The human gene that encodes XRCC1 was cloned nearly thirty years ago but experimental analysis of this fascinating protein is still unveiling new insights into the DNA damage response. XRCC1 is a molecular scaffold protein that interacts with multiple enzymatic components of DNA single-strand break repair (SSBR) including DNA kinase, DNA phosphatase, DNA polymerase, DNA deadenylase, and DNA ligase activities that collectively are capable of accelerating the repair of a broad range of DNA single-strand breaks (SSBs). Arguably the most exciting aspect of XRCC1 function that has emerged in the last few years is its intimate relationship with PARP1 activity and critical role in preventing hereditary neurodegenerative disease. Here, I provide an update on our current understanding of XRCC1, and on the impact of hereditary mutations in this protein and its protein partners on human disease.

• MeSH
• DNA metabolismus   MeSH
• dvouřetězcové zlomy DNA MeSH
• jednořetězcové zlomy DNA MeSH
• lidé MeSH
• oprava DNA * MeSH
• protein XRCC1 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Mammalian DNA base excision repair (BER) is accelerated by poly(ADP-ribose) polymerases (PARPs) and the scaffold protein XRCC1. PARPs are sensors that detect single-strand break intermediates, but the critical role of XRCC1 during BER is unknown. Here, we show that protein complexes containing DNA polymerase β and DNA ligase III that are assembled by XRCC1 prevent excessive engagement and activity of PARP1 during BER. As a result, PARP1 becomes "trapped" on BER intermediates in XRCC1-deficient cells in a manner similar to that induced by PARP inhibitors, including in patient fibroblasts from XRCC1-mutated disease. This excessive PARP1 engagement and trapping renders BER intermediates inaccessible to enzymes such as DNA polymerase β and impedes their repair. Consequently, PARP1 deletion rescues BER and resistance to base damage in XRCC1-/- cells. These data reveal excessive PARP1 engagement during BER as a threat to genome integrity and identify XRCC1 as an "anti-trapper" that prevents toxic PARP1 activity.

• MeSH
• buněčné linie MeSH
• DNA vazebné proteiny metabolismus   MeSH
• DNA-ligasa ATP metabolismus   MeSH
• DNA-polymerasa beta metabolismus   MeSH
• DNA genetika   MeSH
• fibroblasty účinky léků   metabolismus   MeSH
• jednořetězcové zlomy DNA MeSH
• lidé MeSH
• oprava DNA účinky léků   genetika   MeSH
• PARP inhibitory farmakologie   MeSH
• poly-ADP-ribóza-polymeráza 1 metabolismus   MeSH
• poly(ADP-ribosa)-polymerasy metabolismus   MeSH
• poškození DNA účinky léků   genetika   MeSH
• protein XRCC1 metabolismus   MeSH
• vazba proteinů účinky léků   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Genetic defects in the repair of DNA single-strand breaks (SSBs) can result in neurological disease triggered by toxic activity of the single-strand-break sensor protein PARP1. However, the mechanism(s) by which this toxic PARP1 activity triggers cellular dysfunction are unclear. Here we show that human cells lacking XRCC1 fail to rapidly recover transcription following DNA base damage, a phenotype also observed in patient-derived fibroblasts with XRCC1 mutations and Xrcc1-/- mouse neurons. This defect is caused by excessive/aberrant PARP1 activity during DNA base excision repair, resulting from the loss of PARP1 regulation by XRCC1. We show that aberrant PARP1 activity suppresses transcriptional recovery during base excision repair by promoting excessive recruitment and activity of the ubiquitin protease USP3, which as a result reduces the level of monoubiquitinated histones important for normal transcriptional regulation. Importantly, inhibition and/or deletion of PARP1 or USP3 restores transcriptional recovery in XRCC1-/- cells, highlighting PARP1 and USP3 as possible therapeutic targets in neurological disease.

• MeSH
• DNA genetika   MeSH
• genetická transkripce genetika   MeSH
• histony metabolismus   MeSH
• jednořetězcové zlomy DNA * MeSH
• lidé MeSH
• myši knockoutované MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• oprava DNA genetika   MeSH
• oxidační stres genetika   MeSH
• peroxid vodíku toxicita   MeSH
• poly-ADP-ribóza-polymeráza 1 genetika   metabolismus   MeSH
• protein XRCC1 genetika   metabolismus   MeSH
• specifické proteázy ubikvitinu metabolismus   MeSH
• ubikvitinace fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

DNA damage repair (DDR) is a safeguard for genome integrity maintenance. Increasing DDR efficiency could increase the yield of induced pluripotent stem cells (iPSC) upon reprogramming from somatic cells. The epigenetic mechanisms governing DDR during iPSC reprogramming are not completely understood. Our goal was to evaluate the splicing isoforms of histone variant macroH2A1, macroH2A1.1, and macroH2A1.2, as potential regulators of DDR during iPSC reprogramming. GFP-Trap one-step isolation of mtagGFP-macroH2A1.1 or mtagGFP-macroH2A1.2 fusion proteins from overexpressing human cell lines, followed by liquid chromatography-tandem mass spectrometry analysis, uncovered macroH2A1.1 exclusive interaction with Poly-ADP Ribose Polymerase 1 (PARP1) and X-ray cross-complementing protein 1 (XRCC1). MacroH2A1.1 overexpression in U2OS-GFP reporter cells enhanced specifically nonhomologous end joining (NHEJ) repair pathway, while macroH2A1.1 knock-out (KO) mice showed an impaired DDR capacity. The exclusive interaction of macroH2A1.1, but not macroH2A1.2, with PARP1/XRCC1, was confirmed in human umbilical vein endothelial cells (HUVEC) undergoing reprogramming into iPSC through episomal vectors. In HUVEC, macroH2A1.1 overexpression activated transcriptional programs that enhanced DDR and reprogramming. Consistently, macroH2A1.1 but not macroH2A1.2 overexpression improved iPSC reprogramming. We propose the macroH2A1 splicing isoform macroH2A1.1 as a promising epigenetic target to improve iPSC genome stability and therapeutic potential.

• MeSH
• DNA MeSH
• endoteliální buňky metabolismus   MeSH
• histony * metabolismus   MeSH
• indukované pluripotentní kmenové buňky * metabolismus   MeSH
• lidé MeSH
• myši MeSH
• oprava DNA MeSH
• protein XRCC1 genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Genetic polymorphisms in DNA repair genes may influence individual variations in the DNA repair capacity. Polymorphisms in the XRCC1 gene that cause amino acid substitutions may impair the interaction of its proteins (XRCC1) with the other enzymatic proteins and consequently alter DNA repair function, which may be associated with the risk of HIV-1/AIDS disease. In this study, we aimed to determine the frequency of polymorphisms in XRCC1 codon 399 in a sample of Indian population with HIV-1/AIDS to evaluate its association with the disease. Polymerase chain reaction and restriction fragment length polymorphism were used to analyse XRCC1 Arg399Gln polymorphisms in 300 positively diagnosed cases with HIV-1/AIDS and an equal number of negatively diagnosed controls of the matched age. The XRCC1 homozygous variant genotype Gln399Gln was associated with an increased risk of HIV-1/AIDS disease (OR = 1.8, 95% CI 1.10-2.94), while no association was found with the Arg399Gln genotype. Polymorphisms in the XRCC1 homozygous variant genotype for the 399Gln allele were associated with the risk of HIV-1/AIDS disease in a sample of North Indian population.

• MeSH
• AIDS genetika   MeSH
• DNA vazebné proteiny genetika   MeSH
• genetická predispozice k nemoci MeSH
• genotyp MeSH
• HIV-1 patogenita   MeSH
• lidé MeSH
• oprava DNA MeSH
• polymorfismus genetický MeSH
• Check Tag
• lidé MeSH
• Geografické názvy
• Indie MeSH

BACKGROUND: We aimed to detect single nucleotide polymorphisms (SNPs) and mutations in DNA repair genes and their possible association with myelodysplastic syndrome (MDS). METHODS: Targeted enrichment resequencing of 84 DNA repair genes was initially performed on a screening cohort of MDS patients. Real-time polymerase chain reaction was used for genotyping selected SNPs in the validation cohort of patients. RESULTS: A heterozygous frameshift mutation in the XRCC2 gene was identified. It leads to the formation of a truncated non-functional protein and decreased XRCC2 expression level. Decreased expression levels of all DNA repair genes functionally connected with mutated XRCC2 were also present. Moreover, a synonymous substitution in the PRKDC gene and 2 missense mutations in the SMUG1 and XRCC1 genes were also found. In the screening cohort, 6 candidate SNPs were associated with the tendency to develop MDS: rs4135113 (TDG, p = 0.03), rs12917 (MGMT, p = 0.003), rs2230641 (CCNH, p = 0.01), rs2228529 and rs2228526 (ERCC6, p = 0.04 and p = 0.03), and rs1799977 (MLH1, p = 0.04). In the validation cohort, only a polymorphism in MLH1 was significantly associated with development of MDS in patients with poor cytogenetics (p = 0.0004). CONCLUSION: Our study demonstrates that genetic variants are present in DNA repair genes of MDS patients and may be associated with susceptibility to MDS.

• MeSH
• DNA vazebné proteiny genetika   MeSH
• genetická predispozice k nemoci MeSH
• jaderné proteiny genetika   MeSH
• jednonukleotidový polymorfismus MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace * MeSH
• mutační analýza DNA MeSH
• MutL homolog 1 genetika   MeSH
• myelodysplastické syndromy enzymologie   genetika   metabolismus   MeSH
• oprava DNA * MeSH
• protein XRCC1 genetika   MeSH
• proteinkinasa aktivovaná DNA genetika   MeSH
• uracil-DNA-glykosidasa genetika   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Poly(ADP-ribose) is synthesized by PARP enzymes during the repair of stochastic DNA breaks. Surprisingly, however, we show that most if not all endogenous poly(ADP-ribose) is detected in normal S phase cells at sites of DNA replication. This S phase poly(ADP-ribose) does not result from damaged or misincorporated nucleotides or from DNA replication stress. Rather, perturbation of the DNA replication proteins LIG1 or FEN1 increases S phase poly(ADP-ribose) more than 10-fold, implicating unligated Okazaki fragments as the source of S phase PARP activity. Indeed, S phase PARP activity is ablated by suppressing Okazaki fragment formation with emetine, a DNA replication inhibitor that selectively inhibits lagging strand synthesis. Importantly, PARP activation during DNA replication recruits the single-strand break repair protein XRCC1, and human cells lacking PARP activity and/or XRCC1 are hypersensitive to FEN1 perturbation. Collectively, our data indicate that PARP1 is a sensor of unligated Okazaki fragments during DNA replication and facilitates their repair.

• MeSH
• "flap" endonukleasy metabolismus   MeSH
• buněčné linie MeSH
• DNA vazebné proteiny metabolismus   MeSH
• DNA-ligasa ATP metabolismus   MeSH
• DNA genetika   metabolismus   MeSH
• lidé MeSH
• oprava DNA MeSH
• poly-ADP-ribóza-polymeráza 1 metabolismus   MeSH
• poly(ADP-ribosa)-polymerasy genetika   metabolismus   MeSH
• polyadenosindifosfátribosa metabolismus   MeSH
• poškození DNA MeSH
• protein XRCC1 metabolismus   MeSH
• replikace DNA fyziologie   MeSH
• S fáze fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

In response to DNA damage, the histone PARylation factor 1 (HPF1) regulates PARP1/2 activity, facilitating serine ADP-ribosylation of chromatin-associated factors. While PARP1/2 are known for their role in DNA single-strand break repair (SSBR), the significance of HPF1 in this process remains unclear. Here, we investigated the impact of HPF1 deficiency on cellular survival and SSBR following exposure to various genotoxins. We found that HPF1 loss did not generally increase cellular sensitivity to agents that typically induce DNA single-strand breaks (SSBs) repaired by PARP1. SSBR kinetics in HPF1-deficient cells were largely unaffected, though its absence partially influenced the accumulation of SSB intermediates after exposure to specific genotoxins in certain cell lines, likely due to altered ADP-ribosylation of chromatin. Despite reduced serine mono-ADP-ribosylation, HPF1-deficient cells maintained robust poly-ADP-ribosylation at SSB sites, possibly reflecting PARP1 auto-poly-ADP-ribosylation at non-serine residues. Notably, poly-ADP-ribose chains were sufficient to recruit the DNA repair factor XRCC1, which may explain the relatively normal SSBR capacity in HPF1-deficient cells. These findings suggest that HPF1 and histone serine ADP-ribosylation are largely dispensable for PARP1-dependent SSBR in response to genotoxic stress, highlighting the complexity of mechanisms that maintain genomic stability and chromatin remodeling.

• MeSH
• buněčné linie MeSH
• chromatin metabolismus   MeSH
• DNA vazebné proteiny metabolismus   genetika   MeSH
• histony metabolismus   MeSH
• jaderné proteiny metabolismus   genetika   MeSH
• jednořetězcové zlomy DNA * MeSH
• lidé MeSH
• oprava DNA * MeSH
• poly-ADP-ribosylace MeSH
• poly-ADP-ribóza-polymeráza 1 * metabolismus   genetika   MeSH
• poly(ADP-ribosa)-polymerasy metabolismus   genetika   MeSH
• protein XRCC1 metabolismus   genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Úvod: Proteíny XRCC1 a ERCC1 plnia funkciu pri oprave poškodenej DNA. Proteín XRCC1 zohráva úlohu v bázovej excíznej reparácii, kým proteín ERCC1 v nukleotidovej excíznej reparácii. Zvýšená expresia týchto proteínov sa za podmienok nádorového bujnenia podieľa na vzniku liekovej rezistencie. Proteín ERCC1 je dávaný do súvisu so vznikom rezistencie na liečbu platinovými derivátmi. Pacienti a metódy: V práci sme použili 107 vzoriek karcinómov pľúc od pacientov s nemalobunkovým karcinómom. Vo vzorkách sme imunohistochemickou metódou detegovali expresiu DNA reparačných proteínov XRCC1 a ERCC1. Nami detegovanú expresiu proteínov sme porovnali s týmito klinickopatologickými parametrami: histopatologický typ nádoru, grade a TNM štádium ochorenia. Výsledky: Hladiny proteínu XRCC1 boli vo vzorkách skvamocelulárneho a veľkobunkového typu dvojnásobne vyššie (60 % pozitívnych vzoriek) v porovnaní so vzorkami adenokarcinómu (35 % pozitívnych vzoriek). Použitím chí-kvadrátového testu sme ďalej zistili, že vzťah proteín XRCC1 vs typ nádoru bol štatistický významný, nakoľko p = 0,0306. Pri porovnaní expresie proteínu s gradingom a stagingom sme však štatistickú významnosť nepotvrdili. V prípade proteínu ERCC1 sme pozorovali u adenokarcinómu a skvamocelulárneho karcinómu vysokú expresiu proteínu ERCC1. Táto dosahovala hodnoty 64,5 % u adenokarcinómu a 62,5 % u skvamocelulárneho typu. U adenokarcinómu sme ďalej zistili podstatný rozdiel v obsahu proteínov ERCC1 (64,5 % pozitívnych vzoriek) vs XRCC1 (35,5 % pozitívnych vzoriek). V prípade vzoriek veľkobunkového typu bola pozitivita ERCC1 približne rovnaká (45,5 % pozitívnych vzoriek vs 54,5 % negatívnych prípadov). Pri použití chí--kvadrátového testu sme nezaznamenali štatistickú významnosť ani v jednom zo sledovaných parametrov (typ, grading, staging). Záver: Zdá sa, že proteín XRCC1 by mohol predstavovať významný mechanizmus DNA reparácie u skvamocelulárneho a veľkobunkového typu. Okrem toho, jeho expresia bola v korelácii s typom nádoru. V prípade proteínu ERCC1 by sme u pa­cientov s adenokarcinómom a skvamocelulárnym typom mohli predpokladať zvýšenú rezistenciu na liečbu platinovými derivátmi vďaka vysokej pravdepodobnosti expresie proteínu. ­­­Ak by sme však tento predpoklad chceli potvrdiť, potrebovali by sme podstatne zväčšiť náš súbor pacientov. Hladiny ERCC1 však nekorelovali so žiadnymi nami sledovanými klinickopatologickými parametrami. Tento proteín bude pravdepodobne predstavovať u NSCLC prognosticky nezávislý faktor.

Background: Proteins XRCC1 and ERCC1 are involved in DNA repair. XRCC1 plays a role in DNA base excision repair and ERCC1 in nucleotide excision repair pathway. Higher expression profile of both proteins in cancer cells may contribute to development of drug resistance. ERCC1 is involved in removal of platinum adducts and might be a potential predictive and prognostic marker in NSCLC (non-small-cell lung cancer) treated with a cisplatin-based regimen. The purpose of study was determination of XRCC1 and ERCC1 levels and their correlation with basic clini­copathological parameters in NSCLC. Patients and Methods: In this study, 107 tumor samples diagnosed as NSCLC were immunohistochemically examined for expression of XRCC1 and ERCC1 proteins. Our results were compared to basic clinicopathological parameters: type of tumor, tumor grade and stage of disease. For statistical analysis, the chi-square test was used. Results: In squamous cell carcinoma and large cell carcinoma samples, the XRCC1 protein level was twofold higher (60% of positive samples) than in adenocarcinoma samples (35.5% of positive samples). We have found statistical correlation between XRCC1 protein expression and type of tumor (p = 0.0306). On the other hand, the statistical importance between the protein level versus grade and stage was not found. In the case of the ERCC1 protein, we observed the highest protein level in adenocarcinoma (64.5%) and squamous cell carcinoma (62.5%) samples. Next, we determined a significant difference in content of XRCC1 versus ERCC1 (35.5% vs 64.5%) in adenocarcinoma samples. Statistical chi-square test did not reveal any correlation between ERCC1 status and clinicopathological parameters. Conclusion: According to our results, XRCC1 represents an important mechanism of DNA repair in squamous cell and large cell carcinomas. Besides that, expression of XRCC1 was in correlation with type of tumor. In patients with adenocarcinoma and squamous cell carcinoma, we could assume increased resistance to platinum-based therapy because of high expectation of ERCC1 protein expression. However, its levels did not correlate with monitored clinicopathological parameters. The ERCC1 protein will be possibly an independent prognostic factor in NSCLC. To prove a true survival benefit of patients with expression of ERCC1, prospective validation of ERCC1 before clinical implication is needed in the future. Key words: DNA repair proteins – non-small-cell lung carcinoma – immunohistochemistry Submitted: 29. 5. 2012 Accepted: 14. 7. 2012

• Klíčová slova
• XRCC1, ERCC1,
• MeSH
• adenokarcinom genetika   metabolismus   MeSH
• chemorezistence MeSH
• DNA vazebné proteiny * diagnostické užití   genetika   MeSH
• endonukleasy metabolismus   MeSH
• financování organizované MeSH
• imunohistochemie MeSH
• lidé MeSH
• nádory plic genetika   metabolismus   MeSH
• nemalobuněčný karcinom plic * genetika   metabolismus   MeSH
• oprava DNA MeSH
• spinocelulární karcinom genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH

Decreased levels of single-strand breaks in DNA (SSBs), reflecting DNA damage, have previously been observed with increased styrene exposure in contrast to a dose-dependent increase in the base-excision repair capacity. To clarify further the above aspects, we have investigated the associations between SSBs, micronuclei, DNA repair capacity and mRNA expression in XRCC1, hOGG1 and XPC genes on 71 styrene-exposed and 51 control individuals. Styrene concentrations at workplace and in blood characterized occupational exposure. The workers were divided into low (below 50 mg/m³) and high (above 50 mg/m³)) styrene exposure groups. DNA damage and DNA repair capacity were analyzed in peripheral blood lymphocytes by Comet assay. The mRNA expression levels were determined by qPCR. A significant negative correlation was observed between SSBs and styrene concentration at workplace (R=-0.38, p=0.001); SSBs were also significantly higher in men (p=0.001). The capacity to repair irradiation-induced DNA damage was the highest in the low exposure group (1.34±1.00 SSB/10⁹ Da), followed by high exposure group (0.72±0.81 SSB/10⁹ Da) and controls (0.65±0.82 SSB/10⁹ Da). The mRNA expression levels of XRCC1, hOGG1 and XPC negatively correlated with styrene concentrations in blood and at workplace (p<0.001) and positively with SSBs (p<0.001). Micronuclei were not affected by styrene exposure, but were higher in older persons and in women (p<0.001). In this study, we did not confirm previous findings on an increased DNA repair response to styrene-induced genotoxicity. However, negative correlations of SSBs and mRNA expression levels of XRCC1, hOGG1 and XPC with styrene exposure warrant further highly-targeted study.

• MeSH
• DNA vazebné proteiny biosyntéza   genetika   MeSH
• DNA-glykosylasy biosyntéza   genetika   MeSH
• dospělí MeSH
• jednořetězcové zlomy DNA účinky léků   MeSH
• kometový test MeSH
• lidé středního věku MeSH
• lidé MeSH
• lymfocyty účinky léků   MeSH
• messenger RNA biosyntéza   MeSH
• mladý dospělý MeSH
• oprava DNA účinky léků   genetika   MeSH
• pracovní expozice škodlivé účinky   MeSH
• regulace genové exprese účinky léků   MeSH
• styren škodlivé účinky   krev   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH