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

2020 ◽  
Vol 48 (8) ◽  
pp. 4492-4506 ◽  
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.

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.

Quadruplex Folding of DNA Promotes the Condensation of Linker Histones via Liquid-Liquid Phase Separation

2020 ◽  
Author(s):  
Masahiro Mimura ◽  
Shunsuke Tomita ◽  
Yoichi Shinkai ◽  
Kentaro Shiraki ◽  
Ryoji Kurita
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Electrostatic Interactions ◽  
Droplet Formation ◽  
Liquid Phase Separation ◽  
Intrinsically Disordered ◽  
G Quadruplex ◽  
Dna Quadruplex ◽  
Liquid Liquid Phase Separation

<p>Liquid-liquid phase separation (LLPS) of proteins and DNA has recently emerged as a possible mechanism underlying the dynamic organization of chromatin. We herein report the role of DNA quadruplex folding in liquid droplet formation via LLPS induced by interactions between DNA and linker histone H1 (H1), a key regulator of chromatin organization. Fluidity measurements inside the droplets and binding assays using G-quadruplex-selective probes demonstrated that quadruplex DNA structures, such as the G-quadruplex and i-motif, promote droplet formation with H1 and decrease molecular motility within droplets. The dissolution of the droplets in the presence of additives indicated that in addition to electrostatic interactions between the DNA and the intrinsically disordered region of H1, π-π stacking between quadruplex DNAs could potentially drive droplet formation. Given that DNA quadruplex structures are well documented in heterochromatin regions, it is imperative to understand the role of DNA quadruplex folding in the context of intranuclear LLPS.<b></b></p>

scholarly journals Quadruplex Folding of DNA Promotes the Condensation of Linker Histones via Liquid-Liquid Phase Separation

2020 ◽  
Author(s):  
Masahiro Mimura ◽  
Shunsuke Tomita ◽  
Yoichi Shinkai ◽  
Kentaro Shiraki ◽  
Ryoji Kurita
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Electrostatic Interactions ◽  
Droplet Formation ◽  
Liquid Phase Separation ◽  
Intrinsically Disordered ◽  
G Quadruplex ◽  
Dna Quadruplex ◽  
Liquid Liquid Phase Separation

<p>Liquid-liquid phase separation (LLPS) of proteins and DNA has recently emerged as a possible mechanism underlying the dynamic organization of chromatin. We herein report the role of DNA quadruplex folding in liquid droplet formation via LLPS induced by interactions between DNA and linker histone H1 (H1), a key regulator of chromatin organization. Fluidity measurements inside the droplets and binding assays using G-quadruplex-selective probes demonstrated that quadruplex DNA structures, such as the G-quadruplex and i-motif, promote droplet formation with H1 and decrease molecular motility within droplets. The dissolution of the droplets in the presence of additives indicated that in addition to electrostatic interactions between the DNA and the intrinsically disordered region of H1, π-π stacking between quadruplex DNAs could potentially drive droplet formation. Given that DNA quadruplex structures are well documented in heterochromatin regions, it is imperative to understand the role of DNA quadruplex folding in the context of intranuclear LLPS.<b></b></p>

scholarly journals Tandem RNA binding sites induce self-association of the stress granule marker protein TIA-1

2021 ◽  
Author(s):  
Fionna E Loughlin ◽  
Danella L West ◽  
Menachem J Gunzburg ◽  
Saboora Waris ◽  
Simon A Crawford ◽  
...  
Keyword(s):  
Phase Separation ◽  
Nucleic Acid ◽  
Liquid Phase ◽  
Binding Sites ◽  
Rna Binding ◽  
Liquid Phase Separation ◽  
Stress Granule ◽  
Granule Formation ◽  
Self Association ◽  
Liquid Liquid Phase Separation

Abstract TIA-1 is an RNA-binding protein that sequesters target RNA into stress granules under conditions of cellular stress. Promotion of stress granule formation by TIA-1 depends upon self-association of its prion-like domain that facilitates liquid-liquid phase separation and is thought to be enhanced via RNA binding. However, the mechanisms underlying the influence of RNA on TIA-1 self-association have not been previously demonstrated. Here we have investigated the self-associating properties of full-length TIA-1 in the presence of designed and native TIA-1 nucleic acid binding sites in vitro, monitoring phase separation, fibril formation and shape. We show that single stranded RNA and DNA induce liquid-liquid phase separation of TIA-1 in a multisite, sequence-specific manner and also efficiently promote formation of amyloid-like fibrils. Although RNA binding to a single site induces a small conformational change in TIA-1, this alone does not enhance phase separation of TIA-1. Tandem binding sites are required to enhance phase separation of TIA-1 and this is finely tuned by the protein:binding site stoichiometry rather than nucleic acid length. Native tandem TIA-1 binding sites within the 3′ UTR of p53 mRNA also efficiently enhance phase separation of TIA-1 and thus may potentially act as potent nucleation sites for stress granule assembly.

Quadruplex Folding of DNA Promotes the Condensation of Linker Histones via Liquid-Liquid Phase Separation

2020 ◽  
Author(s):  
Masahiro Mimura ◽  
Shunsuke Tomita ◽  
Yoichi Shinkai ◽  
Kentaro Shiraki ◽  
Ryoji Kurita
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Electrostatic Interactions ◽  
Droplet Formation ◽  
Liquid Phase Separation ◽  
Intrinsically Disordered ◽  
G Quadruplex ◽  
Dna Quadruplex ◽  
Liquid Liquid Phase Separation

<p>Liquid-liquid phase separation (LLPS) of proteins and DNA has recently emerged as a possible mechanism underlying the dynamic organization of chromatin. We herein report the role of DNA quadruplex folding in liquid droplet formation via LLPS induced by interactions between DNA and linker histone H1 (H1), a key regulator of chromatin organization. Fluidity measurements inside the droplets and binding assays using G-quadruplex-selective probes demonstrated that quadruplex DNA structures, such as the G-quadruplex and i-motif, promote droplet formation with H1 and decrease molecular motility within droplets. The dissolution of the droplets in the presence of additives indicated that in addition to electrostatic interactions between the DNA and the intrinsically disordered region of H1, π-π stacking between quadruplex DNAs could potentially drive droplet formation. Given that DNA quadruplex structures are well documented in heterochromatin regions, it is imperative to understand the role of DNA quadruplex folding in the context of intranuclear LLPS.<b></b></p>

scholarly journals G-quadruplex structures trigger RNA phase separation

2019 ◽  
Author(s):  
Yueying Zhang ◽  
Minglei Yang ◽  
Susan Duncan ◽  
Xiaofei Yang ◽  
Mahmoud A S Abdelhamid ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Molecular Basis ◽  
Short Root ◽  
Physiological Conditions ◽  
Cellular Processes ◽  
G Quadruplex ◽  
Liquid Liquid Phase Separation ◽  
In Cells

Abstract Liquid–liquid phase separation plays an important role in a variety of cellular processes, including the formation of membrane-less organelles, the cytoskeleton, signalling complexes, and many other biological supramolecular assemblies. Studies on the molecular basis of phase separation in cells have focused on protein-driven phase separation. In contrast, there is limited understanding on how RNA specifically contributes to phase separation. Here, we described a phase-separation-like phenomenon that SHORT ROOT (SHR) RNA undergoes in cells. We found that an RNA G-quadruplex (GQ) forms in SHR mRNA and is capable of triggering RNA phase separation under physiological conditions, suggesting that GQs might be responsible for the formation of the SHR phase-separation-like phenomenon in vivo. We also found the extent of GQ-triggered-phase-separation increases on exposure to conditions which promote GQ. Furthermore, GQs with more G-quartets and longer loops are more likely to form phase separation. Our studies provide the first evidence that RNA can adopt structural motifs to trigger and/or maintain the specificity of RNA-driven phase separation.

scholarly journals CRISPR-based DNA and RNA detection with liquid-liquid phase separation

2018 ◽  
Author(s):  
Willem Kasper Spoelstra ◽  
Jeroen M. Jacques ◽  
Franklin L. Nobrega ◽  
Anna C. Haagsma ◽  
Marileen Dogterom ◽  
...  
Keyword(s):  
Phase Separation ◽  
Nucleic Acid ◽  
Liquid Phase ◽  
Clinical Diagnostics ◽  
Liquid Phase Separation ◽  
Molecular Diagnostic ◽  
Target Sequence ◽  
Dna And Rna ◽  
Wide Range ◽  
Liquid Liquid Phase Separation

AbstractThe ability to detect specific nucleic acid sequences allows for a wide range of applications including identification of pathogens, clinical diagnostics, and genotyping. CRISPR-Cas proteins Cas12a and Cas13a are RNA-guided endonucleases that bind and cleave specific DNA and RNA sequences, respectively. After recognition of a target sequence both enzymes activate a unique, indiscriminate nucleic acid cleavage activity, which has been exploited for detection of sequence specific nucleotides using labelled reporter molecules. We here present a label-free detection approach that uses a readout based on solution turbidity caused by liquid-liquid phase separation (LLPS). Turbidity arises from coacervates of positively charged polyelectrolytes with long poly(dT) or poly(U) oligonucleotides. In the presence of a target sequence, long oligonucleotides are progressively shortened, changing the solution from turbid to transparent. We explain how oligonucleotide cleavage resolves LLPS by using a mathematical model which we validate with poly(dT) phase separation experiments. The deployment of LLPS complements CRISPR-based molecular diagnostic applications and facilitates easy and low-cost nucleotide sequence detection.

G-Quadruplex-Induced Liquid–Liquid Phase Separation in Biomimetic Protocells

2021 ◽  
Author(s):  
Xuejiao Liu ◽  
Yansong Xiong ◽  
Chunjuan Zhang ◽  
Rongji Lai ◽  
Hui Liu ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Liquid Phase Separation ◽  
G Quadruplex ◽  
Liquid Liquid Phase Separation

scholarly journals Quadruplex folding promotes the condensation of linker histones and DNAs via liquid-liquid phase separation

2021 ◽  
Author(s):  
Masahiro Mimura ◽  
Shunsuke Tomita ◽  
Yoichi Shinkai ◽  
Takuya Hosokai ◽  
Hiroyuki Kumeta ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Electrostatic Interactions ◽  
Droplet Formation ◽  
Liquid Phase Separation ◽  
Intrinsically Disordered ◽  
G Quadruplex ◽  
Dna Quadruplex ◽  
Liquid Liquid Phase Separation

Liquid-liquid phase separation (LLPS) of proteins and DNA has recently emerged as a possible mechanism underlying the dynamic organization of chromatin. We herein report the role of DNA quadruplex folding in liquid droplet formation via LLPS induced by interactions between DNA and linker histone H1 (H1), a key regulator of chromatin organization. <a>Fluidity measurements inside the droplets, binding assays using G-quadruplex-selective probes, and structural analyses based on circular dichroism demonstrated that quadruplex DNA structures, such as the G-quadruplex and i-motif, promote droplet formation with H1 and decrease molecular motility within droplets. </a><a></a><a></a><a>The dissolution of the droplets in the presence of additives and the LLPS of the DNA structural units indicated that in addition to electrostatic interactions between the DNA and the intrinsically disordered region of H1, π-π stacking between quadruplex DNAs could potentially drive droplet formation, unlike in the electrostatically driven LLPS of duplex DNA and H1. According to phase diagrams of anionic molecules with various conformations, the high LLPS ability associated with quadruplex folding arises from the formation of interfaces consisting of organized planes of guanine bases and the side surfaces with high charge density. </a>Given that DNA quadruplex structures are well documented in heterochromatin regions, it is imperative to understand the role of DNA quadruplex folding in the context of intranuclear LLPS.<br>

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