MTH1 deficiency selectively increases non-cytotoxic oxidative DNA damage in lung cancer cells: more bad news than good?

Representative images of “Comets” and the corresponding intensity profiles, showing (i) ~ 5% Tail DNA damage, typical of the NSCLC cells treated with no siRNA or scramble siRNA, and analysed by regular Fpg-modified alkaline comet assay (0.8 U Fpg/gel); and (ii) comets showing ~ 10% tail DNA, typical of the NSCLC cells treated with MTH1 siRNA. Superimposed on the Comet images are the image analysis software (Komet 5.5, Andor Technology) determined boundaries demarcating the ‘Comet head’ (pink circle) and ‘tail extent’ (vertical orange line) (Barber RC, Hickenbotham P, Hatch T, Kelly D, Topchiy N, Almeida GM, et al. Radiation-induced transgenerational alterations in genome stability and DNA damage. Oncogene. 2006;25(56):7336–7342). % tail DNA = 100 - % head DNA; % head DNA = (integrated optical head intensity / (integrated optical head intensity + integrated optical tail intensity)) × 100. (PDF 1431 kb

An ab initio and AIM investigation into the hydration of 2-thioxanthine

<p>Abstract</p> <p>Background</p> <p>Hydration is a universal phenomenon in nature. The interactions between biomolecules and water of hydration play a pivotal role in molecular biology. 2-Thioxanthine (2TX), a thio-modified nucleic acid base, is of significant interest as a DNA inhibitor yet its interactions with hydration water have not been investigated either computationally or experimentally. Here in, we reported an <it>ab initio </it>study of the hydration of 2TX, revealing water can form seven hydrated complexes.</p> <p>Results</p> <p>Hydrogen-bond (H-bond) interactions in 1:1 complexes of 2TX with water are studied at the MP2/6-311G(d, p) and B3LYP/6-311G(d, p) levels. Seven 2TX^...H₂O hydrogen bonded complexes have been theoretically identified and reported for the first time. The proton affinities (PAs) of the O, S, and N atoms and deprotonantion enthalpies (DPEs) of different N-H bonds in 2TX are calculated, factors surrounding why the seven complexes have different hydrogen bond energies are discussed. The theoretical infrared and NMR spectra of hydrated 2TX complexes are reported to probe the characteristics of the proposed H-bonds. An improper blue-shifting H-bond with a shortened C-H bond was found in one case. NBO and AIM analysis were carried out to explain the formation of improper blue-shifting H-bonds, and the H-bonding characteristics are discussed.</p> <p>Conclusion</p> <p>2TX can interact with water by five different H-bonding regimes, N-H^...O, O-H^...N, O-H^...O, O-H^...S and C-H^...O, all of which are medium strength hydrogen bonds. The most stable H-bond complex has a closed structure with two hydrogen bonds (N(7)-H^...O and O-H^...O), whereas the least stable one has an open structure with one H-bond. The interaction energies of the studied complexes are correlated to the PA and DPE involved in H-bond formation. After formation of H-bonds, the calculated IR and NMR spectra of the 2TX-water complexes change greatly, which serves to identify the hydration of 2TX.</p

Aberrant repair of etheno-DNA adducts in leukocytes and colon tissue of colon cancer patients

International audienceTo assess the role of lipid peroxidation-induced DNA damage and repair in colon carcinogenesis, the excision rates and levels of 1,N-6-etheno-2'-deoxyadenosine (epsilon dA), 3,N-4-etheno-2'-deoxycytidine (epsilon dC), and 1,N-2-etheno-2'-deoxyguanosine (1,N-2-epsilon dG) were analyzed in polymorphic blood leukocytes (PBL) and resected colon tissues of 54 colorectal carcinoma (CRC) patients and PBL of 56 healthy individuals. In PBL the excision rates of 1,N-6-ethenoadenine (epsilon Ade) and 3,N-4-ethenocytosine (epsilon Cyt), measured by the nicking of oligodeoxynucleotide duplexes with single lesions, and unexpectedly also the levels of epsilon dA and 1,N-2-epsilon dG, measured by LC/MS/MS, were lower in CRC patients than in controls. In contrast the mRNA levels of repair enzymes, alkylpurine- and thymine-DNA glycosylases and abasic site endonuclease (APE1), were higher in PBL of CRC patients than in those of controls, as measured by QPCR. In the target colon tissues epsilon Ade and epsilon Cyt excision rates were higher, whereas the epsilon dA and epsilon dC levels in DNA, measured by P-32-postlabeling, were lower in tumor than in adjacent colon tissue, although a higher mRNA level was observed only for APE1. This suggests that during the onset of carcinogenesis, etheno adduct repair in the colon seems to be under a complex transcriptional and posttranscriptional control, whereby deregulation may act as a driving force for malignancy

Highly mutagenic exocyclic DNA adducts are substrates for the human nucleotide incision repair pathway

Background: Oxygen free radicals induce lipid peroxidation (LPO) that damages and breaks polyunsaturated fatty acids in cell membranes. LPO-derived aldehydes and hydroxyalkenals react with DNA leading to formation of etheno(ε)-bases including 1,N6-ethenoadenine (εA) and 3,N4-ethenocytosine (εC). The εA and εC residues are highly mutagenic in mammalian cells and eliminated in the base excision repair (BER) pathway and/or by AlkB family proteins in the direct damage reversal process. BER initiated by DNA glycosylases is thought to be the major pathway for the removal of non-bulky endogenous base damage. Alternatively, in the nucleotide incision repair (NIR) pathway, the apurinic/apyrimidinic (AP) endonucleases can directly incise DNA duplex 5’ to a damaged base in a DNA glycosylase-independent manner. Methodology/Principal Findings: Here, we characterized the substrate specificity of human major AP endonuclease 1, APE1, towards εA, εC, thymine glycol (Tg) and 7,8-dihydro-8-oxoguanine (8oxoG) residues when present in duplex DNA. APE1 cleaves oligonucleotide duplexes containing εA, εC and Tg, but not those containing 8oxoG. The activity depends strongly on sequence context. The apparent kinetic parameters of the reactions suggest that APE1 has high affinity to DNA containing ε-bases but cleaves DNA duplex at an extremely slow rate. Consistent with this observation, the oligonucleotide duplexes containing an ε-base strongly inhibit AP site nicking activity of APE1 with IC50 values in the range of 5-10 nM. MALDI-TOF MS analysis of the reaction products demonstrated that APE1-catalyzed cleavage of εA•T and εC•G duplexes generates as expected DNA fragments containing 5’-terminal ε-base residue. Conclusions/Significance: The fact that ε-bases and Tg in duplex DNA are recognized and cleaved by APE1 in vitro, suggest that NIR may act as a backup pathway to BER one to remove a large variety of genotoxic base lesions in human cells