Quantitative multiplex sequence analysis of mutational hot spots. Frequency and specificity of mutations induced by a site-specific ethenocytosine in M13 viral DNA

Palejwala, Vaseem A.; Rzepka, Robert W.; Simha, Devendranath; Humayun, M. Zafri

doi:10.1021/bi00066a036

Cited by 58 publications

(87 citation statements)

References 14 publications

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“…Etheno-DNA adducts probably play a causal role in the initiation and progression of liver carcinogenesis. because they produce base pair substitution mutations; ⑀dA can lead to AT 3 CG, AT 3 TA, and AT 3 CG transversions 43,44 ⑀dC can cause CG 3 AT transversions and CG 3 TA transitions, 45,46 and N 2 ,3-ethenodeoxyguanosine, which is also formed in vivo from LPO products, can lead to GC 3 AT transitions. 46 These miscoding properties of etheno-bases strongly implicate them in the initiation of carcinogenesis by vinyl chloride.…”

Section: Discussionmentioning

confidence: 99%

Ethanol-induced cytochrome P4502E1 causes carcinogenic etheno-DNA lesions in alcoholic liver disease

et al. 2009

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Oxidative stress is thought to play a major role in the pathogenesis of hepatocellular cancer (HCC), a frequent complication of alcoholic liver disease (ALD). However, the underlying mechanisms are poorly understood. In hepatocytes of ALD patients, we recently detected by immunohistochemistry significantly increased levels of carcinogenic etheno-DNA adducts that are formed by the reaction of the major lipid peroxidation product, 4-hydroxynonenal (4-HNE) with nucleobases. In the current study, we show that protein-bound 4-HNE and etheno-DNA adducts both strongly correlate with cytochrome P450 2E1 (CYP2E1) expression in patients with ALD (r ‫؍‬ 0.9, P < 0.01). Increased levels of etheno-DNA adducts were also detected in the liver of alcohol-fed lean ( E xcess consumption of alcohol is widespread inWestern countries and, in the United States, alcohol abuse affects approximately 18 million adults. 1 The consumption of more than 80 g ethanol per day for more than 10 years increases the risk for hepatocellular carcinoma (HCC) by approximately fivefold. 2 Worldwide, HCC is one of the most common malignant tumors, ranking as the seventh most prevalent in men and the ninth in women. The risk for HCC in decompensated alcohol-induced cirrhosis is approximately 1% to 2% per year. 2 Although the evolution of alcohol-related HCC involves several factors, 3,4 including the presence of cirrhosis, oxidative stress, disturbed DNA methylation, and defective retinoic acid signaling, the precise mechanisms at a molecular level by which alcohol causes HCC are poorly understood. Recently, the International Agency for Research on Cancer has classified ethanol as a human (group 1) carcinogen, because it induces HCC (among other tumors) in animals and increases the risk for developing HCC in humans. 3,5,6

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Section: Discussionmentioning

confidence: 99%

Ethanol-induced cytochrome P4502E1 causes carcinogenic etheno-DNA lesions in alcoholic liver disease

et al. 2009

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“…Transfection and preparation of pooled progeny phage DNA. The procedures employed for making E. coli cells competent for transfection of εC-DNA or C-DNA, for determining survival effects, and for preparing pooled progeny phage DNA have been described in detail elsewhere (46)(47)(48)(49)53).…”

Section: Methodsmentioning

confidence: 99%

“…The principles and experimental validation of multiplex DNA sequencing technology used to analyze mutation fixation at εC have been described in detail elsewhere (46)(47)(48) and are summarized in Fig. 2.…”

Section: Methodsmentioning

confidence: 99%

“…2, primer extension on each of the five species of DNA within the progeny DNA pool yields a product of a different length. Fractionation of these elongation products by high resolution 16% denaturing polyacrylamide gel electrophoresis followed by computing densitometry of autoradiographs allows the determination of mutation frequency and specificity (46)(47)(48)56).…”

Section: Methodsmentioning

confidence: 99%

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Functional recA, lexA, umuD, umuC, polA, and polB genes are not required for the Escherichia coli UVM response

et al. 1995

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The Escherichia coli UVM response is a recently described phenomenon in which pretreatment of cells with DNA-damaging agents such as UV or alkylating agents significantly enhances mutation fixation at a model mutagenic lesion (3,N 4 -ethenocytosine; C) borne on a transfected M13 single-stranded DNA genome. Since UVM is observed in ⌬recA cells in which SOS induction should not occur, UVM may represent a novel, SOS-independent, inducible response. Here, we have addressed two specific hypothetical mechanisms for UVM: (i) UVM results from a recA-independent pathway for the induction of SOS genes thought to play a role in induced mutagenesis, and (ii) UVM results from a polymerase switch in which M13 replication in treated cells is carried out by DNA polymerase I (or DNA polymerase II) instead of DNA polymerase III. To address these hypotheses, E. coli cells with known defects in recA, lexA, umuDC, polA, or polB were treated with UV or 1-methyl-3-nitro-1-nitrosoguanidine before transfection of M13 single-stranded DNA bearing a site-specific ethenocytosine lesion. Survival of the transfected DNA was measured as transfection efficiency, and mutagenesis at the C residue was analyzed by a quantitative multiplex DNA sequencing technology. Our results show that UVM is observable in ⌬recA cells, in lexA3 (noninducible SOS repressor) cells, in LexA-overproducing cells, and in ⌬umuDC cells. Furthermore, our data show that UVM induction occurs in the absence of detectable induction of dinD, an SOS gene. These results make it unlikely that UVM results from a recAindependent alternative induction pathway for SOS genes. Similarly, UVM is observed in polA (deficient in DNA polymerase I) and polB (deficient in DNA polymerase II) cells, suggesting that neither polymerase plays an indispensable role in UVM induction. Furthermore, our data show that the UVM response is accompanied by enhanced survival (UVM reactivation) of M13 DNA bearing C. The observation of UVM reactivation makes simple repair-suppression models for UVM less attractive and increases the plausibility of mechanisms operating at the level of base insertion. We hypothesize that noncoding lesions fall into two categories. The so-called SOS-dependent (class 1) lesions require SOS functions at the extension (bypass) step, whereas class 2 noncoding lesions do not. It is proposed that UVM, a previously unrecognized damage-inducible response, modulates base insertion at noncoding lesions.Efficient replication past certain DNA lesions depends on factors not required for the replication of normal template DNA. In Escherichia coli, the required additional factors are thought to be encoded by the member genes of the SOS regulon. According to the widely accepted SOS hypothesis, unrepaired DNA damage induces the SOS genes, ultimately resulting in enhanced levels of specific gene products that are proposed to transiently alter the replication machinery.The major features of the regulation of the SOS regulon are well described (20,45,50,65,66,69). Under normal conditions, transcript...

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“…Etheno-adducts are highly promutagenic lesions; edA can lead to ATfiGC transitions as well as ATfiCG and ATfiTA transversions, 18,19 whereas edC can cause CGfiTA transitions and CGfiAT transversions. 18,20 Most sequence changes induced by MDA-DNA adducts are base pair substitutions, with 43% of these substitutions being G>T and 57% being C>T and A>G transversions. 21 Previous studies revealed that such oxidative, stress-related, miscoding etheno-DNA adducts increase in the hepatic tissue of liver cancer-prone patients.…”

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confidence: 99%

Accumulation of lipid peroxidation‐derived DNA lesions in iron‐overloaded thalassemic mouse livers: Comparison with levels in the lymphocytes of thalassemia patients

Meerang¹,

Nair²,

Sirankapracha³

et al. 2009

Intl Journal of Cancer

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In thalassemia patients, iron overload can stimulate lipid peroxidation (LPO), thereby generating miscoding DNA adducts. Adducted DNA was measured in the lymphocytes of b-Thal/Hb E patients and healthy controls and in the organs of thalassemic mice. edA, edC and M 1 dG residues were quantified by 32 P-postlabeling-TLC/ HPLC. M 1 dG levels in lymphocyte DNA from patients were 4 times as high as in controls, while the increase in edA and edC was not significant. Adducted DNA accumulated in the liver of thalassemic mice having >2.7 mg Fe/g tissue dry weight; DNA adducts and iron were highly correlated. edA was not specifically generated in certain mouse liver cell types as revealed by immunohistochemical staining. We found elevated LPO-induced DNA damage in the liver of thalassemic mouse and in lymphocytes, implicating that massive DNA damage occurs in the liver of thalassemia patients. We conclude that promutagenic LPO-derived DNA lesions are involved in the onset of hepatocellular carcinoma in these patients. ' 2009 UICC Key words: iron overload; lipid peroxidation; thalassemia; etheno-DNA adducts Iron overload, one of the major causes of morbidity and mortality in thalassemia patients, results from multiple, life-long transfusions and enhanced iron absorption as a response to increased erythropoiesis. [1][2][3] Iron overload in these patients is indicated by significantly increased levels of serum ferritin, saturated transferrin (the iron carrier protein) 4 and iron deposition in many organs. 5,6 Administration of the iron chelators, desferrioxamine and deferiprone, is commonly used in therapy. These drugs can cause serious side effects and are expensive. For desferrioxamine, a special infusion equipment is needed and, therefore, not all patients can benefit from iron-chelating therapy. In addition, patients who receive iron chelators still encounter a milder or shorter period of iron overload. As a result of iron overload, high levels of plasma iron in the form of nontransferrin bound iron (NTBI) can be found in both transfusion-dependent and transfusion-independent thalassemia patients. 7 NTBI plays a crucial role in the production of hydroxyl radicals (HO • ) in vivo that occurs through the biological Fenton-type and Haber-Weiss reactions. The resulting hydroxyl radicals can induce oxidative stress and, as a consequence, damage of cellular nucleic acids, proteins, lipids and carbohydrates, which in turn can potentiate tissue damage. 8 Previous studies demonstrated increased oxidative stress in thalassemia patients, including increased antioxidant enzyme activities and decreased nonenzymatic antioxidants. 9-12 Moreover, increase in modified DNA bases such as 8-oxodG, resulting from direct attack of hydroxyl radical to DNA, 13 increase in ortho-and meta-tyrosine, which are products of hydroxyl radical attack on phenylalanine, 14 and increase in lipid peroxidation (LPO) products, such as malondialdehyde (MDA) 10 and F 2 -isoprostane, 15 have been reported.LPO end-products such as 4-hydroxy-2-nonenal (HNE) and 1...

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Quantitative multiplex sequence analysis of mutational hot spots. Frequency and specificity of mutations induced by a site-specific ethenocytosine in M13 viral DNA

Cited by 58 publications

References 14 publications

Ethanol-induced cytochrome P4502E1 causes carcinogenic etheno-DNA lesions in alcoholic liver disease

Ethanol-induced cytochrome P4502E1 causes carcinogenic etheno-DNA lesions in alcoholic liver disease

Functional recA, lexA, umuD, umuC, polA, and polB genes are not required for the Escherichia coli UVM response

Accumulation of lipid peroxidation‐derived DNA lesions in iron‐overloaded thalassemic mouse livers: Comparison with levels in the lymphocytes of thalassemia patients

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