scholarly journals Identification of optimal fluorescent probes for G-quadruplex nucleic acids through systematic exploration of mono- and distyryl dye libraries

2019 ◽  
Vol 15 ◽  
pp. 1872-1889 ◽  
Author(s):  
Xiao Xie ◽  
Michela Zuffo ◽  
Marie-Paule Teulade-Fichou ◽  
Anton Granzhan
Keyword(s):  
Rna Structure ◽  
Stokes Shift ◽  
Topological Classification ◽  
Fluorescent Light ◽  
Styryl Dyes ◽  
Dna And Rna ◽  
Double Stranded Dna ◽  
G4 Dna ◽  
G Quadruplex

A library of 52 distyryl and 9 mono-styryl cationic dyes was synthesized and investigated with respect to their optical properties, propensity to aggregation in aqueous medium, and capacity to serve as fluorescence “light-up” probes for G-quadruplex (G4) DNA and RNA structures. Among the 61 compounds, 57 dyes showed preferential enhancement of fluorescence intensity in the presence of one or another G4-DNA or RNA structure, while no dye displayed preferential response to double-stranded DNA or single-stranded RNA analytes employed at equivalent nucleotide concentration. Thus, preferential fluorimetric response towards G4 structures appears to be a common feature of mono- and distyryl dyes, including long-known mono-styryl dyes used as mitochondrial probes or protein stains. However, the magnitude of the G4-induced “light-up” effect varies drastically, as a function of both the molecular structure of the dyes and the nature or topology of G4 analytes. Although our results do not allow to formulate comprehensive structure–properties relationships, we identified several structural motifs, such as indole- or pyrrole-substituted distyryl dyes, as well as simple mono-stryryl dyes such as DASPMI [2-(4-(dimethylamino)styryl)-1-methylpyridinium iodide] or its 4-isomer, as optimal fluorescent light-up probes characterized by high fluorimetric response (I/I 0 of up to 550-fold), excellent selectivity with respect to double-stranded DNA or single-stranded RNA controls, high quantum yield in the presence of G4 analytes (up to 0.32), large Stokes shift (up to 150 nm) and, in certain cases, structural selectivity with respect to one or another G4 folding topology. These dyes can be considered as promising G4-responsive sensors for in vitro or imaging applications. As a possible application, we implemented a simple two-dye fluorimetric assay allowing rapid topological classification of G4-DNA structures.

scholarly journals A Novel G-Quadruplex Binding Protein in Yeast—Slx9

Molecules ◽  
2019 ◽  
Vol 24 (9) ◽  
pp. 1774 ◽  
Author(s):  
Silvia Götz ◽  
Satyaprakash Pandey ◽  
Sabrina Bartsch ◽  
Stefan Juranek ◽  
Katrin Paeschke
Keyword(s):  
Binding Protein ◽  
Sequence Motifs ◽  
Dna And Rna ◽  
G4 Dna ◽  
G Quadruplex ◽  
High Thermodynamic Stability ◽  
Guanine Base ◽  
Regulatory Functions

G-quadruplex (G4) structures are highly stable four-stranded DNA and RNA secondary structures held together by non-canonical guanine base pairs. G4 sequence motifs are enriched at specific sites in eukaryotic genomes, suggesting regulatory functions of G4 structures during different biological processes. Considering the high thermodynamic stability of G4 structures, various proteins are necessary for G4 structure formation and unwinding. In a yeast one-hybrid screen, we identified Slx9 as a novel G4-binding protein. We confirmed that Slx9 binds to G4 DNA structures in vitro. Despite these findings, Slx9 binds only insignificantly to G-rich/G4 regions in Saccharomyces cerevisiae as demonstrated by genome-wide ChIP-seq analysis. However, Slx9 binding to G4s is significantly increased in the absence of Sgs1, a RecQ helicase that regulates G4 structures. Different genetic and molecular analyses allowed us to propose a model in which Slx9 recognizes and protects stabilized G4 structures in vivo.

scholarly journals G-quadruplex DNA inhibits unwinding activity but promotes liquid–liquid phase separation by the DEAD-box helicase Ded1p

2021 ◽  
Author(s):  
Jun Gao ◽  
Zhaofeng Gao ◽  
Andrea A. Putnam ◽  
Alicia K. Byrd ◽  
Sarah L. Venus ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Liquid Phase Separation ◽  
Intrinsically Disordered ◽  
Dead Box ◽  
G4 Dna ◽  
G Quadruplex ◽  
The Dead ◽  
Liquid Liquid Phase Separation

G-quadruplex (G4) DNA inhibits RNA unwinding activity but promotes liquid–liquid phase separation of the DEAD-box helicase Ded1p in vitro and in cells. This highlights multifaceted effects of G4DNA on an enzyme with intrinsically disordered domains.

scholarly journals G-quadruplex DNA drives genomic instability and represents a targetable molecular abnormality in ATRX-deficient malignant glioma

2018 ◽  
Author(s):  
Yuxiang Wang ◽  
Jie Yang ◽  
Wei Wu ◽  
Rachna Shah ◽  
Carla Danussi ◽  
...  
Keyword(s):  
Dna Damage ◽  
Malignant Glioma ◽  
Model Systems ◽  
Copy Number Alterations ◽  
Molecular Alteration ◽  
Quadruplex Dna ◽  
G4 Dna ◽  
G Quadruplex

AbstractMutational inactivation of ATRX (α-thalassemia mental retardation X-linked) represents a defining molecular alteration in large subsets of malignant glioma. Yet the pathogenic consequences of ATRX deficiency remain unclear, as do tractable mechanisms for its therapeutic targeting. Here we report that ATRX loss in isogenic glioma model systems induces replication stress and DNA damage by way of G-quadruplex (G4) DNA secondary structure. Moreover, these effects are associated with the acquisition of disease-relevant copy number alterations over time. We then demonstrate, both in vitro and in vivo, that ATRX deficiency selectively enhances DNA damage and cell death following chemical G4 stabilization. Finally, we show that G4 stabilization synergizes with other DNA-damaging therapies, including ionizing radiation, in the ATRX-deficient context. Our findings reveal novel pathogenic mechanisms driven by ATRX deficiency in glioma, while also pointing to tangible strategies for drug development.

scholarly journals The Functional Consequences of Eukaryotic Topoisomerase 1 Interaction with G-Quadruplex DNA

Genes ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 193
Author(s):  
Alexandra Berroyer ◽  
Nayun Kim
Keyword(s):  
Topoisomerase I ◽  
Genome Instability ◽  
Strand Break ◽  
Dna Topology ◽  
Single Strand Break ◽  
Dna Structures ◽  
G4 Dna ◽  
G Quadruplex

Topoisomerase I in eukaryotic cells is an important regulator of DNA topology. Its catalytic function is to remove positive or negative superhelical tension by binding to duplex DNA, creating a reversible single-strand break, and finally religating the broken strand. Proper maintenance of DNA topological homeostasis, in turn, is critically important in the regulation of replication, transcription, DNA repair, and other processes of DNA metabolism. One of the cellular processes regulated by the DNA topology and thus by Topoisomerase I is the formation of non-canonical DNA structures. Non-canonical or non-B DNA structures, including the four-stranded G-quadruplex or G4 DNA, are potentially pathological in that they interfere with replication or transcription, forming hotspots of genome instability. In this review, we first describe the role of Topoisomerase I in reducing the formation of non-canonical nucleic acid structures in the genome. We further discuss the interesting recent discovery that Top1 and Top1 mutants bind to G4 DNA structures in vivo and in vitro and speculate on the possible consequences of these interactions.

scholarly journals Concurrent binding to DNA and RNA facilitates the pluripotency reprogramming activity of Sox2

2020 ◽  
Vol 48 (7) ◽  
pp. 3869-3887 ◽  
Author(s):  
Linlin Hou ◽  
Yuanjie Wei ◽  
Yingying Lin ◽  
Xiwei Wang ◽  
Yiwei Lai ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Target Genes ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
High Mobility ◽  
Binding Motif ◽  
Dna And Rna ◽  
Somatic Cell Reprogramming ◽  
Double Stranded Dna

Abstract Some transcription factors that specifically bind double-stranded DNA appear to also function as RNA-binding proteins. Here, we demonstrate that the transcription factor Sox2 is able to directly bind RNA in vitro as well as in mouse and human cells. Sox2 targets RNA via a 60-amino-acid RNA binding motif (RBM) positioned C-terminally of the DNA binding high mobility group (HMG) box. Sox2 can associate with RNA and DNA simultaneously to form ternary RNA/Sox2/DNA complexes. Deletion of the RBM does not affect selection of target genes but mitigates binding to pluripotency related transcripts, switches exon usage and impairs the reprogramming of somatic cells to a pluripotent state. Our findings designate Sox2 as a multi-functional factor that associates with RNA whilst binding to cognate DNA sequences, suggesting that it may co-transcriptionally regulate RNA metabolism during somatic cell reprogramming.

scholarly journals Packaging of Genomic RNA in Positive-Sense Single-Stranded RNA Viruses: A Complex Story

Viruses ◽  
2019 ◽  
Vol 11 (3) ◽  
pp. 253 ◽  
Author(s):  
Mauricio Comas-Garcia
Keyword(s):  
Rna Viruses ◽  
Genomic Rna ◽  
Rna Packaging ◽  
Dna And Rna ◽  
Relative Contribution ◽  
Double Stranded Dna ◽  
Positive Sense ◽  
Infectious Cycle

The packaging of genomic RNA in positive-sense single-stranded RNA viruses is a key part of the viral infectious cycle, yet this step is not fully understood. Unlike double-stranded DNA and RNA viruses, this process is coupled with nucleocapsid assembly. The specificity of RNA packaging depends on multiple factors: (i) one or more packaging signals, (ii) RNA replication, (iii) translation, (iv) viral factories, and (v) the physical properties of the RNA. The relative contribution of each of these factors to packaging specificity is different for every virus. In vitro and in vivo data show that there are different packaging mechanisms that control selective packaging of the genomic RNA during nucleocapsid assembly. The goals of this article are to explain some of the key experiments that support the contribution of these factors to packaging selectivity and to draw a general scenario that could help us move towards a better understanding of this step of the viral infectious cycle.

scholarly journals Human Rev1 polymerase disrupts G-quadruplex DNA

2013 ◽  
Vol 42 (5) ◽  
pp. 3272-3285 ◽  
Author(s):  
Sarah Eddy ◽  
Amit Ketkar ◽  
Maroof K. Zafar ◽  
Leena Maddukuri ◽  
Jeong-Yun Choi ◽  
...  
Keyword(s):  
Genome Maintenance ◽  
Quadruplex Dna ◽  
Dna Structures ◽  
G4 Dna ◽  
G Quadruplex ◽  
Steady State Rate ◽  
Dna Substrates ◽  
Successful Replication ◽  
Y Family

Abstract The Y-family DNA polymerase Rev1 is required for successful replication of G-quadruplex DNA (G4 DNA) in higher eukaryotes. Here we show that human Rev1 (hRev1) disrupts G4 DNA structures and prevents refolding in vitro. Nucleotidyl transfer by hRev1 is not necessary for mechanical unfolding to occur. hRev1 binds G4 DNA substrates with Kd,DNA values that are 4–15-fold lower than those of non-G4 DNA substrates. The pre-steady-state rate constant of deoxycytidine monophosphate (dCMP) insertion opposite the first tetrad-guanine by hRev1 is ∼56% as fast as that observed for non-G4 DNA substrates. Thus, hRev1 can promote fork progression by either dislodging tetrad guanines to unfold the G4 DNA, which could assist in extension by other DNA polymerases, or hRev1 can prevent refolding of G4 DNA structures. The hRev1 mechanism of action against G-quadruplexes helps explain why replication progress is impeded at G4 DNA sites in Rev1-deficient cells and illustrates another unique feature of this enzyme with important implications for genome maintenance.

scholarly journals Mammalian CST averts replication failure by preventing G-quadruplex accumulation

2018 ◽  
Author(s):  
Yang Liu ◽  
Miaomiao Zhang ◽  
Bing Wang ◽  
Yingnan Xiao ◽  
Tingfang Li ◽  
...  
Keyword(s):  
Replication Fork ◽  
Genome Integrity ◽  
G4 Dna ◽  
Genome Wide ◽  
G Quadruplex ◽  
Preferential Loss ◽  
Telomere Replication ◽  
Lagging Strand

AbstractHuman CST (CTC1-STN1-TEN1) is an RPA-like complex that associates with G-rich single-strand DNA and helps resolve replication problems both at telomeres and genome-wide. We previously showed that CST binds and disrupts G-quadruplex (G4) DNA in vitro, suggesting that CST may prevent in vivo blocks to replication by resolving G4 structures. Here, we demonstrate that CST binds and unfolds G4 with similar efficiency to RPA. In cells, CST is recruited to telomeric and non-telomeric chromatin upon G4 stabilization. STN1 depletion increases G4 accumulation and slows bulk genomic DNA replication. At telomeres, combined STN1 depletion and G4 stabilization causes multi-telomere FISH signals and telomere loss, hallmarks of deficient telomere duplex replication. Strand-specific telomere FISH indicates preferential loss of C-strand DNA while analysis of BrdU uptake during leading and lagging-strand telomere replication shows preferential under-replication of lagging telomeres. Together these results indicate a block to Okazaki fragment synthesis. Overall, our findings indicate a novel role for CST in maintaining genome integrity through resolution of G4 structures both ahead of the replication fork and on the lagging strand template.

scholarly journals Response of Sulfolobus solfataricus Dpo4 polymerase in vitro to a DNA G-quadruplex

Mutagenesis ◽  
2019 ◽  
Vol 34 (3) ◽  
pp. 289-297 ◽  
Author(s):  
Alexandra Berroyer ◽  
Gloria Alvarado ◽  
Erik D Larson
Keyword(s):  
Repetitive Dna ◽  
Dna Sequences ◽  
Sulfolobus Solfataricus ◽  
Repetitive Dna Sequences ◽  
Polymerase Eta ◽  
G4 Dna ◽  
G Quadruplex ◽  
Dna Polymerase Iv ◽  
Y Family

Abstract Repetitive DNA sequences support the formation of structures that can interrupt replication and repair, leading to breaks and mutagenesis. One particularly stable structure is G-quadruplex (G4) DNA, which is four-stranded and formed from tandemly repetitive guanine bases. When folded within a template, G4 interferes with DNA synthesis. Similar to non-duplex structures, DNA base lesions can also halt an advancing replication fork, but the Y-family polymerases solve this problem by bypassing the damage. In order to better understand how guanine-rich DNA is replicated, we have investigated the activity of the model Y-family polymerase, Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4), on guanine-rich templates in vitro. We find that Dpo4 progression on templates containing either a single GC-rich hairpin or a G4 DNA structure is greatly reduced and synthesis stalls at the structure. Human polymerase eta (hPol eta) showed the same pattern of stalling at G4; however, and in contrast to Klenow, hPol eta and Dpo4 partially synthesise into the guanine repeat. Substitution of the nucleotide selectivity residue in Dpo4 with alanine permitted ribonucleotide incorporation on unstructured templates, but this further reduced the ability of Dpo4 to synthesise across from the guanine repeats. The advancement of Dpo4 on G4 templates was highest when the reaction was supplied with only deoxycytidine triphosphate, suggesting that high-fidelity synthesis is favoured over misincorporation. Our results are consistent with a model where the Y-family polymerases pause upon encountering G4 structures but have an ability to negotiate some synthesis through tetrad-associated guanines. This suggests that the Y-family polymerases reduce mutagenesis by catalysing the accurate replication of repetitive DNA sequences, but most likely in concert with additional replication and structure resolution activities.

scholarly journals Exploring the preferential interaction of quercetin with VEGF promoter G-quadruplex DNA and construction of a pH-dependent DNA-based logic gate

RSC Advances ◽  
2017 ◽  
Vol 7 (59) ◽  
pp. 37230-37240 ◽  
Author(s):  
Snehasish Bhattacharjee ◽  
Pradeep K. Sengupta ◽  
Sudipta Bhowmik
Keyword(s):  
Logic Gate ◽  
Quadruplex Dna ◽  
Preferential Interaction ◽  
Double Stranded Dna ◽  
G4 Dna ◽  
G Quadruplex ◽  
Flavonoid Quercetin ◽  
Ph Dependent

The plant flavonoid quercetin (Que) binds more efficiently to VEGF G-quadruplex DNA (G4–DNA) compared to double stranded DNA as well as other G4–DNAs.

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