scispace - formally typeset
Search or ask a question

Showing papers on "Nuclear DNA published in 2000"


Journal ArticleDOI
TL;DR: The levels of oxidative DNA damage reported in many human tissues or in animal models of carcinogenesis exceed the levels of lesions induced by exposure to exogenous carcinogenic compounds, and it seems likely that oxidativeDNA damage is important in the etiology of many human cancers.
Abstract: A major development of carcinogenesis research in the past 20 years has been the discovery of significant levels of DNA damage arising from endogenous cellular sources. Dramatic improvements in analytical chemistry have provided sensitive and specific methodology for identification and quantitation of DNA adducts. Application of these techniques to the analysis of nuclear DNA from human tissues has debunked the notion that the human genome is pristine in the absence of exposure to environmental carcinogens. Much endogenous DNA damage arises from intermediates of oxygen reduction that either attack the bases or the deoxyribosyl backbone of DNA. Alternatively, oxygen radicals can attack other cellular components such as lipids to generate reactive intermediates that couple to DNA bases. Endogenous DNA lesions are genotoxic and induce mutations that are commonly observed in mutated oncogenes and tumor suppressor genes. Their mutagenicity is mitigated by repair via base excision and nucleotide excision pathways. The levels of oxidative DNA damage reported in many human tissues or in animal models of carcinogenesis exceed the levels of lesions induced by exposure to exogenous carcinogenic compounds. Thus, it seems likely that oxidative DNA damage is important in the etiology of many human cancers. This review highlights some of the major accomplishments in the study of oxidative DNA damage and its role in carcinogenesis. It also identifies controversies that need to be resolved. Unraveling the contributions to tumorigenesis of DNA damage from endogenous and exogenous sources represents a major challenge for the future.

1,825 citations


Journal ArticleDOI
TL;DR: Two redundant parallel pathways may lead to chromatin processing during apoptosis, including one which involves Apaf-1 and caspases, as well as CAD, and leads to oligonucleosomal DNA fragmentation and advanced chromatin condensation and the other which is caspase-independent.
Abstract: Apaf-1−/− or caspase-3−/− cells treated with a variety of apoptosis inducers manifest apoptosis-associated alterations including the translocation of apoptosis-inducing factor (AIF) from mitochondria to nuclei, large scale DNA fragmentation, and initial chromatin condensation (stage I). However, when compared with normal control cells, Apaf-1−/− or caspase-3−/− cells fail to exhibit oligonucleosomal chromatin digestion and a more advanced pattern of chromatin condensation (stage II). Microinjection of such cells with recombinant AIF only causes peripheral chromatin condensation (stage I), whereas microinjection with activated caspase-3 or its downstream target caspase-activated DNAse (CAD) causes a more pronounced type of chromatin condensation (stage II). Similarly, when added to purified HeLa nuclei, AIF causes stage I chromatin condensation and large-scale DNA fragmentation, whereas CAD induces stage II chromatin condensation and oligonucleosomal DNA degradation. Furthermore, in a cell-free system, concomitant neutralization of AIF and CAD is required to suppress the nuclear DNA loss caused by cytoplasmic extracts from apoptotic wild-type cells. In contrast, AIF depletion alone suffices to suppress the nuclear DNA loss contained in extracts from apoptotic Apaf-1−/− or caspase-3−/− cells. As a result, at least two redundant parallel pathways may lead to chromatin processing during apoptosis. One of these pathways involves Apaf-1 and caspases, as well as CAD, and leads to oligonucleosomal DNA fragmentation and advanced chromatin condensation. The other pathway, which is caspase-independent, involves AIF and leads to large-scale DNA fragmentation and peripheral chromatin condensation.

765 citations


Journal ArticleDOI
TL;DR: The data suggest that the kidney barrier in rodents and humans is permeable to DNA molecules large enough to be analyzed by standard genetic methodologies.
Abstract: Background: Cell-free DNA from dying cells recently has been discovered in human blood plasma. In experiments performed on animals and humans, we examined whether this cell-free DNA can cross the kidney barrier and be used as a diagnostic tool. Methods: Mice received subcutaneous injections of either human Raji cells or purified 32P-labeled DNA. DNA was isolated from urine and analyzed by measurement of radioactivity, agarose gel electrophoresis, and PCR. In humans, the permeability of the kidney barrier to polymeric DNA was assessed by detection in urine of sequences that were different from an organism bulk nuclear DNA. Results: In the experiments on laboratory animals, we found that ∼0.06% of injected DNA was excreted into urine within 3 days in a polymeric form and that human-specific Alu sequences that passed through the kidneys could be amplified by PCR. In humans, male-specific sequences could be detected in the urine of females who had been transfused with male blood as well as in DNA isolated from urine of women pregnant with male fetuses. K- ras mutations were detected in the urine of patients with colon adenocarcinomas and pancreatic carcinomas. Conclusions: The data suggest that the kidney barrier in rodents and humans is permeable to DNA molecules large enough to be analyzed by standard genetic methodologies.

380 citations


Journal ArticleDOI
TL;DR: A role of Rad51 in recombinational repair processes of DNA damage present in postreplicative chromatin is supported by ultraviolet microirradiation experiments of small nuclear areas and whole cell ultraviolet C (UVC) irradiation experiments performed with a human fibroblast cell line.
Abstract: Rad51, a eukaryotic RecA homologue, plays a central role in homologous recombinational repair of DNA double-strand breaks (DSBs) in yeast and is conserved from yeast to human. Rad51 shows punctuate nuclear localization in human cells, called Rad51 foci, typically during the S phase (Tashiro, S., N. Kotomura, A. Shinohara, K. Tanaka, K. Ueda, and N. Kamada. 1996. Oncogene. 12:2165–2170). However, the topological relationships that exist in human S phase nuclei between Rad51 foci and damaged chromatin have not been studied thus far. Here, we report on ultraviolet microirradiation experiments of small nuclear areas and on whole cell ultraviolet C (UVC) irradiation experiments performed with a human fibroblast cell line. Before UV irradiation, nuclear DNA was sensitized by the incorporation of halogenated thymidine analogues. These experiments demonstrate the redistribution of Rad51 to the selectively damaged, labeled chromatin. Rad51 recruitment takes place from Rad51 foci scattered throughout the nucleus of nonirradiated cells in S phase. We also demonstrate the preferential association of Rad51 foci with postreplicative chromatin in contrast to replicating chromatin using a double labeling procedure with halogenated thymidine analogues. This finding supports a role of Rad51 in recombinational repair processes of DNA damage present in postreplicative chromatin.

213 citations


Journal ArticleDOI
TL;DR: This chapter discusses mitochondrial respiratory chain diseases, which are a highly diverse group of disorders whose main unifying characteristic is the impairment of mitochondrial function.

174 citations


Journal ArticleDOI
TL;DR: It is found that in the presence of hypoxia-mimicking concentrations of CoCl(2), mitochondrial but not nuclear DNA damage is induced in rat neuronal, PC12 cells, the first documentation of induction of mitochondrial DNA (mtDNA) damage under these conditions.
Abstract: Generation of reactive oxygen species (ROS) and activation of a transcriptional program that mimics the hypoxic response have been documented in cultured cells in the presence of cobalt chloride. We found that in the presence of hypoxia-mimicking concentrations of CoCl(2), mitochondrial but not nuclear DNA damage is induced in rat neuronal, PC12 cells. To our knowledge, this is the first documentation of induction of mitochondrial DNA (mtDNA) damage under these conditions. Likewise, we provide the first evidence for elevation of MYH, the mammalian homolog of the Escherichia coli MutY DNA glycosylase, in mammalian cells. Recently, the human MYH was implicated in repair of oxidative DNA damage and shown to carry a mitochondrial localization sequence. Here, an induction of mtDNA damage and a time-dependent increase in the MYH level were detected with exposure of cells to 100 microM CoCl(2). In addition, the levels of proteins involved in cellular responses to hypoxia, ROS and nuclear DNA damage; hypoxia-inducible factor 1alpha(HIF-1alpha), p53, p21 and PCNA were also modulated temporally. Earlier studies suggested that the mtDNA is a primary target for oxidative damage. Our findings extend these observations and suggest that activation of DNA repair processes is associated with the presence of mtDNA damage.

169 citations


Journal ArticleDOI
TL;DR: It is concluded that it is important to examine many loci when estimating genetic differentiation to infer historical amounts of gene flow and patterns of genetic exchange among populations, less important whether those loci are allozymes or nuclear DNA markers.
Abstract: We examined genetic variation at 21 polymorphic allozyme loci, 15 nuclear DNA loci, and mitochondrial DNA in four spawning populations of sockeye salmon (Oncorhynchus nerka) from Cook Inlet, Alaska, to test for differences in the patterns of divergence among different types of markers. We were specifically interested in testing the suggestion that natural selection at allozyme loci compromises the effectiveness of these markers for describing the amount and patterns of gene flow among populations. We found concordance among markers in the amount of genetic variation within and among populations, with the striking exception of one allozyme locus (sAH), which exhibited more than three times the amount of among-population differentiation as other loci. A consideration of reports of discordance between allozymes and other loci indicates that these differences usually result from one or two exceptional loci. We conclude that it is important to examine many loci when estimating genetic differentiation to infer historical amounts of gene flow and patterns of genetic exchange among populations. It is less important whether those loci are allozymes or nuclear DNA markers.

160 citations


Journal ArticleDOI
TL;DR: Analysis of a test microarray of 400 cDNAs and small random genomic DNA fragments probed with RNAs from two developmental stages of T. brucei demonstrates that the microarray technology can be used to identify batteries of genes differentially expressed during the various life cycle stages of this parasite.

133 citations


Journal ArticleDOI
01 Mar 2000-Genetics
TL;DR: No evidence was found to support hypotheses that predict that oxidative damage or accumulation of errors in nuclear DNA contributes significantly to the aging process, at least in these three somatic tissues.
Abstract: Mutation frequency and specificity were determined as a function of age in nuclear DNA from liver, bladder, and brain of Big Blue lacI transgenic mice aged 1.5-25 months. Mutations accumulated with age in liver and accumulated more rapidly in bladder. In the brain a small initial increase in mutation frequency was observed in young animals; however, no further increase was observed in adult mice. To investigate the origin of mutations, the mutational spectra for each tissue and age were determined. DNA sequence analysis of mutant lacI transgenes revealed no significant changes in mutational specificity in any tissue at any age. The spectra of mutations found in aging animals were identical to those in younger animals, suggesting that they originated from a common set of DNA lesions manifested during DNA replication. The data also indicated that there were no significant age-related mutational changes due to oxidative damage, or errors resulting from either changes in the fidelity of DNA polymerase or the efficiency of DNA repair. Hence, no evidence was found to support hypotheses that predict that oxidative damage or accumulation of errors in nuclear DNA contributes significantly to the aging process, at least in these three somatic tissues.

101 citations


Journal ArticleDOI
TL;DR: The high sensitivity of the nucleus to the medium highlights the problem of reliability limits in estimating genome size and the choice between internal and external standardization and the interpretation of intraspecific variation in ‘nuclear DNA content’ are discussed.

97 citations


Journal ArticleDOI
TL;DR: The present study showed that the levels of damage measured using high‐pressure liquid chromatography/electrochemical detection and an enzymatic/Southern blot assay were comparable, and this assay was used to determine the level of in vivo damage present in rat liver mtDNA both with and without organelle isolation.
Abstract: The oxidatively induced DNA lesion 8-oxo-dG in mitochondrial DNA (mtDNA) is commonly used as a marker for oxidative damage to mitochondria, which in turn is thought to be a fundamental cause of aging For years, mitochondrial levels of 8-oxo-dG were believed to be ∼10-fold higher in mtDNA than in nuclear DNA even in normal, young animals However, studies in our own and other laboratories have shown that this lesion is efficiently repaired Also, mutational consequences specific to 8-oxo-dG (G to T transversions) are rarely reported In the present study, we showed that the levels of damage measured using high-pressure liquid chromatography/electrochemical detection and an enzymatic/Southern blot assay were comparable The latter assay does not require isolation of mitochondria, and so this assay was then used to determine the level of in vivo damage present in rat liver mtDNA both with and without organelle isolation Levels of 8-oxo-dG are approximately threefold higher when measured in mtDNA purified f

Journal ArticleDOI
TL;DR: The results demonstrate both the stability of the T. brucei genome, as illustrated by the conservation of syntenic groups of genes in the two stocks, and the polymorphic nature of the genomic regions involved in antigenic variation.

Journal ArticleDOI
TL;DR: It is concluded that both nuclear and mitochondrial oxidative DNA damages can be simultaneously detected in situ using immunofluorescence labeling with Fab 166 and confocal microscopy.

Journal ArticleDOI
TL;DR: It is shown that approximately 0.2 mole percent of EUGLENA: DNA consists of J, an amount similar to that found in DNA of Trypanosoma brucei, and this adds to the existing evidence for a close phylogenetic relation between kinetoplastids and euglenids.
Abstract: We have analyzed DNA of Euglena gracilis for the presence of the unusual minor base β-D-glucosylhydroxymethyluracil or J, thus far only found in kinetoplastid flagellates and in Diplonema .U sing antibodies specific for J and post-labeling of DNA digests followed by two-dimensional thin-layer chromatography of labeled nucleotides, we show that ~0.2 mole percent of Euglena DNA consists of J, an amount similar to that found in DNA of Trypanosoma brucei. By staining permeabilized Euglena cells with anti-J antibodies, we show that J is rather uniformly distributed in the Euglena nucleus, and does not co-localize to a substantial extent with (GGGTTA) n repeats, the putative telomeric repeats of Euglena. Hence, most of J in Euglena appears to be non-telomeric. Our results add to the existing evidence for a close phylogenetic relation between kinetoplastids and euglenids.

Journal ArticleDOI
TL;DR: The increase or decrease in the copy number of several repetitive sequences examined here may indicate the existence of a directed change in genome size in rice.
Abstract: This paper describes a fluorescence in situ hybridization (FISH) analysis of three different repetitive sequence families, which were mapped to mitotic metaphase chromosomes and extended DNA fibers (EDFs) of the two subspecies of rice (OrYza sativa), indica and japonica (2n = 2x = 24). The repeat families studied were (1) the tandem repeat sequence A (TrsA), a functionally non-significant repeat; (2) the [TTTA-GGG]n telomere sequence, a non-transcribed, tandemly repeated but functionally significant repeat; and (3) the 5S ribosomal RNA (5S rDNA). FISH of the TrsA repeat to metaphase chromosomes of indica and japonica cultivars revealed clear signals at the distal ends of twelve and four chromosomes, respectively. As shown in a previous report, the 17S ribosomal RNA genes (17S rDNA) are located at the nucleolus organizers (NORs) on chromosomes 9 and 10 of the indica cultivar. However, the japonica rice lacked the rDNA signals on chromosome 10. The size of the 5S rDNA repeat block, which was mapped on the chromosome 11 of both cultivars, was 1.22 times larger in the indica than in the japonica genome. The telomeric repeat arrays at the distal ends of all chromosome arms were on average three times longer in the indica genome than in the japonica genome. Flow cytometric measurements revealed that the nuclear DNA content of indica rice is 9.7% higher than that of japonica rice. Our data suggest that different repetitive sequence families contribute significantly to the variation in genome size between indica and japonica rice, though to different extents. The increase or decrease in the copy number of several repetitive sequences examined here may indicate the existence of a directed change in genome size in rice. Possible reasons for this phenomenon of concurrent evolution of various repeat families are discussed.

Journal ArticleDOI
TL;DR: Graphical comparisons indicated that unpartitioned cytochrome b has evolved at 5-10 times the rate of scnDNA, and third-position transversions appeared to offer the most useful sequence partition for phylogenetic analysis.
Abstract: Mitochondrial cytochrome b sequence data from 15 species of herons (Aves: Ardeidae), representing 13 genera, were compared with DNA hybridization data of single-copy nuclear DNA (scnDNA) from the same species in a taxonomic congruence assessment of heron phylogeny. The two data sets produced a partially resolved, completely congruent estimate of phylogeny with the following basic structure: (Tigrisoma, Cochlearius, (((Zebrilus, (Ixobrychus, Botaurus)), (((Ardea, Casmerodius), Bubulcus), ((Egretta thula, Egretta caerulea, Egretta tricolor), Syrigma), Butorides, Nycticorax, Nyctanassa)))). Because congruence indicated similar phylogenetic information in the two data sets, we used the relatively unsaturated DNA hybridization distances as surrogates of time to examine graphically the patterns and rates of change in cytochrome b distances. Cytochrome b distances were computed either from whole sequences or from partitioned sequences consisting of transitions, transversions, specific codon site positions, or specific protein-coding regions. These graphical comparisons indicated that unpartitioned cytochrome b has evolved at 5-10 times the rate of scnDNA. Third-position transversions appeared to offer the most useful sequence partition for phylogenetic analysis because of their relatively fast rate of substitution (two times that of scnDNA) and negligible saturation. We also examined lineage-based rates of evolution by comparing branch length patterns between the nuclear and cytochrome b trees. The degree of correlation in corresponding branch lengths between cytochrome b and DNA hybridization trees depended on DNA sequence partitioning. When cytochrome b sequences were not partitioned, branch lengths in the cytochrome b and DNA hybridization trees were not correlated. However, when cytochrome b sequences were reduced to third-position transversions (i.e., unsaturated, relatively fast changing data), branch lengths were correlated. This finding suggests that lineage-based rates of DNA evolution in nuclear and mitochondrial genomes are influenced by common causes.

Journal ArticleDOI
TL;DR: The large spectrum of deletions identified in this study highlights the ubiquitous nature and the high mutational load of mitochondrial DNA associated with ultraviolet exposure and chronologic aging.

Journal ArticleDOI
01 May 2000-Genetics
TL;DR: The results showed that mouse rho(0) cells could receive mtDNA from a different mouse species, M. spretus, or even mt DNA from the rat, Rattus norvegicus, and that the introduced rat mtDNA caused mitochondrial dysfunction, even though rat mt DNA could restore normal mitochondrial translation in the cybrids.
Abstract: By the fusion of mtDNA-less (rho(0)) cells of Mus musculus domesticus with platelets from different species, mtDNA repopulated cybrids were obtained for finding the mtDNA species that could induce mitochondrial abnormalities. Expression of mitochondrial dysfunction might be expected in these cybrids due to incompatibility between nuclear and mitochondrial genomes from different species. The results showed that mouse rho(0) cells could receive mtDNA from a different mouse species, M. spretus, or even mtDNA from the rat, Rattus norvegicus, and that the introduced rat mtDNA, but not M. spretus mtDNA, caused mitochondrial dysfunction, even though rat mtDNA could restore normal mitochondrial translation in the cybrids. Considering that mitochondrial respiratory complexes consist of nuclear DNA- and mtDNA-coded polypeptides, these observations suggest that the nuclear and mitochondrial interactions required for replication, transcription, and translation of introduced rat mtDNA must be less stringently controlled than those required for formation of normal respiratory complexes. As no procedure for introduction of mutagenized mouse mtDNA into living cells has yet been established, these findings provide important insights into generating mtDNA-knockout mice.

Journal ArticleDOI
TL;DR: The evidence for efficient photorepair of organellar DNA contrasts with previous studies of irradiated 5-day-old seedlings, and with the apparent absence of Arabidopsis photolyases bearing transit peptides.
Abstract: Replication of Arabidopsis nuclear, mitochondrial and chloroplast DNA (ncDNA, mtDNA, cpDNA) was assayed by measuring respective changes in copies per leaf, employing quantitative PCR (QPCR) analysis with genome-specific primer pairs. All three genomes showed parallel increases during growth of cotyledons and 5th leaves in planta, maintaining approximately 13 mtDNA copies and 280 cpDNA copies per haploid nuclear genome. Detached 5th leaves, which showed good growth and DNA replication on agar plates, were irradiated at (DNA-effective) UV-B fluences of 1.3-5.0 kJ m-2 and incubated under blue (photorepair-active) plus gold light or gold light only. Under blue light, replication of all genomes after all UV fluences was approximately as efficient as replication in unirradiated leaves. UV-irradiated leaves showed little growth under gold light only; 5 kJ m-2 stopped replication of all three genomes, 2.5 kJ m-2 stopped only cpDNA replication, and 1.3 kJ m-2 only delayed cpDNA replication. Immunoassays showed that 5 kJ m-2 induced about 1.2 cyclobutane pyrimidine dimers and 0.1 [6-4]photoproducts per kbp of bulk DNA, and that both photoproducts were completely removed during 2-3 days under blue light, suggesting efficient photorepair of at least ncDNA and cpDNA. The evidence for efficient photorepair of organellar DNA contrasts with previous studies of irradiated 5-day-old seedlings, and with the apparent absence of Arabidopsis photolyases bearing transit peptides.

Journal ArticleDOI
TL;DR: Examination of genetic diversity of P. grisea in Arkansas indicated that two races predominated in the regional collection, and the data indicate that certain cultivars grown in Arkansas may serve as a "bottleneck", selecting out specific lineages in the local population.
Abstract: MGR586 DNA fingerprinting has been widely used to characterize population diversity of the rice blast pathogen, Pyricularia grisea. However, the frequency and distribution of particular haplotypes (individuals) within MGR-delimited lineages has not been examined in the United States. MGR586 DNA fingerprinting, mitochondrial DNA (mtDNA) restriction fragment length polymorphism (RFLPs), and virulence phenotyping were used to examine genetic diversity of P. grisea in Arkansas. A total of 470 monoconidial isolates were recovered from eight rice cultivars in 18 commercial fields in nine counties in Arkansas. All isolates were examined for nuclear DNA RFLPs with the MGR586 DNA fingerprint probe, and both the MGR lineage (isolates with >80% similarity) and the haplotype frequencies were determined. Four distinct MGR586 DNA fingerprint lineages (designated A, B, C, and D) were identified among the 470 field isolates. All four lineages were found in 9 of the 18 locations. Three lineages were found in four...

Journal ArticleDOI
TL;DR: Covalent cross-linking of DNA to HMG I/Y by FR900482 in vivo is reported which represents the first example of a covalent DNA-drug-protein cross-link with a minor groove-binding oncoprotein and a potential novel mechanism through which these compounds exert their anti-tumor activity.

Journal ArticleDOI
TL;DR: Data showed that the subcellular site of nucleoside analog phosphorylation is an important determinant for incorporation of nucleOSide analogs into nuclear or mitochondrial DNA.

Journal ArticleDOI
TL;DR: The results demonstrate that the spectral changes between normal and neoplastic tissue are mostly due to an enhanced signature of DNA in neoplasia, which is sufficiently large to suggest that it is most likely due to a increased detectability of DNA, rather than an increase in concentration.
Abstract: In our efforts to understand the infrared spectral features of cells and tissues, and the spectral changes occurring between normal and disease states, we reported previously a detailed correlation between histochemical/immunohistochemical and spectral results These results suggested an increase of nucleic acid spectral contributions in neoplastic, as compared to normal, tissue samples In the present paper, these studies are extended to report the spectral features of DNA and RNA separately in these tissue samples This was accomplished by selectively digesting either DNA or RNA from tissue sections, leaving behind the protein matrix with nuclear/cytoplasmic RNA or the protein matrix with nuclear DNA, respectively These results demonstrate that the spectral changes between normal and neoplastic tissue are mostly due to an enhanced signature of DNA in neoplastic tissue This enhancement is sufficiently large to suggest that it is most likely due to an increased detectability of DNA, rather than an increase in concentration Index Headings: Infrared microspectroscopy; Human tissue; Liver; Cancer; Enzyme digestion

Journal ArticleDOI
TL;DR: It is concluded that sub‐alpine Abies veitchii and A. homolepis produce natural hybrids, and that their systematic relationship should be re‐evaluated.
Abstract: Sub-alpine Abies veitchii and A. homolepis are distributed in the central part of Honshu Island, Japan, and their habitats are segregated vertically. These species sometimes form a mixed forest in the overlapping area of the two species, that is, in the upper limit of the A. homolepis habitat and the lower limit of A. veitchii. These species have been considered to be distantly related because they were classified into different sections by most conventional classifications. No natural hybridization has been reported between the two species. The aim of this study was to demonstrate, through the use of molecular markers, whether natural hybridization takes place between these two species at two experimental sites on Mt. Fuji, where the species occur naturally. DNA markers from paternally inherited chloroplast DNA (cpDNA), maternally inherited mitochondrial DNA (mtDNA) and biparentally inherited nuclear DNA (nDNA), were used for this study. As organelle DNA markers, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) markers were developed to determine the maternal and paternal species for each individual. Two of 334 individuals possessed a cpDNA haplotype derived from A. homolepis and a mtDNA haplotype from A. veitchii. Furthermore, the nDNA of these two individuals was analysed using the random amplified polymorphic DNA (RAPD) assay to investigate their genomic composition. RAPD analysis indicated that the nuclear genomes of the two individuals were derived from both species. We conclude that A. veitchii and A. homolepis produce natural hybrids, and that their systematic relationship should be re-evaluated.

Journal ArticleDOI
TL;DR: Although 8-NO(2)-Gua was generated in isolated DNA by different nitrating systems, other types of damage were formed in abundance, and the lesion could not be found reliably in nuclear DNA, suggesting that the biological importance is limited.

Journal Article
TL;DR: It is shown that the intermediate filament protein vimentin is associated with the nuclear DNA of MDA-MB-231 breast cancer cells, an estrogen receptor-negative, hormone-independent breast cancer cell line with high metastatic potential, and may play important roles in breast tumorigenesis.
Abstract: The nuclear matrix is a dynamic RNA-protein complex that organizes chromatin and regulates nuclear DNA metabolism. Nuclear matrix proteins informative in the diagnosis of cancer have been identified. Here, the nuclear matrix breast cancer proteins (NMBCs) cross-linked to nuclear DNA in situ with cisplatin in human breast cancer cell lines were analyzed by two-dimensional gel electrophoresis. We identified NMBCs that were differentially associated with nuclear DNA of hormone-dependent and -independent breast cancer cell lines. Three DNA cross-linked NMBCs were found to be exclusive to estrogen receptor-positive, hormone-dependent breast cancer cells, whereas two NMBCs were observed only in estrogen receptor-negative, hormone-independent breast cancer cells. Changes in these NMBCs were observed when hormone-dependent breast cancer cells became hormone independent. Furthermore, we show that the intermediate filament protein vimentin is associated with the nuclear DNA of MDA-MB-231 breast cancer cells, an estrogen receptor-negative, hormone-independent breast cancer cell line with high metastatic potential. These nuclear matrix DNA-binding proteins may play important roles in breast tumorigenesis.

Journal ArticleDOI
TL;DR: Polymerase chain reaction (PCR) and RFLP of the intergenic spacer region-1 (IGS-1) generated banding patterns for nine Armillaria species, and new banded patterns are presented for A. ostoyae, A. gemina, and A. mellea.
Abstract: Twenty-six isolates representing nine North American Armillaria species were investigated with flow cytometry and RFLP (restriction fragment length polymorphism) analyses to determine their nuclear DNA content and RFLP profile. Three pu- tatively diploid isolates of A. ostoyae, A. gemina, A. calvescens, A. sinapina, A. mellea, A. gallica, A. nabs- nona, and North American Biological Species (NABS) X were analyzed, and two putatively diploid isolates of NABS XI also were analyzed. Nuclear DNA contents of Armillaria species were 0.11-0.17 pg per nucleus (55-84 X 10" bp/C), depending on species. Among the nine North American Armillaria species tested, A. ostoyae, A. gemirza, and A. mellea possessed relatively small nuclear DNA contents (0.1 1-0.12 pg per nucleus), whereas A. gallica possessed a relatively large nuclear DNA content (0.17 pg per nucleus). A. nabsno,na has a slightly larger nuclear DNA content (0.13pg per nucleus) than A. ostojae, A. gemina, and A. mellea. Other species (A. calvescens, A. sinapina, NABS X, and NABS XI) possessed moderate nuclear DNA contents (ca 0.15 pg per nucleus). Polymerase chain reaction (PCR) and RFLP of the intergenic spacer region-1 (IGS-1) generated banding patterns for nine Armillaria species. In addition to previously reported banding patterns, new banding patterns are

Journal ArticleDOI
TL;DR: It is verified the existence of the T3250C and T3291C mutations, which have been found in patients with mitochondrial myopathy, in the authentic mitochondrial genome.
Abstract: Technical advancements in molecular genetics have shown various mitochondrial DNA (mtDNA) abnormalities in patients with mitochondrial myopathies. Recently, it has been revealed that, in these patients, the nuclear DNA carries sequences similar to those of the mtDNA (nuclear pseudogene) and it has several point mutations previously reported to be pathogenic. We verified the existence of the T3250C and T3291C mutations, which we have found in patients with mitochondrial myopathy, in the authentic mitochondrial genome. A long polymerase chain reaction provides a powerful tool for avoiding nuclear pseudogene amplification and for ruling out ambiguity in the detection of the mutation for diagnosis.

Journal ArticleDOI
TL;DR: Testing for Ku activity in mitochondrial extracts prepared from the xrs-5 hamster cell line that lacks Ku80 mRNA expression shows consistent with the hypothesis that a C-terminally truncated form of Ku80 is localized in mammalian mitochondria where it functions in a DNA end-binding activity.
Abstract: Mammalian mitochondrial DNA end-binding activity is nearly indistinguishable from that of nuclear Ku. This observation led to the hypothesis that mitochondrial DNA end-binding activity is in part dependent upon Ku80 gene expression. To test this hypothesis, we assayed for Ku activity in mitochondrial extracts prepared from the xrs-5 hamster cell line that lacks Ku80 mRNA expression. Mitochondrial protein extracts prepared from this cell line lacked the DNA end-binding activity found in similar extracts prepared from wild-type cells. Azacytidine-reverted xrs-5 cells that acquired nuclear DNA end-binding activity also acquired mitochondrial DNA end-binding activity. Western blot analysis of human mitochondrial protein extracts using a monoclonal antibody specific for an N-terminal epitope of Ku80 identified a protein with an apparent molecular weight of 68 kDa. This mitochondrial protein was not detected by a monoclonal antibody specific for an epitope at the C-terminal end of Ku80. Consistently, while both the N- and C-terminal Ku80 monoclonal antibodies supershifted the nuclear DNA end-binding complex on an electrophoretic mobility shift assay, only the N-terminal monoclonal antibody supershifted the mitochondrial DNA end-binding complex. To confirm that the 68 kDa Ku protein was not a consequence of nuclear protein contamination of mitochondrial preparations, highly purified intact nuclei and mitochondria were treated with proteinase K which traverses the pores of intact nuclei but gains limited access into intact mitochondria. Ku80 in purified intact nuclei was sensitive to treatment with this protease, while the 68 kDa Ku protein characteristic of purified intact mitochondria was resistant. Further, immunocytochemical analysis revealed the co-localization of the N-terminal specific Ku80 monoclonal antibody with a mitochondrial-targeted green fluorescence protein. Mitochondrial localization of the C-terminal Ku80 monoclonal antibody was not observed. These data are consistent with the hypothesis that a C-terminally truncated form of Ku80 is localized in mammalian mitochondria where it functions in a DNA end-binding activity.

Journal ArticleDOI
TL;DR: The results suggest that the accessory subunit of mtDNA polymerase plays an important role in the control of mt DNA replication in Drosophila.