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At least 217 records · Page 12Linked to original sources
PubMed1997

The terminal DNA structure of mammalian chromosomes.

In virtually all eukaryotic organisms, telomeric DNA is composed of a variable number of short direct repeats. While the primary sequence of telomeric repeats has been determined for a great variety of species, the actual physical DNA structure at the ends of a bona fide metazoan chromosome with a centromere is unknown. It is shown here that an overhang of the strand forming the 3' ends of the chromosomes, the G-rich strand, is found at mammalian chromosome ends. Moreover, on at least some telomeres, the overhangs are > or = 45 bases long. Such surprisingly long overhangs were present on chromosomes derived from fully transformed tissue culture cells and normal G0-arrested peripheral leukocytes. Thus, irrespective of whether the cells were actively dividing or arrested, a very similar terminal DNA arrangement was found. These data suggest that the ends of mammalian and possibly all vertebrate chromosomes consist of an overhang of the G-rich strand and that these overhangs may be considerably larger than previously anticipated.

Animals↗
PubMed1999

Ultra-high resolution DNA structures.

This paper describes the progress in our efforts at producing ultra-high resolution (< 0.8 A) DNA structures using advanced cryo-crystallography and synchrotron. Our work is aimed at providing reliable geometric (bond length and bond angle), electronic and motional information of DNA molecules in different conformational contexts. These highly-reliable, new structures will be the basis for constructing better DNA force-field parameters, which will benefit the structural refinement of DNA, protein-DNA complexes, and ligand-DNA complexes.

Antibiotics, Antineoplastic↗
PubMed2000

Targeting DNA secondary structures.

DNA secondary structures containing regions of single-stranded DNA have now been identified in the genomic DNA of a number of prokaryotic and eukaryotic species, including humans. Many of these secondary structures are associated with regions of DNA involved in regulation of transcription: promoters or upstream elements. The secondary structures involved appear likely to be hairpin or cruciform structures that may be recognition sites for binding of transcription factors. In the case of the coliphage N4 virion RNA polymerase, a defined hairpin in the polymerase promoter necessary for binding of the polymerase and regulation of transcription has been shown to be extruded under physiological conditions in plasmid DNA. The presence of single-stranded DNA in the promoters of several species suggests that regulatory hairpins may be involved in transcription of a number of genes. In support of this, hairpin- or cruciform-binding proteins have been identified from several species. These results imply that secondary structures in regulatory regions may be targets for drugs that bind and either block or enhance binding of proteins involved in transcription. In this review, we discuss the evidence for DNA secondary structures, particularly hairpins and cruciforms, in genomic DNA and review the studies to date of development of small molecules that can selectively bind these structures.

Animals↗
PubMed1984

Patterns of DNA structural polymorphism and their evolutionary implications.

The pattern of sites within purified DNA that are highly susceptible to double-stranded cleavage by micrococcal nuclease has been analyzed in the vicinity of over 20 genes from widely separated loci in Drosophila. These genes have uniformly exhibited a distinctive organization of cleavage sites such that at early times of digestion major sites are observed in the spacer regions surrounding the genes, but not within the protein coding regions themselves. Examples examined include Drosophila genes for heat-shock proteins, cytoplasmic actin, ribosomal protein 49, alcohol dehydrogenase, Sgs 4 glue protein, and other developmentally regulated transcripts, a human beta-globin gene, and mouse alpha 3-globin pseudogene. It seems probable that this gene/spacer pattern will be a general one in the genomes of eucaryotes, but not in the genomes of procaryotes, since neither pBR322 nor phage lambda DNA display such a pattern. One observes a nonrandom spacing of strong cleavage sites in Drosophila DNA, with the most frequent intervals being 195 bp and 411 bp. Such a pattern of variation in DNA structure may have evolved to facilitate the packaging of eucaryotic DNA into chromatin.

Animals↗
PubMed1997

A particular DNA structure is required for the function of a cis-acting component of the Epstein-Barr virus OriLyt origin of replication.

OriLyt, thecis-acting element of Epstein-Barr virus lytic origin of replication, consists of upstream and downstream components. The upstream component plays a dual role in transcription and replication. The downstream component contains a homopurine-homopyrimidine sequence which forms an H palindrome. We show that the downstream component can adopt a triple helix structure in vitro, that the 5' border of the homopyrimidine sequence is sensitive to P1 nuclease when carried by a supercoiled plasmid and that an oligonucleotide complementary to the homopyrimidine strand is taken up by a plasmid carrying the OriLyt H palindrome. We also show that all mutations which alter the H palindrome impair both oligonucleotide uptake and OriLyt-dependent replication. Interestingly, compensatory mutations which restore an H palindrome also restore oligonucleotide uptake by the mutated plasmids and their OriLyt-dependent replication. Thus, there is a strong correlation between the inability of the OriLyt H palindrome to form a non-B-DNA structure in vitro and impairment of OriLyt-dependent replication. This suggests that the presence of a non-B-DNA structure in the OriLyt downstream component is required for OriLyt-dependent replication.

Base Sequence↗
PubMed1994

Triple-helix formation interferes with the transcription and hinged DNA structure of the interferon-inducible 6-16 gene promoter.

The interferon responsive element (IRE) of the 6-16 gene lies within two 39-bp elements in tandem. A purine-rich oligodeoxynucleotide, oligo(dN), was found to be able to pair with the purine-rich strand of the IRE in an antiparallel orientation which led to triple-helix formation with Mg2+ being necessary for triplex stability. Footprinting analysis confirmed these results. The interaction between the IRE and the oligo(dN) was reversible and had a Kd equal to 20 nM. The two repeats of the 6-16 gene IRE can form a hinged DNA structure through pairing of their purine-rich regions; exonuclease III experiments support this model. The hybrid DNA structure leads to a parallel pairing of the purine strands of the 6-16 gene IRE and this conformation was shown to be destabilized by triplex formation. When co-transfected with a reporter gene whose promoter was under the control of the 6-16 gene IRE, the triple-helix-forming oligo(dN)s inhibit the interferon-induced stimulation of the reporter gene with complete inhibition being obtained with 1 microM oligo(dN) at the time of transfection. When added to the cell culture medium after transfection, the concentrations of oligo(dN) needed to obtain 50% inhibition of the interferon effect on gene transcription must be 50-100 times higher. Besides the existence of a peculiar structure for the 6-16 gene IRE, the possibility of interfering with gene expression by means of oligo(dN)s is demonstrated.

Base Sequence↗
PubMedDate not supplied

[Interrelations between the content of reactive oxygen species and the state of DNA structure in bone marrow cells of mice after whole body gamma-irradiation].

The aim of the research is a further analysis of a problem concerning two (regulatory and damaging) functions of reactive oxygen species (ROS) in viability of organism cells under acute exposure to ionizing radiation. For this purpose the ROS content and the state of DNA structure in bone marrow cells of male CBA and SHK mice have been studied in dynamics, from 15 minutes up to 185 day after acute exposure to a sublethal dose (1.5 Gy) of ionizing radiation. The analysis of dependencies between these parameters in the norm, immediately after irradiation and in later cell descendants showed the direct correlation between the ROS content and the DNA nativity in the norm; 185 days after irradiation the correlation disappeared. It was suggested that the correlation occurred in the norm indicates participation of the ROS (as a sensory link) in a system of reactions (under the control of the corresponding genetic program), that ensure the DNA structure and, ultimately, the genome stability. The loss of such connection after acute exposure to ionizing radiation indicates actuation of another module of reactions sustaining stability of cellular genome in new conditions, without regulatory participation of ROS, that can promote or demonstrate the development of radiation-induced genome instability.

Animals↗
PubMed2001

The effect of DNA structure on the catalytic efficiency and fidelity of human DNA polymerase beta on templates with platinum-DNA adducts.

DNA adducts formed by platinum-based anticancer drugs interfere with DNA replication. The carrier ligand of the platinum compound is likely to affect the conformation of the Pt-DNA adducts. In addition, the conformation of the adduct can also change upon binding of damaged DNA to the active site of DNA polymerase. From the crystal structures of pol beta ternary complexes it is evident that undamaged gapped and primed single-stranded (non-gapped) DNA templates exist in very different conformations when bound to pol beta. Therefore, one might expect that the constraints imposed on the damaged templates by binding to the polymerase active site should also affect the conformation of the Pt-DNA adducts and their ability to inhibit DNA replication. In support of this hypothesis we have found that the efficiency, carrier ligand specificity, site of discrimination (3'-G versus 5'-G of the Pt-GG adducts), and fidelity of translesion synthesis past Pt-DNA adducts by pol beta are strongly affected by the structure of the DNA template. Previous studies have suggested that the conformation of Pt-DNA adducts may be affected by the sequence context of the adduct. In support of this hypothesis, our data show that sequence context affects the efficiency, fidelity, and pattern of misincorporation by pol beta.

Antineoplastic Agents↗
PubMedDate not supplied

[Evolution of the DNA structure: direction, mechanism, rate].

On the basis of the analysis of frequencies of occurence of pyrimidines of different length, the degree of clustering of DNA of a hundred species belonging to different taxons has been determined. A tendency towards increase in the index of DNA clustering was revealed in the sequence: bacteria, invertebrates, fishes, amphibians, reptiles, birds, mammals. A mechanism is postulated, according to which an increase in the degree of clustering of DNA in the process of progressive evolution of species may be due to accumulation of mutations, Pyr in equilibrium Pur transversions, resulting in an increase in the degree of asymmetry of the complementary chains of DNA. That this mechanism does exist is proved by a positive correlation between the degree of clustering of DNA and the degree of asymmetry of natural DNA chains. The mean frequency of mutation of vertebrates is about 4,6-10(-8) substitutions per nucleotide per year. Evolution of different groups of organisms may be accompanied with an increase in the rate of evolution of DNA structure. With the help of a special computer program, proceeding from the amino acid sequence of cytochromes c in 40 species belonging to different taxons, the degree of clustering of pyrimidines and the degree of asymmetry of complementary chains of DNA cistrons coding for cytochrome c was determined. A general tendency towards an increase in the mean values of the corresponding parametres of structure was found in the following: bacteria, invertebrates, fishes, amphibians, reptiles, birds and mammals. Thus, it was established that "neutral" amino acid substitutions in cytochromes are based on the selection of mutations leading to accumulation of pyrimidines in sense H-chain of DNA, and purines--in the corresponding mRNA. The frequency of mutation in cytochrome c of chordates is about 5,2-10(-8) of amino acid residues per year. It is assumed that the evolution modification of DNA structure may be due to increase in the disturbance stability of translation.

Amino Acids↗
PubMed1988

[Action of khanerol on the superhelical DNA structure of S-37 cells].

The damaging effects on the structure of a supramolecular DNA complex of sarcoma-37 cells in mice following the use of the therapeutic doses of khanerol were found. Capillary elastoviscosimetry and nucleoid sedimentation procedure showed that the damage of the supramolecular DNA complex structure becomes apparent already after 4 hours, and increases by 24 hour. In studies of the binding of khanerol with DNA by Cd and Tmelt. the procedure showed direct interaction of khanerol with DNA. The role of conformational changes of the supramolecular DNA complex in the cytotoxic action of antitumour agents is discussed.

Animals↗
PubMed2004

Highly polymorphic repeat region in the CETP promoter induces unusual DNA structure.

Genetic variation in the human cholesteryl ester transfer protein (CETP) promoter is associated with HDL cholesterol levels and cardiovascular disease with much of the genetic variation in CETP attributed to the promoter region. In this region, there are several single nucleotide polymorphisms as well as a variable length tandem repeat located 1946 base pairs upstream of the CETP transcription start that is highly polymorphic with respect to both length and sequence. There are more than 10 different long alleles and these vary in their repeat structure. We find that the short allele of this repeat is associated with high HDL cholesterol levels in vivo (P<0.0001). In males, this association is independent of the nearby -629 polymorphism. In addition, the variable length GAAA repeat can stimulate an adjacent GGGGA repeat to form a structure that hinders DNA amplification and sequencing. This structure also has an effect in vivo as shown by orientation effects and cloning efficiency in Escherichia coli.

Animals↗
PubMed2000

Unusual DNA structure of the diabetes susceptibility locus IDDM2 and its effect on transcription by the insulin promoter factor Pur-1/MAZ.

One of the loci responsible for genetic susceptibility to insulin-dependent diabetes mellitus (IDDM) is the insulin-linked polymorphic region (ILPR, also known as IDDM2). This polymorphic G-rich minisatellite, located in the promoter region of the human insulin gene, comprises a variable number of tandemly repeating sequences related to ACAGGGGTGTGGGG. An interesting characteristic of the ILPR is its ability to form unusual DNA structures in vitro, presumably through formation of G-quartets. This ability to form G-quartets raises the intriguing possibility that transcriptional activity of the insulin gene may in fact be influenced by the quaternary DNA topology of the ILPR. We now show that single nucleotide differences in the ILPR known to affect insulin transcription are correlated with ability to form unusual DNA structures. Through the design and testing of two high transcriptional activity ILPR repeats, we demonstrate that both inter- and intramolecular G-quartet formation in the ILPR can influence transcriptional activity of the human insulin gene, and thus, may contribute to that portion of diabetes susceptibility attributed to the IDDM2 locus.

DNA↗
PubMed1988

Bent DNA structures associated with several origins of replication are recognized by a unique enzyme from trypanosomatids.

Sequence-directed bending of the DNA double helix is a conformational variation found in both prokaryotic and eukaryotic organisms. The utilization of bent DNA structures from various sources as specific signals recognized by an enzyme is demonstrated here using a unique endonuclease purified from trypanosomatid cells. Crithidia fasciculata nicking enzyme was previously shown to recognize specifically the bent structure found in kinetoplast DNA minicircles. The binding constant measured for this specific interaction is of two orders of magnitude higher than that measured for the binding of the enzyme to a non-curved sequence. As determined by binding competition and mobility shift electrophoresis analyses, this enzyme recognizes the sequence-directed bends associated with the origins of replication of bacteriophage lambda and simian virus 40 (SV40), as well as that located within the autonomously replicating sequence (ARS1) region of the yeast S. cerevisiae.

Animals↗
PubMed1994

The yeast KEM1 gene encodes a nuclease specific for G4 tetraplex DNA: implication of in vivo functions for this novel DNA structure.

We have previously reported the identification of a G4-DNA-dependent nuclease from S. cerevisiae that recognizes a tetrastranded G4-DNA structure and cuts in a single-stranded region 5' to the G4 structure. We purify this activity to homogeneity and show it to be the product of the S. cerevisiae KEM1 gene, which is also known as SEP1, DST2, XRN1, and RAR5. Since a homozygous deletion of the KEM1 gene blocks meiotic cells at the 4N stage, the finding of these G4-dependent DNA binding and cleavage activities for the KEM1 gene product supports the hypothesis that G4-DNA may play a role in meiosis.

Amino Acid Sequence↗
PubMed2002

Visualization of hemiknot DNA structure with an atomic force microscope.

The hemiknot, a novel type of DNA structure in which a loop is stabilized by threading one end of the duplex through another, has been studied in this paper. The hemiknot was obtained by reassociation of a DNA fragment with (CA/TG)n inserts of different lengths. Slow and fast migrating products were purified by gel electrophoresis and imaged by atomic force microscopy (AFM) using the aminopropylsilatrane-mica technique for sample preparation. Slow migrating product was characterized by the formation of small blobs for the short insert (60 bp) and clear loops and other morphologies for the long insert (188 bp). These structural features were found in almost 100% of the molecules of the slow migrating sample and were not present in the control sample. Measurements showed that the location of the blobs coincided with the positions of the inserts. The sample with the 188 bp insert in the 573 bp fragment had large structural irregularities. The majority of the molecules (77%) had asymmetrically located loops. The location of the loop in the molecules correlated well with the position of the insert in the fragment. The measured sizes of the loops were in agreement with the insert size. Altogether, these data support the hypothesis for hemiknot formation suggested earlier. In addition to looped structures, other morphologies of the hemiknot were identified in AFM images. Possible models for hemiknot formation and structure are discussed.

Animals↗
PubMed2004

Stability and strand asymmetry in the non-B DNA structure at the bcl-2 major breakpoint region.

The t(14;18) translocation involving the Ig heavy chain locus and the BCL-2 gene is the single most common chromosomal translocation in human cancer. Recently we reported in vitro and in vivo chemical probing data indicating that the 150-bp major breakpoint region (Mbr), which contains three breakage subregions (hotspots) (known as peaks I, II, and III), has single-stranded character and hence a non-B DNA conformation. Although we could document the non-B DNA structure formation at the bcl-2 Mbr, the structural studies were limited to chemical probing. Therefore, in the present study, we used multiple methods including circular dichroism to detect the non-B DNA at the bcl-2 Mbr. We established a new gel shift method to detect the altered structure at neutral pH on shorter DNA fragments containing the bcl-2 Mbr and analyzed the fine structural features. We found that the single-stranded region in the non-B DNA structure observed is stable for days and is asymmetric with respect to the Watson and Crick strands. It could be detected by oligomer probing, a bisulfite modification assay, or a P1 nuclease assay. We provide evidence that two different non-B conformations exist at peak I in addition to the single one observed at peak III. Finally we used mutagenesis and base analogue incorporation to show that the non-B DNA structure formation requires Hoogsteen pairing. These findings place major constraints on the location and nature of the non-B conformations assumed at peaks I and III of the bcl-2 Mbr.

Base Sequence↗
PubMed1984

Osmium-induced alteration in DNA structure.

In the presence of pyridine and other ligands osmium tetroxide binds covalently to pyrimidine bases in DNA. Properties of osmium-modified native and denatured calf thymus DNA, and plasmid Co1E1 DNA were investigated by means of differential pulse polarography, absorption spectrophotometry, circular dichroism, agarose gel electrophoresis, and nuclease S1 digestion. A great difference in the reaction kinetics of native and denatured DNAs with osmium, pyridine was observed. On the ground of the slow stepwise reaction kinetics of native DNA in the initial stage of its modification by osmium it has been suggested that the primary reaction sites do not include bases contained in the intact double helix. Osmium binding to sporadic primary reaction sites (represented e.g. by bases in the vicinity of a single-strand break) in native calf thymus DNA resulted in local changes in DNA conformation limited to a close neighbourhood of the binding site. At higher osmium/nucleotide ratios disordering of the DNA structure over a region extending beyond the immediate binding site was observed. With denatured DNA the same type of structure disordering was detected already in the initial stage of the reaction at osmium/nucleotide ratios as low as 0.01. Osmium binding to the supercoiled Co1E1 DNA resulted in its relaxation without nicking and it increased its sensitivity to linearization by cleavage with nuclease S1. The behaviour of Co1E1 DNA has been explained by the formation of a denatured region in the molecule (accompanied by a coupled loss of duplex and superhelical turns). It has been suggested that osmium can be used to label and to visualize distorted regions in the DNA double helix.

Animals↗