Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cigarette smoke can cause DNA single strand breaks in cultured human lung cells (T. Nakayama et al., Nature, 314 (1985) 462-464) but the mechanisms behind this DNA damage have not been clearly elucidated. In the present study we have investigated the possibility that one of the major constituents in cigarette smoke, hydroquinone, may be important for mediating smoke-induced DNA damage in the human epithelial lung cell line, A 549, and the mechanisms behind this damage. Cells were exposed to cigarette smoke, hydrogen peroxide, or hydroquinone, in the absence and presence of different inhibitors, and the resulting DNA damage was assessed either as DNA single strand break formation or formation of the oxidative DNA adduct, 8-hydroxydeoxyguanosine. It was found that (i) exposure to cigarette smoke, hydrogen peroxide or hydroquinone causes a rapid decrease in the intracellular thiol level and a considerable DNA single strand break formation, (ii) the formation of DNA single strand breaks in cells exposed to cigarette smoke is inhibited by catalase, dimethylthiourea, and o-phenantroline, suggesting that hydroxyl radicals generated from iron-catalyzed hydrogen peroxide dissociation are involved in the DNA damage, (iii) hydroquinone causes considerable DNA strand break formation that is blocked by aurintricarboxylic acid, an inhibitor of endonuclease activation, and by BAPTA, an intracellular calcium chelator, (iv) addition of hydroquinone to a smoke condensate greatly enhances its ability to cause DNA single strand breaks, and (v) smoke, but not hydroquinone, causes formation of 8-hydroxydeoxyguanosine, a DNA damage product induced by the action of hydroxyl radicals on the DNA base, deoxyguanosine. These findings suggest that the ability of cigarette smoke to cause DNA single strand breaks in cultured lung cells is due to mechanisms involving hydroxyl radical attack on DNA and endonuclease activation. They also suggest that hydroquinone is an important contributor to the DNA damaging effect of cigarette smoke on human lung cells.
 ...
PMID:Cigarette smoke-induced DNA damage in cultured human lung cells: role of hydroxyl radicals and endonuclease activation. 130 85

We have conducted studies to obtain practical knowledge regarding the stability, digestion, and analytical determination of the content of 8-hydroxy-2-deoxy-guanosine (8-OHdG) in oxidatively damaged DNA. Utilizing H2O2 plus uv light to form oxidatively damaged DNA, we found that storage of the DNA at -20 degrees C at alkaline pH caused a significant loss of 8-OHdG, whereas storage at -20 degrees C at neutral or acidic pH prevented loss of 8-OHdG. The 8-OHdG within DNA is stable at 100 degrees C for at least 15 min. Formation of 8-OHdG within DNA using uv light and H2O2 as a hydroxyl free radical-generating system yields the highest amounts when low levels of phosphate buffer are used; but the use of Tris or citrate buffers causes a lower yield of 8-OHdG because these buffers act as scavengers for the hydroxyl free radicals. Independent assessment of hydroxyl free radical flux by the use of salicylate trapping allows assessment of competitive radical reactions. Ethanol washing of plastic microfuge tubes prior to DNA enzymatic digestion improved the yield of 8-OHdG and reduced the variability between samples. Digestion of the oxidatively damaged DNA by the use of a method involving DNase I, endonuclease, phosphodiesterase, and alkaline phosphatase produced the highest yield of 8-OHdG.
 ...
PMID:Conditions influencing yield and analysis of 8-hydroxy-2'-deoxyguanosine in oxidatively damaged DNA. 222 56

Hydroxyurea is a chemotherapeutic agent used for the treatment of myeloproliferative disorders (MPD) and solid tumors. The mutagenic and carcinogenic potential of hydroxyurea has not been established, although hydroxyurea has been associated with an increased risk of leukemia in MPD patients. To clarify whether hydroxyurea has potential carcinogenicity, we examined site-specific DNA damage induced by hydroxyurea using (32)P-5'-end-labeled DNA fragments obtained from the human p53 and p16 tumor suppressor genes and the c-Ha-ras-1 protooncogene. Hydroxyurea caused Cu(II)-mediated DNA damage especially at thymine and cytosine residues. NADH efficiently enhanced hydroxyurea-induced DNA damage. The DNA damage was almost entirely inhibited by catalase and bathocuproine, a Cu(I)-specific chelator, suggesting the involvement of hydrogen peroxide (H(2)O(2)) and Cu(I). Typical free hydroxyl radical scavengers did not inhibit DNA damage by hydroxyurea, but methional did. These results suggest that crypto-hydroxyl radicals such as Cu(I)-hydroperoxo complex (Cu(I)-OOH) cause DNA damage. Formation of 8-hydroxy-2'-deoxyguanosine (8-OHdG) was induced by hydroxyurea in the presence of Cu(II). An electron spin resonance spectroscopic study using N-(dithiocarboxy)sarcosine as a nitric oxide (NO)-trapping reagent demonstrated that NO was generated from hydroxyurea in the presence and absence of catalase. In addition, the generation of formamide was detected by both gas chromatography-mass spectrometry (GC-MS) and time-of-flight-mass spectrometry (TOF-MS). A high concentration of hydroxyurea induced depurination at DNA bases in an H(2)O(2)-independent manner, and endonuclease IV treatment led to chain cleavages. These results suggest that hydroxyurea could induce base oxidation as the major pathway of DNA modification and depurination as a minor pathway. Therefore, it is considered that DNA damage by hydroxyurea participates in not only anti-cancer activity, but also carcinogenesis.
 ...
PMID:Hydroxyurea induces site-specific DNA damage via formation of hydrogen peroxide and nitric oxide. 1171 40

Ischemia/reperfusion is known to result in DNA fragmentation and cell death in kidney tubular epithelium, but the endonucleases responsible for this DNA damage have not been identified. DNA substrate gel analysis of extracts from normal rat kidney cortex revealed the presence of a DNase with an apparent molecular mass of 30 to 34 kD. This enzyme is not a dimer of the previously described nuclear 15-kD endonuclease in kidney cells. Partially purified DNase exhibited characteristics similar to those of rat DNase I. The DNase was able to digest circular DNA (endonuclease), required both Ca(2+) and Mg(2+) ions, and was inhibited by Zn(2+) and by aurintricarboxylic acid; it was not inhibited by G-actin. Rat kidneys were subjected to 40 min of ischemia, followed by 0, 1, 4, 16, or 48 h of reperfusion. The activity of the DNase in cytosolic and nuclear extracts, the 200-bp ladder-generating activity, and 3'OH strand breaks in nuclear DNA were simultaneously increased after ischemia, during the first hours of reperfusion. Oxidative DNA damage, measured as 8-hydroxydeoxyguanosine content, did not coincide with endonuclease-generated DNA breaks. Oxidative DNA damage was increased during ischemia and gradually decreased during reperfusion. Phosphorothioated DNase I antisense oligodeoxynucleotide introduced into cultured NRK-52E rat kidney epithelial cells inhibited DNA fragmentation and attenuated cell death induced by hypoxia/reoxygenation in vitro. The data indicate that the DNase I-like endonuclease may contribute to DNA fragmentation in reperfused rat kidneys.
 ...
PMID:DNase I-like endonuclease in rat kidney cortex that is activated during ischemia/reperfusion injury. 1191 59

In order to improve 8-hydroxyguanine (8-OH-Gua) detection in DNA, we digested isolated DNA with nuclease P1 and analyzed for 8-hydroxydeoxyguanosine 5'-monophosphate (8-OH-dGMP) using a high-performance liquid chromatography system equipped with an electrochemical detector (HPLC-ECD). The amount of 8-OH-Gua in the DNA was expressed as the ratio of 8-OH-dGMP to deoxycytidine monophosphate (dCMP). Using this analysis, the background level of 8-OH-Gua in DNA from human lung carcinoma cells (A549) was several-fold lower than that obtained by a previous method. A549 cells were exposed to 20-60 Gy of gamma-radiation and an increase in 8-OH-Gua concentration was observed with increasing gamma-ray dose (0.3 residues per 10(7) dCMP per Gy). Moreover, by an immunohistochemical procedure using a commercial FITC-kit, 8-OH-Gua was clearly detected in A549 cells and the fluorescence intensity of cells with oxidative DNA damage increased with the doses of gamma-irradiation. Using an endonuclease nicking assay, we also found that gamma-rays decreased 8-OH-Gua repair activity. The results indicate that 8-OH-dGMP is a useful and sensitive marker for estimating oxidative damage in DNA.
 ...
PMID:Analysis of 8-hydroxydeoxyguanosine 5'-monophosphate (8-OH-dGMP) as a reliable marker of cellular oxidative DNA damage after gamma-irradiation. 1280 3

Our previous studies have demonstrated that DNA injury occurs in the brain after intracerebral hemorrhage (ICH). DNA damage can result from at least two pathways, either endonuclease-mediated DNA fragmentation or oxidative injury. The present study investigated the occurrence of the latter after ICH and the role of iron in such injury. Male Sprague-Dawley rats received an infusion of autologous whole blood or ferrous iron into the right basal ganglia. Control rats just had a needle insertion (sham). The rats were sacrificed 1, 3, or 7 days later. 8-Hydroxyl-2'-deoxyguanosine (8-OHdG) was analyzed by immunohistochemistry while the number of apurnic/apyrimidinic abasic sites (AP sites) was also quantified. 8-OHdG and AP sites are two hallmarks of DNA oxidation. Dinitrophenyl (DNP) was measured by Western blotting to compare the time course of protein oxidative damage to that of DNA. DNA repair Ku proteins were measured by Western blot analysis. DNA damage was also examined using DNA polymerase I-mediated biotin-dATP nick translation (PANT) labeling. An increase of 8-OHdG, AP sites and DNP levels and a decrease of Ku levels were observed. Abundant PANT-positive cells were also observed in the perihematomal area 3 days after ICH. In addition, intracerebral infusion of iron increased brain DNP levels and resulted in DNA injury. These results suggest that oxidative stress contributes to DNA damage and brain injury after ICH. Reducing DNA oxidative damage (for example, through iron chelation) may be a therapeutic target for ICH.
 ...
PMID:Oxidative DNA injury after experimental intracerebral hemorrhage. 1578 Oct 43

Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) is the redox regulator of multiple stress-inducible transcription factors, such as NF-kappaB, and the major 5'-endonuclease in base excision repair (BER). We utilized mice containing a heterozygous gene-targeted deletion of APE1/Ref-1 (Apex(+/-)) to determine the impact of APE1/Ref-1 haploinsufficiency on the processing of oxidative DNA damage induced by 2-nitropropane (2-NP) in the liver tissue of mice. APE1/Ref-1 haploinsufficiency results in a significant decline in NF-kappaB DNA-binding activity in response to oxidative stress in liver. In addition, loss of APE1/Ref-1 increases the apoptotic response to oxidative stress, in which significant increases in GADD45g expression, p53 protein stability, and caspase activity are observed. Oxidative stress displays a differential impact on monofunctional (UNG) and bifunctional (OGG1) DNA glycosylase-initiated BER in the liver of Apex(+/-) mice. APE1/Ref-1 haploinsufficiency results in a significant decline in the repair of oxidized bases (e.g., 8-OHdG), whereas removal of uracil is increased in liver nuclear extracts of mice using an in vitro BER assay. Apex(+/-) mice exposed to 2-NP displayed a significant decline in 3'-OH-containing single-strand breaks and an increase in aldehydic lesions in their liver DNA, suggesting an accumulation of repair intermediates of failed bifunctional DNA glycosylase-initiated BER.
 ...
PMID:Oxidative stress alters base excision repair pathway and increases apoptotic response in apurinic/apyrimidinic endonuclease 1/redox factor-1 haploinsufficient mice. 1926 24

Apurinic/apyrimidinic endonuclease (APE) acts as a regulator of p53 or vice versa in the cellular response to oxidative stress. Since oxidative stress-induced apoptosis is suggested in the pathophysiology of diabetic nephropathy, we proposed that APE may have a feasible role in the progression of diabetic complications. We investigated the interrelationship between APE and p53 in streptozotocin-induced diabetic rat kidneys. Variable parameters on kidneys were checked 12 weeks after streptozotocin administration with or without chitosan oligosaccharide (COS) treatment. Streptozotocin administration caused changes as seen in early diabetic nephropathy with increased kidney size, increased p53, decreased APE, and increased cleaved caspase-3. COS was not suspected as being detrimental to renal measurements, and caused the augmentation of APE after streptozotocin administration. The augmented APE, in association with increased p53, suppressed cleaved caspase-3. 8-OHdG was mainly immunolocalized in the distal tubules, but also in the proximal tubules after streptozotocin administration without COS treatment, while APE was observed in proximal tubules in all groups. These results suggested that p53-dependent apoptosis resulting in suppressed APE might be an underlying mechanism of streptozotocin-induced nephropathy.
 ...
PMID:The role of apurinic/apyrimidinic endonuclease on the progression of streptozotocin-induced diabetic nephropathy in rats. 2217 8

Vitiligo is an acquired depigmentation disorder, and reactive oxygen species play an important role in melanocyte damage. Base excision repair is the major pathway responsible for removing reactive oxygen species-induced DNA damage, in which APE1, ADPRT, and XRCC1 play key roles. To investigate the association between genetic variations of these genes and the risk of vitiligo in Chinese populations, we genotyped APE1-Asp148Glu, ADPRT-Val762Ala, and XRCC1-Arg399Gln polymorphisms and measured serum 8-OHdG levels in a hospital-based case-control study. We found that a significantly increased risk of vitiligo was associated with the APE1 Asp/Glu (adjusted odds ratio (OR) 1.24; 95% confidence interval (CI) 1.02-1.52) and Glu/Glu genotypes (adjusted OR 1.48; 95% CI 1.13-1.93), compared with the APE1 Asp/Asp genotype, whereas no vitiligo risk was associated with the genotypes ADPRT-Val762Ala and XRCC1-Arg399Gln. Furthermore, serum 8-OHdG levels were elevated in the APE1-148Glu allele carriers (Asp/Glu+Glu/Glu), in an allele dose-response manner, with the risk of vitiligo (Ptrend<0.05). In addition, we found that the APE1-148Glu variant increased the 8-OHdG levels of cultured human melanocytes treated with H2O2, without any impact on the endonuclease activity. These data suggest that the APE1-Asp148Glu polymorphism aggravates oxidative stress in human melanocytes and contributes to genetic predisposition to vitiligo in Chinese people.
 ...
PMID:Genetic variants of the APE1 gene and the risk of vitiligo in a Chinese population: a genotype-phenotype correlation study. 2336 58

The authors describe a fluorometric method for improving the determination of the cancer biomarker 8-hydroxy-2'-deoxyguanosine (8-OHdG). A nicking endonuclease (NEase)-powered 3-D DNA nanomachine was constructed by assembling hundreds of carboxyfluorescein-labeled single strand oligonucleotides (acting as signal reporter) and tens of swing arms (acting as single-foot DNA walkers) on a gold nanoparticle (AuNP). The activity of this DNA nanomachine was controlled by introducing the protecting oligonucleotides. In the presence of aptamer against 8-OHdG, the protecting oligonucleotides are removed from the swing arms by toehold-mediated strand displacement reaction. In the next step, detached DNA walker hybridizes to the labelled DNA so that the DNA nanomachine becomes activated. Special sequences of signal reporter in the formed duplex can be recognized and cleaved by NEase. As a result, the DNA walker autonomously and progressively moves along the surface of the AuNP, thereby releasing hundreds of signal reporters and causing a rapid increase in green fluorescence. This 3-D nanomachine is highly efficient because one aptamer can release hundreds of signal reporters. These unique properties allowed for the construction of a DNA nanomachine-based method for sensitively detecting 8-OHdG in concentrations as low as 4 pM. This is three orders of magnitude lower compared to previously reported methods. Graphical abstract Schematic of a fluorometric method for determination of the cancer biomarker 8-hydroxy-2'-deoxyguanosine. A nicking endonuclease powered 3D-DNA nanomachine was used to improve the sensitivity. Limit of detection is three orders of magnitude lower than reported methods.
 ...
PMID:Improving the fluorometric determination of the cancer biomarker 8-hydroxy-2'-deoxyguanosine by using a 3D DNA nanomachine. 3028 93


1 2 Next >>