Potential Quadruplex Sequences Research Articles

Telomeric retrotransposons show propensity to form G-quadruplexes in various eukaryotic species

BackgroundCanonical telomeres (telomerase-synthetised) are readily forming G-quadruplexes (G4) on the G-rich strand. However, there are examples of non-canonical telomeres among eukaryotes where telomeric tandem repeats are invaded by specific retrotransposons. Drosophila melanogaster represents an extreme example with telomeres composed solely by three retrotransposons—Het-A, TAHRE and TART (HTT). Even though non-canonical telomeres often show strand biased G-distribution, the evidence for the G4-forming potential is limited.ResultsUsing circular dichroism spectroscopy and UV absorption melting assay we have verified in vitro G4-formation in the HTT elements of D. melanogaster. Namely 3 in Het-A, 8 in TART and 2 in TAHRE. All the G4s are asymmetrically distributed as in canonical telomeres. Bioinformatic analysis showed that asymmetric distribution of potential quadruplex sequences (PQS) is common in telomeric retrotransposons in other Drosophila species. Most of the PQS are located in the gag gene where PQS density correlates with higher DNA sequence conservation and codon selection favoring G4-forming potential. The importance of G4s in non-canonical telomeres is further supported by analysis of telomere-associated retrotransposons from various eukaryotic species including green algae, Diplomonadida, fungi, insects and vertebrates. Virtually all analyzed telomere-associated retrotransposons contained PQS, frequently with asymmetric strand distribution. Comparison with non-telomeric elements showed independent selection of PQS-rich elements from four distinct LINE clades.ConclusionOur findings of strand-biased G4-forming motifs in telomere-associated retrotransposons from various eukaryotic species support the G4-formation as one of the prerequisites for the recruitment of specific retrotransposons to chromosome ends and call for further experimental studies.

Taq-Polymerase Stop Assay to Determine Target Selectivity of G4 Ligands in Native Promoter Sequences of MYC, TERT, and KIT Oncogenes.

Computational and high-throughput experimental methods predict thousands of potential quadruplex sequences (PQSs) in the human genome. Often these PQSs contain more than four G-runs, which introduce additional uncertainty into the conformational polymorphism of the G4 DNA. G4-specific ligands, which are currently being actively developed as potential anticancer agents or tools for studying G4 structures in genomes, may preferentially bind to specific G4 structures over the others that can be potentially formed in the extended G-rich genomic region. We propose a simple technique that identifies the sequences that tend to form G4 in the presence of potassium ions or a specific ligand. Thermostable DNA Taq-polymerase stop assay can detect the preferential position of the G4 -ligand binging within a long PQS-rich genomic DNA fragment. This technique was tested for four G4 binders PDS, PhenDC3, Braco-19, and TMPyP4 at three promoter sequences of MYC, KIT, and TERT that contain several PQSs each. We demonstrate that the intensity of polymerase pausing reveals the preferential binding of a ligand to particular G4 structures within the promoter. However, the strength of the polymerase stop at a specific site does not always correlate with the ligand-induced thermodynamic stabilization of the corresponding G4 structure.

Genome-wide analysis of 8-oxo-7,8-dihydro-2'-deoxyguanosine at single-nucleotide resolution unveils reduced occurrence of oxidative damage at G-quadruplex sites.

8-Oxo-7,8-dihydro-2′-deoxyguanosine (OG), one of the most common oxidative DNA damages, causes genome instability and is associated with cancer, neurological diseases and aging. In addition, OG and its repair intermediates can regulate gene transcription, and thus play a role in sensing cellular oxidative stress. However, the lack of methods to precisely map OG has hindered the study of its biological roles. Here, we developed a single-nucleotide resolution OG-sequencing method, named CLAPS-seq (Chemical Labeling And Polymerase Stalling Sequencing), to measure the genome-wide distribution of both exogenous and endogenous OGs with high specificity. Our data identified decreased OG occurrence at G-quadruplexes (G4s), in association with underrepresentation of OGs in promoters which have high GC content. Furthermore, we discovered that potential quadruplex sequences (PQSs) were hotspots of OGs, implying a role of non-G4-PQSs in OG-mediated oxidative stress response.

G-Quadruplexes in the Archaea Domain.

The importance of unusual DNA structures in the regulation of basic cellular processes is an emerging field of research. Amongst local non-B DNA structures, G-quadruplexes (G4s) have gained in popularity during the last decade, and their presence and functional relevance at the DNA and RNA level has been demonstrated in a number of viral, bacterial, and eukaryotic genomes, including humans. Here, we performed the first systematic search of G4-forming sequences in all archaeal genomes available in the NCBI database. In this article, we investigate the presence and locations of G-quadruplex forming sequences using the G4Hunter algorithm. G-quadruplex-prone sequences were identified in all archaeal species, with highly significant differences in frequency, from 0.037 to 15.31 potential quadruplex sequences per kb. While G4 forming sequences were extremely abundant in Hadesarchaea archeon (strikingly, more than 50% of the Hadesarchaea archaeon isolate WYZ-LMO6 genome is a potential part of a G4-motif), they were very rare in the Parvarchaeota phylum. The presence of G-quadruplex forming sequences does not follow a random distribution with an over-representation in non-coding RNA, suggesting possible roles for ncRNA regulation. These data illustrate the unique and non-random localization of G-quadruplexes in Archaea.

Stable G-Quadruplex Structures of Oncogene Promoters Induce Potassium-Dependent Stops of Thermostable DNA Polymerase.

Amplification of GC-rich regions of genomic DNA is hindered either by high stability of DNA double helix or as a result of alternative structure formation by a guanine-rich DNA strand. Such potential G-quadruplex (G4) sequences are fairly common in promoters of the human genome. The efficiency of PCR amplification of promoter sequences for several human oncogenes (MYC, NRAS, TERT, KRAS, KIT) was studied. We demonstrate that the efficiency of DNA polymerase is reduced in the presence of potassium ions. The primer-extension technique localized DNA polymerase stops at the 3'-ends of potential quadruplex sequences. The structural and thermodynamic properties of short G-rich oligonucleotides corresponding to the stops of DNA polymerase were analyzed. These oligonucleotides formed stable parallel G4 in the presence of potassium ions. Correlation between the stability of G4 structure and efficiency of DNA polymerase stops was revealed. The results provide a method for detecting new G4 structures in extended genomic sequences and also clarify the mechanism of inhibition of DNA polymerase in G-rich regions of DNA.

Guanine quadruplexes are formed by specific regions of human transposable elements

BackgroundTransposable elements form a significant proportion of eukaryotic genomes. Recently, Lexa et al. (Nucleic Acids Res 42:968-978, 2014) reported that plant long terminal repeat (LTR) retrotransposons often contain potential quadruplex sequences (PQSs) in their LTRs and experimentally confirmed their ability to adopt four-stranded DNA conformations.ResultsHere, we searched for PQSs in human retrotransposons and found that PQSs are specifically localized in the 3’-UTR of LINE-1 elements, in LTRs of HERV elements and are strongly accumulated in specific regions of SVA elements. Circular dichroism spectroscopy confirmed that most PQSs had adopted monomolecular or bimolecular guanine quadruplex structures. Evolutionarily young SVA elements contained more PQSs than older elements and their propensity to form quadruplex DNA was higher. Full-length L1 elements contained more PQSs than truncated elements; the highest proportion of PQSs was found inside transpositionally active L1 elements (PA2 and HS families).ConclusionsConservation of quadruplexes at specific positions of transposable elements implies their importance in their life cycle. The increasing quadruplex presence in evolutionarily young LINE-1 and SVA families makes these elements important contributors toward present genome-wide quadruplex distribution.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-1032) contains supplementary material, which is available to authorized users.

Quadruplex DNA: sequence, topology and structure

G-quadruplexes are higher-order DNA and RNA structures formed from G-rich sequences that are built around tetrads of hydrogen-bonded guanine bases. Potential quadruplex sequences have been identified in G-rich eukaryotic telomeres, and more recently in non-telomeric genomic DNA, e.g. in nuclease-hypersensitive promoter regions. The natural role and biological validation of these structures is starting to be explored, and there is particular interest in them as targets for therapeutic intervention. This survey focuses on the folding and structural features on quadruplexes formed from telomeric and non-telomeric DNA sequences, and examines fundamental aspects of topology and the emerging relationships with sequence. Emphasis is placed on information from the high-resolution methods of X-ray crystallography and NMR, and their scope and current limitations are discussed. Such information, together with biological insights, will be important for the discovery of drugs targeting quadruplexes from particular genes.

Highly prevalent putative quadruplex sequence motifs in human DNA

We report here the results of a systematic search for the existence and prevalence of potential intramolecular G-quadruplex forming sequences in the human genome. We have also examined the tendency for particular sequences of ‘loop’ regions to occur in particular positions with respect to the G-tracts in a quadruplex. Using arithmetic ratio and probability techniques we have discovered frequent and systematic occurrence of certain sequence types, the most prominent being a potential quadruplex containing CCTGT in the first ‘loop’ position. Being able to highlight types of potential quadruplex sequences in G-rich regions is an important step in searching for biologically relevant sequences and finding their function.