scholarly journals DNA repair by Rad52 liquid droplets

2019 ◽  
Author(s):  
Roxanne Oshidari ◽  
Richard Huang ◽  
Maryam Medghalchi ◽  
Elizabeth Y.W. Tse ◽  
Nasser Ashgriz ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Dna Repair ◽  
Phase Separation ◽  
Liquid Phase ◽  
Nuclear Periphery ◽  
Liquid Phase Separation ◽  
Liquid Droplets ◽  
Cellular Processes ◽  
Dna Repair Proteins

Cellular processes are influenced by liquid phase separation, but its role in DNA repair is unclear. Here, we show that in Saccharomyces cerevisiae, Rad52 DNA repair proteins at different DNA damage sites assemble liquid droplets that fuse into a repair centre droplet. This larger droplet concentrates tubulin and projects short aster-like microtubule filaments, which tether the droplet to longer microtubule filaments mediating the mobilization of damaged DNA to the nuclear periphery for repair.

scholarly journals FUS-dependent liquid–liquid phase separation is important for DNA repair initiation

2021 ◽  
Vol 220 (5) ◽  
Author(s):  
Brunno R. Levone ◽  
Silvia C. Lenzken ◽  
Marco Antonaci ◽  
Andreas Maiser ◽  
Alexander Rapp ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Phase Separation ◽  
Liquid Phase ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Liquid Phase Separation ◽  
Structured Illumination Microscopy ◽  
Liquid Liquid Phase Separation

RNA-binding proteins (RBPs) are emerging as important effectors of the cellular DNA damage response (DDR). The RBP FUS is implicated in RNA metabolism and DNA repair, and it undergoes reversible liquid–liquid phase separation (LLPS) in vitro. Here, we demonstrate that FUS-dependent LLPS is necessary for the initiation of the DDR. Using laser microirradiation in FUS-knockout cells, we show that FUS is required for the recruitment to DNA damage sites of the DDR factors KU80, NBS1, and 53BP1 and of SFPQ, another RBP implicated in the DDR. The relocation of KU80, NBS1, and SFPQ is similarly impaired by LLPS inhibitors, or LLPS-deficient FUS variants. We also show that LLPS is necessary for efficient γH2AX foci formation. Finally, using superresolution structured illumination microscopy, we demonstrate that the absence of FUS impairs the proper arrangement of γH2AX nanofoci into higher-order clusters. These findings demonstrate the early requirement for FUS-dependent LLPS in the activation of the DDR and the proper assembly of DSB repair complexes.

scholarly journals Perturbation of liquid droplets of P-granule protein LAF-1 by the antimicrobial peptide LL-III

2020 ◽  
Vol 56 (78) ◽  
pp. 11577-11580
Author(s):  
Rosario Oliva ◽  
Sanjib K. Mukherjee ◽  
Zamira Fetahaj ◽  
Simone Möbitz ◽  
Roland Winter
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Antimicrobial Peptides ◽  
Antimicrobial Peptide ◽  
Droplet Formation ◽  
Liquid Phase Separation ◽  
Liquid Droplets ◽  
Cellular Organization ◽  
P Protein ◽  
Liquid Liquid Phase Separation

Protein/RNA droplet formation by liquid–liquid phase separation has emerged as a key mechanism for cellular organization. We show that binding of antimicrobial peptides such as LL-III can lead to loss of droplet function.

scholarly journals Transcription Regulators and Membraneless Organelles Challenges to Investigate Them

2021 ◽  
Vol 22 (23) ◽  
pp. 12758
Author(s):  
Katarzyna Sołtys ◽  
Andrzej Ożyhar
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Liquid Phase Separation ◽  
Cellular Processes ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Experimental Systems ◽  
Transcription Regulators ◽  
Crucial Information ◽  
Liquid Liquid Phase Separation

Eukaryotic cells are composed of different bio-macromolecules that are divided into compartments called organelles providing optimal microenvironments for many cellular processes. A specific type of organelles is membraneless organelles. They are formed via a process called liquid–liquid phase separation that is driven by weak multivalent interactions between particular bio-macromolecules. In this review, we gather crucial information regarding different classes of transcription regulators with the propensity to undergo liquid–liquid phase separation and stress the role of intrinsically disordered regions in this phenomenon. We also discuss recently developed experimental systems for studying formation and properties of membraneless organelles.

scholarly journals Dynamic Formation of Liquid Droplets Triggered by Sequential Enzymatic Reactions

2020 ◽  
Author(s):  
Tomoto Ura ◽  
Shunsuke Tomita ◽  
Kentaro Shiraki
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Enzyme Reaction ◽  
Model System ◽  
Liquid Phase Separation ◽  
Enzymatic Reactions ◽  
Liquid Droplets ◽  
Dynamic Formation ◽  
Liquid Liquid Phase Separation

<p>A model system was developed that dynamically generates two different liquid droplets via liquid–liquid phase separation coupled with a sequential glycolytic reaction. The sequential two-enzyme reaction triggers the formation/dissolution of the liquid droplets. The droplets, in turn, compartmentalize each enzymatic step and generate feedback to accelerate the overall reaction.</p>

scholarly journals FUS-dependent liquid-liquid phase separation is an early event in double-strand break repair

2019 ◽  
Author(s):  
Brunno R. Levone ◽  
Silvia C. Lenzken ◽  
Marco Antonaci ◽  
Andreas Maiser ◽  
Alexander Rapp ◽  
...  
Keyword(s):  
Dna Damage ◽  
Phase Separation ◽  
Liquid Phase ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Liquid Phase Separation ◽  
Early Event ◽  
Structured Illumination Microscopy ◽  
Liquid Liquid Phase Separation

AbstractRNA-binding proteins (RBPs) are emerging as important effectors of the cellular DNA damage response (DDR). The RBP FUS is implicated in RNA metabolism and DNA repair, and it undergoes reversible liquid-liquid phase separation (LLPS) in vitro. Here, we demonstrate that FUS-dependent LLPS is necessary for the initiation of the DDR. Using laser microirradiation in FUS-knockout cells, we show that FUS is required for the recruitment to DNA damage sites of the DDR factors KU80, NBS1, 53BP1, and of SFPQ, another RBP implicated in the DDR. The relocation of KU80, NBS1, and SFPQ is similarly impaired by LLPS inhibitors, or LLPS-deficient FUS variants. We also show that LLPS is necessary for efficient γH2AX foci formation. Finally, using super-resolution structured illumination microscopy, we demonstrate that the absence of FUS impairs the proper arrangement of γH2AX nano-foci into higher-order clusters. These findings demonstrate the early requirement for FUS-dependent LLPS in the activation of the DDR and the proper assembly of DSBs repair complexes.

Dynamic behavior of liquid droplets with enzyme compartmentalization triggered by sequential glycolytic enzyme reactions

2021 ◽  
Author(s):  
Tomoto Ura ◽  
Shunsuke Tomita ◽  
Kentaro Shiraki
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Dynamic Behavior ◽  
Droplet Formation ◽  
Glycolytic Enzyme ◽  
Liquid Phase Separation ◽  
Biological Processes ◽  
Liquid Droplets ◽  
Enzyme Reactions ◽  
Liquid Liquid Phase Separation

Dynamic droplet formation via liquid-liquid phase separation (LLPS) is believed to be involved in the regulation of various biological processes. Here, a model LLPS system coupled with a sequential glycolytic...

A Short Peptide Synthon for Liquid-Liquid Phase Separation

2020 ◽  
Author(s):  
Manzar Abbas ◽  
Wojciech P. Lipiński ◽  
Karina K. Nakashima ◽  
Wilhelm T.S. Huck ◽  
Evan Spruijt
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Living Cells ◽  
Liquid Phase Separation ◽  
Disordered Proteins ◽  
Liquid Droplets ◽  
Short Peptide ◽  
Peptide Analogues ◽  
Single Peptide ◽  
Liquid Liquid Phase Separation

Liquid-liquid phase separation of disordered proteins has emerged as a ubiquitous route to membraneless compartments in living cells, and similar coacervates may have played a role when the first cells formed. However, existing coacervates are typically made of multiple macromolecular components, and designing short peptide analogues capable of self-coacervation has proven difficult. Here, we present a short peptide synthon for phase separation, made of only two dipeptide stickers linked via a flexible, hydrophilic spacer. These small-molecule compounds self-coacervate into micrometre-sized liquid droplets at sub-mM concentrations, which retain up to 75 weight-% water. The design is general and we derive guidelines for the required sticker hydrophobicity and spacer polarity. To illustrate their potential as protocells, we create a disulphide-linked derivative that undergoes reversible compartmentalisation controlled by redox chemistry. The resulting coacervates sequester and melt nucleic acids, and act as microreactors that catalyse two different anabolic reactions yielding molecules of increasing complexity. This provides a stepping stone for new protocells made of single peptide species.<br>

A Short Peptide Synthon for Liquid-Liquid Phase Separation

2020 ◽  
Author(s):  
Manzar Abbas ◽  
Wojciech P. Lipiński ◽  
Karina K. Nakashima ◽  
Wilhelm T.S. Huck ◽  
Evan Spruijt
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Living Cells ◽  
Liquid Phase Separation ◽  
Disordered Proteins ◽  
Liquid Droplets ◽  
Short Peptide ◽  
Peptide Analogues ◽  
Single Peptide ◽  
Liquid Liquid Phase Separation

Liquid-liquid phase separation of disordered proteins has emerged as a ubiquitous route to membraneless compartments in living cells, and similar coacervates may have played a role when the first cells formed. However, existing coacervates are typically made of multiple macromolecular components, and designing short peptide analogues capable of self-coacervation has proven difficult. Here, we present a short peptide synthon for phase separation, made of only two dipeptide stickers linked via a flexible, hydrophilic spacer. These small-molecule compounds self-coacervate into micrometre-sized liquid droplets at sub-mM concentrations, which retain up to 75 weight-% water. The design is general and we derive guidelines for the required sticker hydrophobicity and spacer polarity. To illustrate their potential as protocells, we create a disulphide-linked derivative that undergoes reversible compartmentalisation controlled by redox chemistry. The resulting coacervates sequester and melt nucleic acids, and act as microreactors that catalyse two different anabolic reactions yielding molecules of increasing complexity. This provides a stepping stone for new protocells made of single peptide species.<br>

scholarly journals Small molecules as potent biphasic modulators of protein liquid-liquid phase separation

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
W. Michael Babinchak ◽  
Benjamin K. Dumm ◽  
Sarah Venus ◽  
Solomiia Boyko ◽  
Andrea A. Putnam ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Small Molecules ◽  
Rational Design ◽  
De Novo ◽  
Liquid Phase Separation ◽  
Disulfonic Acid ◽  
Liquid Droplets ◽  
Small Molecule Modulators ◽  
Liquid Liquid Phase Separation

Abstract Liquid-liquid phase separation (LLPS) of proteins that leads to formation of membrane-less organelles is critical to many biochemical processes in the cell. However, dysregulated LLPS can also facilitate aberrant phase transitions and lead to protein aggregation and disease. Accordingly, there is great interest in identifying small molecules that modulate LLPS. Here, we demonstrate that 4,4’-dianilino-1,1’-binaphthyl-5,5’-disulfonic acid (bis-ANS) and similar compounds are potent biphasic modulators of protein LLPS. Depending on context, bis-ANS can both induce LLPS de novo as well as prevent formation of homotypic liquid droplets. Our study also reveals the mechanisms by which bis-ANS and related compounds modulate LLPS and identify key chemical features of small molecules required for this activity. These findings may provide a foundation for the rational design of small molecule modulators of LLPS with therapeutic value.

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