Homozygous mutation of MTPAP causes cellular radiosensitivity and persistent DNA double-strand breaks

The study of rare human syndromes characterized by radiosensitivity has been instrumental in identifying novel proteins and pathways involved in DNA damage responses to ionizing radiation. In the present study, a mutation in mitochondrial poly-A-polymerase (MTPAP), not previously recognized for its role in the DNA damage response, was identified by exome sequencing and subsequently associated with cellular radiosensitivity. Cell lines derived from two patients with the homozygous MTPAP missense mutation were radiosensitive, and this radiosensitivity could be abrogated by transfection of wild-type mtPAP cDNA into mtPAP-deficient cell lines. Further analysis of the cellular phenotype revealed delayed DNA repair, increased levels of DNA double-strand breaks, increased reactive oxygen species (ROS), and increased cell death after irradiation (IR). Pre-IR treatment of cells with the potent anti-oxidants, α-lipoic acid and n-acetylcysteine, was sufficient to abrogate the DNA repair and clonogenic survival defects. Our results firmly establish that mutation of the MTPAP gene results in a cellular phenotype of increased DNA damage, reduced repair kinetics, increased cell death by apoptosis, and reduced clonogenic survival after exposure to ionizing radiation, suggesting a pathogenesis that involves the disruption of ROS homeostasis.     

The GO system prevents ROS-induced mutagenesis and killing in Pseudomonas aeruginosa

Inactivation of the Pseudomonas aeruginosa mutM, mutY , or mutT gene conferred a 2.4-, 17.2-, or 38.1-fold increase in spontaneous mutation frequency, respectively. Importantly, the mutY and mutT strains each displayed a robust H₂O₂-induced mutation frequency. In addition, the mutM, mutY , and mutT mutations severely sensitized P. aeruginosa to killing by H₂O₂, suggesting that these gene products act to repair one or more cytotoxic lesions in P. aeruginosa . Nucleotide sequence analysis of a fragment of the rpoB gene from rifampicin resistant mutM -, mutY -, and, mutT -deficient strains was consistent with this conclusion. These findings are discussed in terms of possible roles for mutM, mutY , and mutT in contributing to survival and mutagenesis of P. aeruginosa colonizing the airways of cystic fibrosis patients.     

DNA氧化性损伤与端粒缩短

末端复制问题（the end replication problem）不能完全解释端粒在某些细胞分裂过程中迅速缩短的现象.40％的高压氧下细胞传代次数降低,端粒缩短速率增大,细胞出现衰老特征,端粒DNA上单链断裂积累.推测端粒缩短的主要原因在于衰老过程中或氧胁迫下端粒DNA单链断裂增多,使端粒末端单链片段在DNA复制时丢失.端粒酶和活性氧对端粒长度的正负调控作用的准确机制还有待于更深入的研究.     

Genotoxicity of streptozotocin in normal and cancer cells and its modulation by free radical scavengers

Streptozotocin (STZ) is an antibiotic which can be used to induce diabetes in experimental animals in order to have an insight into pathogenesis of this disease. To use STZ as a diabetogenic substance, its molecular mode of action should be elucidated. Using the alkaline comet assay, we showed that STZ at concentrations in the range 0.01-100 micromol/L induced DNA damage in normal human lymphocytes and HeLa cancer cells in a dose-dependent manner. Lymphocytes were able to remove damage to their DNA within a 30-min repair incubation, whereas HeLa cells completed the repair in 60 min. Vitamins C and E at 10 and 50 micromol/L diminished the extent of DNA damage induced by 50 micromol/L STZ. Pretreatment of the lymphocytes with the nitrone spin trap, alpha-(4-pyridil-1-oxide)-N-tert-butylnitrone (POBN) or ebselen, which mimics glutathione peroxidase, or pyrrolidine dithiocarbamate (PDTC) reduced the extent of DNA damage evoked by STZ. The cells exposed to STZ and treated with endonuclease III (Endo III), formamidopyrimidine-DNA glycosylase (Fpg) and 3-methyladenine-DNA glycosylase II (AlkA), the enzymes recognizing oxidized and alkylated bases, displayed greater extent of DNA damage than those not treated with these enzymes. These results suggest that free radicals may be involved in the formation of DNA lesions induced by streptozotocin. The drug can also alkylate DNA bases. This broad range of DNA damage induced by STZ indicates that the drug may seriously affect genomic stability in normal and pathological cells.     

Regulation of the activity and expression of ERK8 by DNA damage

We have investigated the agonists that activate transfected extracellular signal-regulated kinase 8 (ERK8) in cells, and have found that the most potent activators are hydrogen peroxide, DNA alkylating and cross-linking agents and the poly (ADP-ribose) polymerase inhibitor KU-0058948. The feature shared by all these agents is that they lead to the accumulation of single strand breaks in DNA, suggesting a role for ERK8 in the response to, or repair of, DNA single strand breaks. The DNA alkylating agent MMS also induced the disappearance of endogenous ERK8 by a proteasome-dependent mechanism.     

邻啡罗啉-Cu对DNA的损伤及其化学核酸酶活性

邻啡罗啉-Cu是一种具有核酸酶活性的配合物,可产生多种形式的DNA损伤,包括碱基修饰、异常碱基位点、链断裂及交联等.近年来,邻啡罗啉-Cu因其切割核酸的性质及作为一种可产生活性氧的模型,引起了学者们浓厚的兴趣,在自由基生物学与医学及核酸化学领域受到广泛的关注.     

Interaction of stannous chloride leads to alteration in DNA, triphosphate nucleotides and isolated bases

Stannous chloride (SnCl₂) is a reducing chemical agent used in several man-made products. SnCl₂ can generate reactive oxygen species (ROS); therefore, studies have been carried out in order to better understand its damaging action in biological systems. In this work, calf thymus DNA, triphosphate nucleotides and isolated bases were incubated with SnCl₂ and the results were analyzed through UV spectrophotometry. The presence of stannous ions altered the absorption spectra of all three isolates. The amount of stannous ions associated to DNA was measured by atomic absorption spectrophotometry. Data showed that more than 40% of the initial SnCl₂ concentration was present in the samples. Our results are in accordance with the damaging potential of this salt and present evidence that stannous ions can complex with DNA, inducing ROS in its vicinity, which may be responsible for the observed lesions. (Mol Cell Biochem xxx: 173–179, 2005)     

Aluminium-induced production of oxygen radicals, lipid peroxidation and DNA damage in seedlings of rice (Oryza sativa)

The effect of aluminium (Al) on seedlings of two rice cultivars, Pusa Basmati and Vikas was investigated after different hours of exposure to 80 mol/L of external Al supply. With increasing time of exposure, the growing seedlings readily absorbed Al and its localization was greater in roots than shoots. Prolonged exposure to Al intensified lipid peroxidation, changed the activities of SOD and peroxidase and caused DNA damage. However, differential responses were observed between the seedlings of two rice cultivars under Al stress. A close inverse relationship existed between decreased root growth and increased Al accumulation, lipid peroxidation, SOD, peroxidase activities and DNA damage. The results demonstrate that roots are the major sites of Al localization and accumulation of Al promoted oxygen free radicals mediated peroxidation of membranes as evidenced by increased MDA levels and the activities of SOD and peroxidase. Our results for the first time showed that Al can cause DNA damage in rice.     

水分胁迫引起的两种不同生态型芦苇的DNA损伤与修复

利用DNA解链荧光分析（FADU）法检测两种不同生态型芦苇（Phragmites communis T.)在PEG6000胁迫处理后的DNA损伤。结果表明：无论是20％还是30％PEG6000胁迫处理,耐旱性强的沙丘芦苇的DNA损伤都比耐旱性弱的沼泽芦苇较轻。利用不同浓度的二乙基二硫代谢氨基甲酸钠（DDC）、H2O2、FeSO4以增加芦苇的3种活性氧（O2^-、H2O2、OH）的实验也同样显示出沙丘芦苇抵抗水分胁迫引起的DNA损伤的能力较强。同时,当加入外源活性氧清除剂二甲基亚砜（DMSO）、抗坏血酸（Vc）时,水分胁迫处理的芦苇DNA损伤表现出不同程度的减轻。当PEG胁迫处理的芦苇复水后,DNA损伤随复水时间延长而逐渐减轻,但沙丘芦苇的DNA损伤修复较快而完全。实验初步证明：水分胁迫可引起植物体内DNA损伤且该损伤与活性氧有关,植物的抗旱性与DNA损伤及修复密切相关。     

Conference overview: Molecular mechanisms of metal toxicity and carcinogenesis

Chronic exposure to many heavy metals and metal-derivatives is associated with an increased risk of cancer, although the mechanisms of tumorigenesis are largely unknown. Approximately 125 scientists attended the 3rd Conference on Molecular Mechanisms of Metal Toxicity and Carcinogenesis and presented the latest research concerning these mechanisms. Major areas of focus included exposure assessment and biomarker identification, roles of ROS and antioxidants in carcinogenesis, mechanisms of metal-induced DNA damage, metal signalling, and the development of animal models for use in metal toxicology studies. Here we highlight some of the research presented, and summarize the conference proceedings.     

Interplay between DNA repair and inflammation,and the link to cancer

Abstract

DNA damage and repair are linked to cancer. DNA damage that is induced endogenously or from exogenous sources has the potential to result in mutations and genomic instability if not properly repaired, eventually leading to cancer. Inflammation is also linked to cancer. Reactive oxygen and nitrogen species (RONs) produced by inflammatory cells at sites of infection can induce DNA damage. RONs can also amplify inflammatory responses, leading to increased DNA damage. Here, we focus on the links between DNA damage, repair, and inflammation, as they relate to cancer. We examine the interplay between chronic inflammation, DNA damage and repair and review recent findings in this rapidly emerging field, including the links between DNA damage and the innate immune system, and the roles of inflammation in altering the microbiome, which subsequently leads to the induction of DNA damage in the colon. Mouse models of defective DNA repair and inflammatory control are extensively reviewed, including treatment of mouse models with pathogens, which leads to DNA damage. The roles of microRNAs in regulating inflammation and DNA repair are discussed. Importantly, DNA repair and inflammation are linked in many important ways, and in some cases balance each other to maintain homeostasis. The failure to repair DNA damage or to control inflammatory responses has the potential to lead to cancer.     

辐射及活性氧对DNA的损伤以及芥子碱的保护作用

在X射线照射下,小牛胸腺DNA的碱基损伤及链断裂随着剂量升高而增加,其损伤主要集中于链断裂;活性氧可以引起DNA损伤,H2O2仅造成少量伤害,当在含有H2O2的体系中加入微量的Cu2＋、Fe2＋时损伤急剧增加,这是由反应产生的·OH所致,Cu2＋的致损伤效果明显高于Fe2＋。·OH清除剂芥子碱具有很强的抗辐射及抗氧化作用,且对DNA无伤害。这说明·OH在DNA的氧化损伤中起重要作用。