Phase Separation in Living Cells: Benefits and Risks

Preface
Contents
Part I: Physics and Chemistry
	Chapter 1: FUS Aggregation by Shear Stress on Pipetting and Its Suppression by Non-coding RNA
		1.1 FUS Protein and Long Non-coding RNA That Binds to FUS
			1.1.1 FUS Protein Implicated in Neurodegenerative Diseases
			1.1.2 Several Different States of FUS
			1.1.3 Long Non-coding RNA That Binds to FUS
		1.2 Large Conformational Change of FUS from a Compact to an Extended Structure upon Binding of Non-coding RNA
			1.2.1 FRET Analysis
			1.2.2 AFM Analysis
		1.3 FUS Aggregation by Shear Stress on Pipetting
			1.3.1 Fluorescence Spectroscopy Analysis
			1.3.2 Fluorescence Microscopy Analysis
			1.3.3 TEM Analysis
			1.3.4 The Mechanism of the FUS Aggregation on Pipetting
			1.3.5 The Relationship with Other Studies Including LLPS One
		1.4 Suppression of FUS Aggregation by Long Non-coding RNA in a Sequence-Dependent Manner
			1.4.1 Fluorescence Spectroscopy Analysis
			1.4.2 Fluorescence Microscopy Analysis
			1.4.3 The Mechanism of Suppression of FUS Aggregation by Long Non-coding RNA in a Sequence-Dependent Manner
		1.5 Conclusions
		References
	Chapter 2: Basics and Recent Advances in Computational and Theoretical Methods for Understanding the Liquid-Liquid Phase Separ...
		2.1 Introduction
		2.2 Theoretical Approaches
			2.2.1 Flory-Huggins Theory: Basic Theory for Understanding the LLPS Phenomena
			2.2.2 Theory of Viscoelastic Phase Separation
		2.3 Computational Approaches
			2.3.1 Molecular Dynamics Simulation
			2.3.2 Coarse-Grained Molecular Dynamics Simulation
			2.3.3 Informatics: Database, Machine Learning, and AI
		2.4 Summary
		References
	Chapter 3: Promotion of Liquid-Liquid Phase Separation by G-Quadruplex DNA and RNA
		3.1 Introduction
		3.2 G4s to Promote the Formation of Hydrogels
		3.3 G4s Binding Proteins to Promote the Formation of Hydrogels
		3.4 RNA Granules G4 Related to Disease
		3.5 Application of G4-Mediated Hydrogel
		3.6 Conclusion
		References
	Chapter 4: Chaperons Against Self-Association for Phase-Separating RNA-Binding Proteins
		4.1 Introduction
		4.2 Kapβ Family
		4.3 Kapβ Inhibits Self-Association of Substrate
		4.4 Mechanism for LLPS Inhibition
		4.5 Dysfunction of Kapβ2 and Disease
		4.6 HSP as Phase Regulator
		4.7 Perspective
		References
Part II: Molecular Biology
	Chapter 5: Positive and Negative Aspects of Protein Aggregation Induced by Phase Separation
		5.1 Introduction
		5.2 Intrinsically Disordered Protein Induces Phase Separation
		5.3 Harmful Aggregates Generated by the Phase Separation
		5.4 Regulation of Phase Separation and Resultant Aggregation
		5.5 Good Aggregates, Functional Amyloid for Memory Formation
		5.6 Future Prospects of Phase Separation and Aggregation
		References
	Chapter 6: Molecular Mechanisms Defining the Structural Basis for Self-Association of the FUS Low-Complexity Domain
		6.1 Introduction
		6.2 RNA-Binding Protein FUS
		6.3 Anatomy of the FUS RNA-Binding Proteins
		6.4 Hydrogel Formation by the FUS LCD
		6.5 Hydrogel Binding Assays
		6.6 Structures of Cross-β Polymers Formed by the FUS LCD
		6.7 Liquid-Liquid Phase Separation by the FUS LCD
		6.8 Maturation of Phase-Separated Liquid-like Droplets into Hydrogels
		6.9 A Two-Core System Controls Formation of Labile Cross-β Interactions of the FUS LCD
		6.10 Effects of Disease-Causing Mutations on Self-Association of FUS LC Domain
		6.11 Conclusion
		References
	Chapter 7: Winding and Tangling. An Initial Phase of Membrane-Less Organelle Formation
		7.1 Introduction to Cajal Body
			7.1.1 Nucleolar Accessory Body, Coiled Body, and Cajal Body
			7.1.2 Functions of Cajal Body
				7.1.2.1 snRNP Biogenesis
				7.1.2.2 Biogenesis of Other RNPs; snoRNP, scaRNP, and Telomerase RNP
				7.1.2.3 Processing of Histone mRNA
				7.1.2.4 Cajal Body as a Hub for Genome Organization
					Cajal Body Is Assembled on an Actively Transcribed snRNA Locus
					Cajal Body Assembly Organizes a Higher Order Structure of the Genome
			7.1.3 Physiological Relevance of Cajal Body
		7.2 Multiple Faces of Coilin and the Cajal Body
			7.2.1 The Close Relationship of Coilin with the Nucleolus
			7.2.2 Cajal Body and Gem
			7.2.3 Residual Bodies in the Absence of Coilin
			7.2.4 Phosphorylation Regulates Coilin Localization
			7.2.5 Roles of Arginine (di)Methylation
		7.3 Possible Mechanisms of Cajal Body Formation
			7.3.1 Nopp140, an Essential Component for Canonical Cajal Body Assembly
			7.3.2 Nopp140 Forms the Condensate
			7.3.3 The Role of Interaction between Coilin and Nopp140 for Cajal Body Assembly
				7.3.3.1 Characteristic Features of the IDRs of some Cajal Body Components
					Roles of IDR Charged Residues in Nuclear Body Assembly
				7.3.3.2 Characteristic Features of Cajal Body Component Proteins
				7.3.3.3 The Amplitude of the CW Motif Is Vital to Sort Proteins to the Specific Nuclear Body
			7.3.4 Roles of RNA in Phase Separation of Proteins with Charged Patches
			7.3.5 Effects of Post-Translational Modifications on the Property of IDR in Coilin
			7.3.6 A Dual-Pathway Model of Cajal Body Assembly
		7.4 Perspective
			7.4.1 RNPs in the Cajal Body Are a Mosaic Complex of Proteins with Different Motifs
			7.4.2 How scaRNPs Mediate Cajal Body Localization
		References
	Chapter 8: Formation and Function of Phase-Separated Nuclear Bodies Directed by Architectural Noncoding RNA
		8.1 Introduction
		8.2 General Molecular Features of arcRNAs
			8.2.1 Definition and Molecular Species of arcRNAs
			8.2.2 Advantages of Using RNAs as Architectural Scaffolds of MLOs
			8.2.3 Disadvantages of RNA-Mediated Phase Separation in Diseases
		8.3 Specific Molecular Features of NEAT1_2 arcRNA
			8.3.1 Overview of NEAT1_2 arcRNA and Paraspeckles
			8.3.2 The Paraspeckle Components
			8.3.3 Functional RNA Domains of NEAT1_2
			8.3.4 Paraspeckles Are Formed Through Micellization
			8.3.5 Significance of the Internal Core-Shell Structure of Paraspeckles
		8.4 Conclusion and Future Perspectives
		References
Part III: Biology
	Chapter 9: Force-Dependent Remodeling of Cell-to-Cell Adhesion Through the Regulation of Phase Separation
		9.1 Introduction
		9.2 Cell-to-Cell Adhesion Becomes Enhanced by Forces
		9.3 Droplets in the Epithelial Cells Are the Product of LLPS
		9.4 Molecular Nature of ZO-1 Protein
		9.5 The Mechano-Response of Embryonic Cells Is Conserved across Species
		9.6 Conclusions and Perspectives
		References
	Chapter 10: Regulation of Neuronal RNA Granule Dynamics Through Phase Separation in Memory Formation and Disease
		10.1 Introduction
		10.2 Constituents of Neuronal RNA Granules
		10.3 Transport of RNA Granules to Dendrites in Neurons
		10.4 Synaptic Input-Dependent Regulation of Local Translation
		10.5 Control of RNA Granule Formation and Dynamics through LLPS
		10.6 Long-Term Memory Formation Mediated by RNA Granule Components
		10.7 Neurodegenerative Diseases Associated with Aggregation of FUS and TDP-43 in RNA Granules
		10.8 Conclusion
		References
	Chapter 11: The Role of Liquid-Liquid Phase Separation in the Structure and Function of Nucleolus
		11.1 Overview
		11.2 Structure and Function of the Nucleolus
			11.2.1 Nucleolar Sub-Compartments
			11.2.2 Ribosome Biogenesis in the Nucleolus
				11.2.2.1 Production of Precursor rRNAs
				11.2.2.2 rRNA Maturation (Processing and Modification)
				11.2.2.3 Assembly of Precursor Ribosomal Subunits
			11.2.3 Multiple Liquid Phases
			11.2.4 Structural Dynamics of the Nucleolus during Cell Cycle
		11.3 Biochemical Properties of Nucleolar Proteins
			11.3.1 Nucleolar Proteins Are Disordered
			11.3.2 Charged Residues
			11.3.3 Role of rRNA in LLPS
		11.4 Nucleolus as a Potential Stress Sensor and Biomarker for Disease and Aging
			11.4.1 Heat Stress
			11.4.2 Viral Infection
			11.4.3 Cancer and Aging
		References
Part IV: Medicine
	Chapter 12: Phase Separation Orchestrates Cancer Signaling: Stress Granules as a Promising Target for Cancer Therapy
		12.1 Introduction
		12.2 Stress Granules at a Glance
		12.3 Phase Separation of MAPK Signaling Regulators
		12.4 Phase Separation of mTOR Signaling
			12.4.1 Phase Separation Inhibits mTORC1 Signaling
		12.5 Phase Separation of PKA Signaling
		12.6 SGs Regulate Cancer
			12.6.1 SG Assembly Mediated by Cancer Signaling Pathways
				12.6.1.1 TORC1 Signaling Regulates SG Assembly
				12.6.1.2 RAS Signaling Upregulates SGs
			12.6.2 SGs as a Therapeutic Target of Cancer Treatment
				12.6.2.1 Targeting Components of SGs and Cancer
				12.6.2.2 SG Nucleating Factors
				12.6.2.3 Targeting Post-translational Modifications Related to SG Assembly
			12.6.3 SGs and Cancer Chemotherapy
		12.7 Concluding Remarks and Future Perspectives
		References
	Chapter 13: The Multifaceted Regulation of TDP-43 Condensates at the Intersection of Physiology and Pathology: Implications fo...
		13.1 Introduction
		13.2 The Complexities of ALS and FTLD: The Interplay of Genetics, Molecular Processes
		13.3 Structural Organization of TDP-43 and Their Influence on Neurodegenerative Diseases
		13.4 The Multifaceted Nature of LLPS: Biomolecular Interactions and Condensate Formation
		13.5 Complex Regulatory Mechanisms of IDRs in RNA-Binding Proteins
		13.6 Understanding TDP-43 Dysfunction and Its Impact on TDP-43 Isoforms on Condensate Formation
		13.7 The Complex Relationship Between Physiological Condensates and Pathological Aggregates in Neurodegenerative Diseases
		13.8 Neuronal System Specificity in Neurodegenerative Diseases and the Multifaceted Impact of Condensates
		13.9 Challenges in Understanding Intracellular Condensates in Neurodegenerative Diseases
		13.10 Conclusion
		References
	Chapter 14: Functional Properties of Phase Separation and Intranuclear Complex of FUS in the Pathogenesis of ALS/FTLD
		14.1 Introduction
		14.2 Functional Properties of FUS
		14.3 Relation to the Disease
		14.4 LLPS
		14.5 Association with SFPQ
		14.6 ASO Therapeutics to Target the RBPs and the LLPS
		14.7 Future Direction
		References
	Chapter 15: Microglia Lipid Droplets in Physiology and Neurodegeneration
		15.1 Introduction to Lipid Droplets
		15.2 Conditions in Which Lipid Droplet Accumulating Microglia Are Found
		15.3 Transcriptional Regulation of Microglial Lipid Droplets
		15.4 Proteins Involved in Microglial Lipid Droplet Formation
		15.5 Lipid Droplet Degradation in Microglia
		15.6 Functional Impacts of Lipid Droplet Accumulating Microglia
		15.7 Conclusion
		References
	Chapter 16: Emerging Role of Phase Separation in COVID-19
		16.1 Introduction
		16.2 Structure of SARS-CoV-2
		16.3 Life Cycle of SARS-CoV-2
		16.4 SARS-CoV-2 Proliferation and Phase Separation
		16.5 Innate Immune Response to SARS-CoV-2 and Phase Separation
		16.6 Conclusion
		References
Index