):

Dna Repair ◽

Excision Repair ◽

Uv Light ◽

Repair Process ◽

Dna Lesions ◽

Repair Pathway ◽

Cyclobutane Pyrimidine Dimers ◽

Dna Excision Repair ◽

Pyrimidine Dimers

DNA lesions induced by UV light, cyclobutane pyrimidine dimers, and (6-4)pyrimidine pyrimidones are known to be repaired by the process of nucleotide excision repair (NER). However, in the fission yeast Schizosaccharomyces pombe, studies have demonstrated that at least two mechanisms for excising UV photo-products exist; NER and a second, previously unidentified process. Recently we reported that S. pombe contains a DNA endonuclease, SPDE, which recognizes and cleaves at a position immediately adjacent to cyclobutane pyrimidine dimers and (6-4)pyrimidine pyrimidones. Here we report that the UV-sensitive S. pombe rad12-502 mutant lacks SPDE activity. In addition, extracts prepared from the rad12-502 mutant are deficient in DNA excision repair, as demonstrated in an in vitro excision repair assay. DNA repair activity was restored to wild-type levels in extracts prepared from rad12-502 cells by the addition of partially purified SPDE to in vitro repair reaction mixtures. When the rad12-502 mutant was crossed with the NER rad13-A mutant, the resulting double mutant was much more sensitive to UV radiation than either single mutant, demonstrating that the rad12 gene product functions in a DNA repair pathway distinct from NER. These data directly link SPDE to this alternative excision repair process. We propose that the SPDE-dependent DNA repair pathway is the second DNA excision repair process present in S. pombe.

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DNA damage stabilizes interaction of CSB with the transcription elongation machinery

The Journal of Cell Biology ◽

10.1083/jcb.200401056 ◽

2004 ◽

Vol 166 (1) ◽

pp. 27-36 ◽

Cited By ~ 84

Author(s):

Vincent van den Boom ◽

Elisabetta Citterio ◽

Deborah Hoogstraten ◽

Angelika Zotter ◽

Jean-Marc Egly ◽

...

Keyword(s):

Molecular Weight ◽

Dna Damage ◽

Dna Repair ◽

Rna Polymerase ◽

Rna Polymerase Ii ◽

High Molecular Weight ◽

Transcription Elongation ◽

Cockayne Syndrome ◽

Dna Lesions ◽

High Molecular Weight Complex

The Cockayne syndrome B (CSB) protein is essential for transcription-coupled DNA repair (TCR), which is dependent on RNA polymerase II elongation. TCR is required to quickly remove the cytotoxic transcription-blocking DNA lesions. Functional GFP-tagged CSB, expressed at physiological levels, was homogeneously dispersed throughout the nucleoplasm in addition to bright nuclear foci and nucleolar accumulation. Photobleaching studies showed that GFP-CSB, as part of a high molecular weight complex, transiently interacts with the transcription machinery. Upon (DNA damage-induced) transcription arrest CSB binding these interactions are prolonged, most likely reflecting actual engagement of CSB in TCR. These findings are consistent with a model in which CSB monitors progression of transcription by regularly probing elongation complexes and becomes more tightly associated to these complexes when TCR is active.

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