Hard/Soft hybrid modeling of temperature-induced unfolding processes involving G-quadruplex and i-motif nucleic acid structures

Abstract G-quadruplexes are noncanonical nucleic acid structures formed by stacked guanine tetrads. They are capable of a range of functions and thought to play widespread biological roles. This diversity raises an important question: what determines the biochemical specificity of G-quadruplex structures? The answer is particularly important from the perspective of biological regulation because genomes can contain hundreds of thousands of G-quadruplexes with a range of functions. Here we analyze the specificity of each sequence in a 496-member library of variants of a reference G-quadruplex with respect to five functions. Our analysis shows that the sequence requirements of G-quadruplexes with these functions are different from one another, with some mutations altering biochemical specificity by orders of magnitude. Mutations in tetrads have larger effects than mutations in loops, and changes in specificity are correlated with changes in multimeric state. To complement our biochemical data we determined the solution structure of a monomeric G-quadruplex from the library. The stacked and accessible tetrads rationalize why monomers tend to promote a model peroxidase reaction and generate fluorescence. Our experiments support a model in which the sequence requirements of G-quadruplexes with different functions are overlapping but distinct. This has implications for biological regulation, bioinformatics, and drug design.

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Oxadiazole/Pyridine-Based Ligands: A Structural Tuning for Enhancing G-Quadruplex Binding

Molecules ◽

10.3390/molecules23092162 ◽

2018 ◽

Vol 23 (9) ◽

pp. 2162 ◽

Cited By ~ 5

Author(s):

Filippo Doria ◽

Valentina Pirota ◽

Michele Petenzi ◽

Marie-Paule Teulade-Fichou ◽

Daniela Verga ◽

...

Keyword(s):

Nucleic Acid ◽

Structural Transition ◽

Binding Mode ◽

Telomeric Sequence ◽

Recognition Element ◽

New Family ◽

G Quadruplex ◽

Preferential Binding ◽

High Cytotoxicity ◽

Nucleic Acid Structures

Non-macrocyclic heteroaryls represent a valuable class of ligands for nucleic acid recognition. In this regard, non-macrocyclic pyridyl polyoxazoles and polyoxadiazoles were recently identified as selective G-quadruplex stabilizing compounds with high cytotoxicity and promising anticancer activity. Herein, we describe the synthesis of a new family of heteroaryls containing oxadiazole and pyridine moieties targeting DNA G-quadruplexes. To perform a structure–activity analysis identifying determinants of activity and selectivity, we followed a convergent synthetic pathway to modulate the nature and number of the heterocycles (1,3-oxazole vs. 1,2,4-oxadiazole and pyridine vs. benzene). Each ligand was evaluated towards secondary nucleic acid structures, which have been chosen as a prototype to mimic cancer-associated G-quadruplex structures (e.g., the human telomeric sequence, c-myc and c-kit promoters). Interestingly, heptapyridyl-oxadiazole compounds showed preferential binding towards the telomeric sequence (22AG) in competitive conditions vs. duplex DNA. In addition, G4-FID assays suggest a different binding mode from the classical stacking on the external G-quartet. Additionally, CD titrations in the presence of the two most promising compounds for affinity, TOxAzaPy and TOxAzaPhen, display a structural transition of 22AG in K-rich buffer. This investigation suggests that the pyridyl-oxadiazole motif is a promising recognition element for G-quadruplexes, combining seven heteroaryls in a single binding unit.

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G-Quadruplex-Forming Aptamers—Characteristics, Applications, and Perspectives

Molecules ◽

10.3390/molecules24203781 ◽

2019 ◽

Vol 24 (20) ◽

pp. 3781 ◽

Cited By ~ 27

Author(s):

Carolina Roxo ◽

Weronika Kotkowiak ◽

Anna Pasternak

Keyword(s):

Nucleic Acid ◽

Production Costs ◽

Biological Processes ◽

Disease Treatment ◽

Diagnostic Potential ◽

G Quadruplex ◽

Fast Development ◽

Systematic Evolution ◽

Nucleic Acid Structures ◽

Exponential Enrichment

G-quadruplexes constitute a unique class of nucleic acid structures formed by G-rich oligonucleotides of DNA- or RNA-type. Depending on their chemical nature, loops length, and localization in the sequence or structure molecularity, G-quadruplexes are highly polymorphic structures showing various folding topologies. They may be formed in the human genome where they are believed to play a pivotal role in the regulation of multiple biological processes such as replication, transcription, and translation. Thus, natural G-quadruplex structures became prospective targets for disease treatment. The fast development of systematic evolution of ligands by exponential enrichment (SELEX) technologies provided a number of G-rich aptamers revealing the potential of G-quadruplex structures as a promising molecular tool targeted toward various biologically important ligands. Because of their high stability, increased cellular uptake, ease of chemical modification, minor production costs, and convenient storage, G-rich aptamers became interesting therapeutic and diagnostic alternatives to antibodies. In this review, we describe the recent advances in the development of G-quadruplex based aptamers by focusing on the therapeutic and diagnostic potential of this exceptional class of nucleic acid structures.

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In-depth Bioinformatic Analyses of Human SARS-CoV-2, SARS-CoV, MERS-CoV, and Other Nidovirales Suggest Important Roles of Noncanonical Nucleic Acid Structures in Their Lifecycles

10.1101/2020.04.09.031252 ◽

2020 ◽

Author(s):

Martin Bartas ◽

Václav Brázda ◽

Natália Bohálová ◽

Alessio Cantara ◽

Adriana Volná ◽

...

Keyword(s):

Nucleic Acid ◽

Genomic Sequence ◽

Inverted Repeats ◽

Genomic Rna ◽

Molecular Processes ◽

Rna Packaging ◽

Binding Motifs ◽

G Quadruplex ◽

Human Proteins ◽

Nucleic Acid Structures

AbstractNoncanonical nucleic acid structures play important roles in the regulation of molecular processes. Considering the importance of the ongoing coronavirus crisis, we decided to evaluate genomes of all coronaviruses sequenced to date (stated more broadly, the order Nidovirales) to determine if they contain noncanonical nucleic acid structures. We discovered much evidence of putative G-quadruplex sites and even much more of inverted repeats (IRs) loci, which in fact are ubiquitous along the whole genomic sequence and indicate a possible mechanism for genomic RNA packaging. The most notable enrichment of IRs was found inside 5′UTR for IRs of size 12+ nucleotides, and the most notable enrichment of putative quadruplex sites (PQSs) was located before 3′UTR, inside 5′UTR, and before mRNA. This indicates crucial regulatory roles for both IRs and PQSs. Moreover, we found multiple G-quadruplex binding motifs in human proteins having potential for binding of SARS-CoV-2 RNA. Noncanonical nucleic acids structures in Nidovirales and in novel SARS-CoV-2 are therefore promising druggable structures that can be targeted and utilized in the future.

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S phase R-loop formation is restricted by PrimPol-mediated repriming

10.1101/318220 ◽

2018 ◽

Cited By ~ 2

Author(s):

Saša Šviković ◽

Alastair Crisp ◽

Sue Mei Tan-Wong ◽

Thomas A. Guilliam ◽

Aidan J. Doherty ◽

...

Keyword(s):

Nucleic Acid ◽

Secondary Structure ◽

Genetic Instability ◽

S Phase ◽

Loop Formation ◽

Dna Motifs ◽

G Quadruplex ◽

Nucleic Acid Structures ◽

Careful Management

SummaryDuring DNA replication, conflicts with ongoing transcription are frequent and require careful management to avoid genetic instability. R-loops, three stranded nucleic acid structures comprising a DNA:RNA hybrid and displaced single stranded DNA, are important drivers of damage arising from such conflicts. How R-loops stall replication and the mechanisms that restrain their formation during S phase are incompletely understood. Here we show in vivo how R-loop formation drives a short purine-rich repeat, (GAA)10, to become a replication impediment that requires the repriming activity of the primase-polymerase PrimPol for its processive replication. Further, we show that loss of PrimPol results in a significant increase in R-loop formation around the repeat during S phase. We extend this observation by showing that PrimPol suppresses R-loop formation in genes harbouring secondary structure-forming sequences, exemplified by G quadruplex and H-DNA motifs, across the genome in both avian and human cells. Thus, R-loops promote the creation of replication blocks at susceptible sequences, while PrimPol-dependent repriming limits the extent of unscheduled R-loop formation at these sequences, mitigating their impact on replication.

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Structure specific recognition of telomeric repeats containing RNA by the RGG-box of hnRNPA1

Nucleic Acids Research ◽

10.1093/nar/gkaa134 ◽

2020 ◽

Vol 48 (8) ◽

pp. 4492-4506 ◽

Cited By ~ 1

Author(s):

Meenakshi Ghosh ◽

Mahavir Singh

Keyword(s):

Phase Separation ◽

Nucleic Acid ◽

Liquid Phase ◽

Cell Interaction ◽

Protein Motifs ◽

G Quadruplex ◽

Separation Of Proteins ◽

Nucleic Acid Structures ◽

Dna Maintenance ◽

Liquid Liquid Phase Separation

Abstract The telomere repeats containing RNA (TERRA) is transcribed from the C-rich strand of telomere DNA and comprises of UUAGGG nucleotides repeats in humans. The TERRA RNA repeats can exist in single stranded, RNA-DNA hybrid and G-quadruplex forms in the cell. Interaction of TERRA RNA with hnRNPA1 has been proposed to play critical roles in maintenance of telomere DNA. hnRNPA1 contains an N-terminal UP1 domain followed by an RGG-box containing C-terminal region. RGG-motifs are emerging as key protein motifs that recognize the higher order nucleic acid structures as well as are known to promote liquid-liquid phase separation of proteins. In this study, we have shown that the RGG-box of hnRNPA1 specifically recognizes the TERRA RNA G-quadruplexes that have loops in their topology, whereas it does not interact with the single-stranded RNA. Our results show that the N-terminal UP1 domain in the presence of the RGG-box destabilizes the loop containing TERRA RNA G-quadruplex efficiently compared to the RNA G-quadruplex that lacks loops, suggesting that unfolding of G-quadruplex structures by UP1 is structure dependent. Furthermore, we have compared the telomere DNA and TERRA RNA G-quadruplex binding by the RGG-box of hnRNPA1 and discussed its implications in telomere DNA maintenance.

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Polyetheroaryl Oxadiazole/Pyridine-Based Ligands: A Structural Tuning for Enhancing G-Quadruplex Binding

10.20944/preprints201807.0110.v1 ◽

2018 ◽

Author(s):

Filippo Doria ◽

Valentina Pirota ◽

Michele Petenzi ◽

Marie-Paule Teulade Fichou ◽

Daniela Verga ◽

...

Keyword(s):

Nucleic Acid ◽

Binding Mode ◽

Telomeric Sequence ◽

Recognition Element ◽

Pyridine Ligands ◽

New Family ◽

G Quadruplex ◽

Preferential Binding ◽

High Cytotoxicity ◽

Nucleic Acid Structures

Acyclic olygoheteroaryl-based compounds represent a valuable class of ligands for nucleic acid recognition. In this regard, acyclic pyridyl polyoxazoles and polyoxadiazoles were recently identified as selective G-quadruplex stabilizing compounds with high cytotoxicity and promising anticancer activity. Herein, we describe the synthesis of a new family of polyheteroaryl oxadiazole/pyridine-ligands targeting DNA G-quadruplexes. In order to perform a structure-activity analysis identifying determinants of activity and selectivity, we followed a convergent synthetic pathway to modulate the nature and number of the heterocycles (1,3-oxazole vs 1,2,4-oxadiazole and pyridine vs benzene). Each ligand was evaluated towards secondary nucleic acid structures, which have been chosen as a prototype to mimic cancer-associated G-quadruplex structures (e.g., the human telomeric sequence, c-myc and c-kit promoters). Interesting, heptapyridyl-oxadiazole compounds showed preferential binding towards the telomeric sequence (22AG) in competitive conditions vs duplex DNA. In addition, G4-FID assays suggest a different binding mode from the classical stacking on the external G-quartet. Additionally, CD titrations in the presence of the two most promising compounds for affinity, TOxAzaPy and TOxAzaPhen, display a structural transition of 22AG in K-rich buffer. This investigation suggests that the pyridyl-oxadiazole motif is a promising recognition element for G-quadruplexes, combining seven heteroaryls in a single binding unit.

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Nucleic acid structures containing g-quadruplex and duplex motifs : a structural study and potential targeting approach

10.32657/10356/73601 ◽

2018 ◽

Author(s):

◽

Thi Quynh Ngoc Nguyen

Keyword(s):

Nucleic Acid ◽

Structural Study ◽

G Quadruplex ◽

Nucleic Acid Structures

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Natural Alkaloids and Heterocycles as G-Quadruplex Ligands and Potential Anticancer Agents

Molecules ◽

10.3390/molecules23020493 ◽

2018 ◽

Vol 23 (2) ◽

pp. 493 ◽

Cited By ~ 11

Author(s):

◽

...

Keyword(s):

Nucleic Acid ◽

Anticancer Agents ◽

Cancer Therapeutics ◽

Mechanisms Of Action ◽

Replication Initiation ◽

Untranslated Regions ◽

Promoter Regions ◽

Functional Regions ◽

G Quadruplex ◽

Nucleic Acid Structures

G-quadruplexes are four-stranded nucleic acid secondary structures that are formed in guanine-rich sequences. G-quadruplexes are widely distributed in functional regions of the human genome and transcriptome, such as human telomeres, oncogene promoter regions, replication initiation sites, and untranslated regions. Many G-quadruplex-forming sequences are found to be associated with cancer, and thus, these non-canonical nucleic acid structures are considered to be attractive molecular targets for cancer therapeutics with novel mechanisms of action. In this mini review, we summarize recent advances made by our lab in the study of G-quadruplex-targeted natural alkaloids and their derivatives toward the development of potential anticancer agents.

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