Genomic Instability of G-Quadruplex Sequences in : Roles of DinG, RecG, and RecQ Helicases.

Guanine-rich DNA can fold into highly stable four-stranded DNA structures called G-quadruplexes (G4). Originally identified in sequences from telomeres and oncogene promoters, they can alter DNA metabolism. Indeed, G4-forming sequences represent obstacles for the DNA polymerase, with important consequences for cell life as they may lead to genomic instability.

NACDDB: Nucleic Acid Circular Dichroism Database.

The Nucleic Acid Circular Dichroism Database (NACDDB) is a public repository that archives and freely distributes circular dichroism (CD) and synchrotron radiation CD (SRCD) spectral data about nucleic acids, and the associated experimental metadata, structural models, and links to literature. NACDDB covers CD data for various nucleic acid molecules, including DNA, RNA, DNA/RNA hybrids, and various nucleic acid derivatives. The entries are linked to primary sequence and experimental structural data, as well as to the literature.

Crucial Role of the C-Terminal Domain of Hfq Protein in Genomic Instability.

G-rich DNA repeats that can form G-quadruplex structures are prevalent in bacterial genomes and are frequently associated with regulatory regions of genes involved in virulence, antigenic variation, and antibiotic resistance. These sequences are also inherently mutagenic and can lead to changes affecting cell survival and adaptation. Transcription of the G-quadruplex-forming repeat (GT) in , when mRNA comprised the G-rich strand, promotes G-quadruplex formation in DNA and increases rates of deletion of G-quadruplex-forming sequences.

Role of Hfq in Genome Evolution: Instability of G-Quadruplex Sequences in .

Certain G-rich DNA repeats can form quadruplex in bacterial chromatin that can present blocks to DNA replication and, if not properly resolved, may lead to mutations. To understand the participation of quadruplex DNA in genomic instability in (), mutation rates were measured for quadruplex-forming DNA repeats, including (GT), (GT), and a RET oncogene sequence, cloned as the template or nontemplate strand. We evidence that these alternative structures strongly influence mutagenesis rates.

Novel computational study on π-stacking to understand mechanistic interactions of Tryptanthrin analogues with DNA.

Based on recently published initial experimental results on the intercalation of a class of broad spectrum antiparasitic compounds, we present a purely theoretical approach for determining if these compounds may preferentially intercalate with guanosine/cytosine (GC)-rich or adenosine/thymidine (TA)-rich regions of DNA. The predictive model presented herein is based upon utilization of density functional theory (DFT) to determine a priori how the best intercalator may energetically and sterically interact with each of the nucleoside base pairs. A potential new method using electrostatic potential maps (EPMs) to visually select the best poses is introduced and compared to the existing brute-force center of mass (COM) approach.

Replication fork stalling and checkpoint activation by a PKD1 locus mirror repeat polypurine-polypyrimidine (Pu-Py) tract.

DNA sequences prone to forming noncanonical structures (hairpins, triplexes, G-quadruplexes) cause DNA replication fork stalling, activate DNA damage responses, and represent hotspots of genomic instability associated with human disease. The 88-bp asymmetric polypurine-polypyrimidine (Pu-Py) mirror repeat tract from the human polycystic kidney disease (PKD1) intron 21 forms non-B DNA secondary structures in vitro. We show that the PKD1 mirror repeat also causes orientation-dependent fork stalling during replication in vitro and in vivo.

On an unbiased and consistent estimator for mutation rates.

Spontaneous mutations are stochastic events. The mutation rate, defined as mutations per genome per replication, is generally very low, and it is widely accepted that spontaneous mutations occur at defined, but different, rates in bacteriophage and in bacterial, insect, and mammalian cells. The calculation of mutation rates has proved to be a significant problem.

Replication-dependent instability at (CTG) x (CAG) repeat hairpins in human cells.

Instability of (CTG) x (CAG) microsatellite trinucleotide repeat (TNR) sequences is responsible for more than a dozen neurological or neuromuscular diseases. TNR instability during DNA synthesis is thought to involve slipped-strand or hairpin structures in template or nascent DNA strands, although direct evidence for hairpin formation in human cells is lacking. We have used targeted recombination to create a series of isogenic HeLa cell lines in which (CTG) x (CAG) repeats are replicated from an ectopic copy of the Myc (also known as c-myc) replication origin.

Antimicrobial activity of tryptanthrins in Escherichia coli.

Tryptanthrins have potential therapeutic activity against a wide variety of pathogenic organisms, although little is known about their mechanism. Activity against Escherichia coli, however, has not been examined. The effects of tryptanthrin (indolo[2,1-b]quinazolin-6,12-dione) and nine derivatives on growth, survival, and mutagenesis in E.

A Z-DNA sequence reduces slipped-strand structure formation in the myotonic dystrophy type 2 (CCTG) x (CAGG) repeat.

All DNA repeats known to undergo expansion leading to human neurodegenerative disease can form one, or several, alternative conformations, including hairpin, slipped strand, triplex, quadruplex, or unwound DNA structures. These alternative structures may interfere with the normal cellular processes of transcription, DNA repair, replication initiation, or polymerase elongation and thereby contribute to the genetic instability of these repeat tracts. We show that (CCTG) x (CAGG) repeats, in the first intron of the ZNF9 gene associated with myotonic dystrophy type 2, form slipped-strand DNA structures in a length-dependent fashion upon reduplexing.

Unstable spinocerebellar ataxia type 10 (ATTCT*(AGAAT) repeats are associated with aberrant replication at the ATX10 locus and replication origin-dependent expansion at an ectopic site in human cells.

Spinocerebellar ataxia type 10 (SCA10) is associated with expansion of (ATTCT)n repeats (where n is the number of repeats) within the ataxin 10 (ATX10/E46L) gene. The demonstration that (ATTCT)n tracts can act as DNA unwinding elements (DUEs) in vitro has suggested that aberrant replication origin activity occurs at expanded (ATTCT)n tracts and may lead to their instability. Here, we confirm these predictions.

Slipped strand DNA structures.

Slipped strand DNA structures are formed when complementary strands comprising direct repeats pair in a misaligned, or slipped, fashion along the DNA helix axis. Although slipped strand DNA may form in almost any direct repeat, to date, these structures have only been detected in short DNA repeats, termed unstable DNA repeats, in which expansion is associated with many neurodegenerative diseases. This alternative DNA structure, or a similar slipped intermediate DNA that may form during DNA replication or repair, may be a causative factor in the instability of the DNA sequences that can form these structures.

Target DNA structure plays a critical role in RAG transposition.

Antigen receptor gene rearrangements are initiated by the RAG1/2 protein complex, which recognizes specific DNA sequences termed RSS (recombination signal sequences). The RAG recombinase can also catalyze transposition: integration of a DNA segment bounded by RSS into an unrelated DNA target. For reasons that remain poorly understood, such events occur readily in vitro, but are rarely detected in vivo.

(CAG)*(CTG) repeats associated with neurodegenerative diseases are stable in the Escherichia coli chromosome.

(CAG)(n)*(CTG)(n) expansion is associated with many neurodegenerative diseases. Repeat instability has been extensively studied in bacterial plasmids, where repeats undergo deletion at high rates. We report an assay for (CAG)(n)*(CTG)(n) deletion from the chloramphenicol acetyltransferase gene integrated into the Escherichia coli chromosome.

Replication restart: a pathway for (CTG).(CAG) repeat deletion in Escherichia coli.

(CTG)n.(CAG)n repeats undergo deletion at a high rate in plasmids in Escherichia coli in a process that involves RecA and RecB. In addition, DNA replication fork progression can be blocked during synthesis of (CTG)n.

DNA strand arrangement within the SfiI-DNA complex: atomic force microscopy analysis.

The SfiI restriction enzyme binds to DNA as a tetramer holding two usually distant DNA recognition sites together before cleavage of the four DNA strands. To elucidate structural properties of the SfiI-DNA complex, atomic force microscopy (AFM) imaging of the complexes under noncleaving conditions (Ca2+ instead of Mg2+ in the reaction buffer) was performed. Intramolecular complexes formed by protein interaction between two binding sites in one DNA molecule (cis interaction) as well as complexes formed by the interaction of two sites in different molecules (trans interaction) were analyzed.

Generation of long tracts of disease-associated DNA repeats.

The generation of long uninterrupted DNA repeats is important for the study of repeat instability associated with several human genetic diseases, including myotonic dystrophy type 1. However, obtaining defined lengths of long repeats in vitro has been problematic. Strand slippage and/or DNA secondary structure formation may prevent efficient ligation.

Duplications between direct repeats stabilized by DNA secondary structure occur preferentially in the leading strand during DNA replication.

To ascertain a leading or lagging strand preference for duplication mutations, several short DNA sequences, i.e. mutation inserts, were designed that should demonstrate an asymmetric propensity for duplication mutations in the two complementary DNA strands during replication.

Chemotherapeutic deletion of CTG repeats in lymphoblast cells from DM1 patients.

Myotonic dystrophy type 1 (DM1) is caused by the expansion of a (CTG).(CAG) repeat in the DMPK gene on chromosome 19q13.3.

Genetic recombination destabilizes (CTG)n.(CAG)n repeats in E. coli.

The expansion of trinucleotide repeats has been implicated in 17 neurological diseases to date. Factors leading to the instability of trinucleotide repeat sequences have thus been an area of intense interest. Certain genes involved in mismatch repair, recombination, nucleotide excision repair, and replication influence the instability of trinucleotide repeats in both Escherichia coli and yeast.

Interaction of the Zalpha domain of human ADAR1 with a negatively supercoiled plasmid visualized by atomic force microscopy.

Interest to the left-handed DNA conformation has been recently boosted by the findings that a number of proteins contain the Zalpha domain, which has been shown to specifically recognize Z-DNA. The biological function of Zalpha is presently unknown, but it has been suggested that it may specifically direct protein regions of Z-DNA induced by negative supercoiling in actively transcribing genes. Many studies, including a crystal structure in complex with Z-DNA, have focused on the human ADAR1 Zalpha domain in isolation.

Supercoiling-induced DNA bending.

Local DNA bending is a critical factor for numerous DNA functions including recognition of DNA by sequence-specific regulatory binding proteins. Negative DNA supercoiling increases both local and global DNA dynamics, and this dynamic flexibility can facilitate the formation of DNA-protein complexes. We have recently shown that apexes of supercoiled DNA molecules are sites that can promote the formation of an alternative DNA structure, a cruciform, suggesting that these positions in supercoiled DNA are under additional stress and perhaps have a distorted DNA geometry.