Phase separation as a possible means of nuclear compartmentalization.

The nucleus is perhaps the most familiar organelle within eukaryotic cells, serving as a compartment to house the genetic material. The nuclear volume is subdivided into a variety of functional and dynamic nuclear bodies not separated from the nucleoplasm by membranes. It has been hypothesized that aqueous phase separation brought about by macromolecular crowding may be in part responsible for these intranuclear compartments. This chapter discusses macromolecular solution chemistry with regard to several common types of phase separation in polymer solutions as well as to recent evidence that suggests that cytoplasmic and nuclear bodies may exist as liquid phases. We then examine the functional significance of phase separation and how it may serve as a means of compartmentalizing various nuclear activities, and describe recent studies that have used simple model systems to generate coexisting aqueous phase compartments, concentrate molecules within them, and perform localized biochemical reactions.

[1]  C. G. D. Kruif,et al.  Polysaccharide protein interactions , 2001 .

[2]  Ying Wang,et al.  Phase transitions in human IgG solutions. , 2013, The Journal of chemical physics.

[3]  M. C. Stuart,et al.  Interfacial tension between a complex coacervate phase and its coexisting aqueous phase , 2010 .

[4]  Andrew C. Miklos,et al.  Protein crowding tunes protein stability. , 2011, Journal of the American Chemical Society.

[5]  P. Schurtenberger,et al.  Phase separation in binary eye lens protein mixtures , 2011 .

[6]  F. Tjerneld,et al.  Driving forces for phase separation and partitioning in aqueous two-phase systems. , 1998, Journal of chromatography. B, Biomedical sciences and applications.

[7]  P. Albertsson,et al.  Partition of Cell Particles and Macromolecules , 1986 .

[8]  J. Gall,et al.  Cajal bodies, nucleoli, and speckles in the Xenopus oocyte nucleus have a low-density, sponge-like structure. , 2004, Molecular biology of the cell.

[9]  D. Thirumalai,et al.  Molecular crowding enhances native state stability and refolding rates of globular proteins. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[10]  Christine D. Keating,et al.  Complete Budding and Asymmetric Division of Primitive Model Cells To Produce Daughter Vesicles with Different Interior and Membrane Compositions , 2011, Journal of the American Chemical Society.

[11]  J. Daban Physical constraints in the condensation of eukaryotic chromosomes. Local concentration of DNA versus linear packing ratio in higher order chromatin structures. , 2000, Biochemistry.

[12]  J Ovádi,et al.  Macromolecular compartmentation and channeling. , 2000, International review of cytology.

[13]  Paul S. Russo,et al.  Phase Transitions in the Assembly of MultiValent Signaling Proteins , 2016 .

[14]  A. Minton,et al.  Quantitative assessment of the relative contributions of steric repulsion and chemical interactions to macromolecular crowding. , 2013, Biopolymers.

[15]  T. H. Lilley,et al.  Theory of phase equilibria for multicomponent aqueous solutions: applications to aqueous polymer two-phase systems , 1993 .

[16]  Rosa Bernardi,et al.  Structure, dynamics and functions of promyelocytic leukaemia nuclear bodies , 2007, Nature Reviews Molecular Cell Biology.

[17]  T. Misteli,et al.  In vivo kinetics of Cajal body components , 2004, The Journal of cell biology.

[18]  V. Bloomfield DNA condensation by multivalent cations. , 1997, Biopolymers.

[19]  Donald E. Brooks,et al.  Partitioning in Aqueous Two-Phase Systems: Theory, Methods, Uses, and Applications to Biotechnology , 1986 .

[20]  S. Friberg A review of: “AQUEOUS TWO-PHASE PARTITIONING: Physical Chemistry and Bioanalytical Applications,” Boris Y. Zaslavsky, ed., Marcel Dekker, NY, 1995. , 1995 .

[21]  A. Estevez-Torres,et al.  DNA compaction: fundamentals and applications , 2011 .

[22]  Damien Hall,et al.  Macromolecular crowding: qualitative and semiquantitative successes, quantitative challenges. , 2003, Biochimica et biophysica acta.

[23]  Andrew S. LaCroix,et al.  Separation of preferential interaction and excluded volume effects on DNA duplex and hairpin stability , 2011, Proceedings of the National Academy of Sciences.

[24]  A. Piruska,et al.  Enhanced transcription rates in membrane-free protocells formed by coacervation of cell lysate , 2013, Proceedings of the National Academy of Sciences.

[25]  R. Hancock,et al.  Internal organisation of the nucleus: assembly of compartments by macromolecular crowding and the nuclear matrix model , 2004, Biology of the cell.

[26]  C. Filipe,et al.  In vivo formation of protein based aqueous microcompartments. , 2009, Journal of the American Chemical Society.

[27]  T Misteli,et al.  Protein dynamics: implications for nuclear architecture and gene expression. , 2001, Science.

[28]  T Misteli,et al.  Functional architecture in the cell nucleus. , 2001, The Biochemical journal.

[29]  Christine D. Keating,et al.  Biocatalyzed mineralization in an aqueous two-phase system: effect of background polymers and enzyme partitioning. , 2013, Journal of materials chemistry. B.

[30]  W. Loh,et al.  Calorimetric Investigation of the Formation of Aqueous Two-Phase Systems in Ternary Mixtures of Water, Poly(ethylene oxide) and Electrolytes (Or Dextran) , 2000 .

[31]  S. Mahalingam,et al.  Nuclear transport of Ras-associated tumor suppressor proteins: different transport receptor binding specificities for arginine-rich nuclear targeting signals. , 2007, Journal of molecular biology.

[32]  A. Hyman,et al.  Active liquid-like behavior of nucleoli determines their size and shape in Xenopus laevis oocytes , 2011, Proceedings of the National Academy of Sciences.

[33]  Marco Rito-Palomares,et al.  Practical experiences from the development of aqueous two‐phase processes for the recovery of high value biological products , 2008 .

[34]  F. Weinbreck,et al.  Complex coacervation of proteins and anionic polysaccharides , 2004 .

[35]  Bin Zhang,et al.  Biogenesis and function of nuclear bodies. , 2011, Trends in genetics : TIG.

[36]  Huan‐Xiang Zhou Influence of crowded cellular environments on protein folding, binding, and oligomerization: Biological consequences and potentials of atomistic modeling , 2013, FEBS letters.

[37]  Stephen Mann,et al.  Peptide-nucleotide microdroplets as a step towards a membrane-free protocell model. , 2011, Nature chemistry.

[38]  V. Tolstoguzov Compositions and phase diagrams for aqueous systems based on proteins and polysaccharides. , 2000, International review of cytology.

[39]  Huaying Zhao,et al.  The role of macromolecular crowding in the evolution of lens crystallins with high molecular refractive index , 2011, Physical biology.

[40]  C. Keating,et al.  Partitioning and assembly of metal particles and their bioconjugates in aqueous two-phase systems. , 2005, Langmuir : the ACS journal of surfaces and colloids.

[41]  C. Keating,et al.  Budding and asymmetric protein microcompartmentation in giant vesicles containing two aqueous phases. , 2008, Journal of the American Chemical Society.

[42]  P. Lichter,et al.  Macromolecular crowding and its potential impact on nuclear function. , 2008, Biochimica et biophysica acta.

[43]  C. Keating,et al.  Dynamic microcompartmentation in synthetic cells , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[44]  Clifford P. Brangwynne,et al.  Soft active aggregates: mechanics, dynamics and self-assembly of liquid-like intracellular protein bodies , 2011 .

[45]  M. Dundr,et al.  De Novo Formation of a Subnuclear Body , 2008, Science.

[46]  Dominique Durand,et al.  Particles trapped at the droplet interface in water-in-water emulsions. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[47]  D. Hernandez-Verdun Nucleolus: from structure to dynamics , 2005, Histochemistry and Cell Biology.

[48]  Taco Nicolai,et al.  Stabilization of water-in-water emulsions by addition of protein particles. , 2013, Langmuir : the ACS journal of surfaces and colloids.

[49]  Y. Sugita,et al.  Variable interactions between protein crowders and biomolecular solutes are important in understanding cellular crowding. , 2012, The journal of physical chemistry. B.

[50]  H. Bohidar,et al.  Systematic of alcohol-induced simple coacervation in aqueous gelatin solutions. , 2003, Biomacromolecules.

[51]  Tom Misteli,et al.  Biogenesis of nuclear bodies. , 2010, Cold Spring Harbor perspectives in biology.

[52]  A. Minton,et al.  The Influence of Macromolecular Crowding and Macromolecular Confinement on Biochemical Reactions in Physiological Media* , 2001, The Journal of Biological Chemistry.

[53]  Carol K. Hall,et al.  Interfacial tension of polyethyleneglycol-dextran-water systems: influence of temperature and polymer molecular weight , 1990 .

[54]  R. P. Aaronson,et al.  Organization in the cell nucleus: divalent cations modulate the distribution of condensed and diffuse chromatin , 1981, The Journal of cell biology.

[55]  R. Hancock,et al.  A role for macromolecular crowding effects in the assembly and function of compartments in the nucleus. , 2004, Journal of structural biology.

[56]  S. Sandler,et al.  Effect of ionic strength and ionic species on partitioning behavior of hydrophobic and hydrophilic polystyrene latex beads in aqueous two-phase polymer systems , 1996 .

[57]  C. Gespach,et al.  Nuclear bodies and compartments: functional roles and cellular signalling in health and disease. , 2004, Cellular signalling.

[58]  V. Juvekar,et al.  Quantification of thermodynamics of aqueous solutions of poly(ethylene glycols): Role of calorimetry , 2009 .

[59]  S. Takayama,et al.  Patchy surfaces stabilize dextran-polyethylene glycol aqueous two-phase system liquid patterns. , 2013, Langmuir : the ACS journal of surfaces and colloids.

[60]  S. Turgeon,et al.  Protein-polysaccharide interactions: phase-ordering kinetics, thermodynamic and structural aspects , 2003 .

[61]  Allen P. Minton,et al.  The effect of volume occupancy upon the thermodynamic activity of proteins: some biochemical consequences , 2004, Molecular and Cellular Biochemistry.

[62]  T. H. Lilley,et al.  A new excluded volume theory and its application to the coexistence curves of aqueous polymer two-phase systems , 1993 .

[63]  F. Weinbreck,et al.  Rheological properties of whey protein/gum arabic coacervates , 2004 .

[64]  K. Luby‐Phelps The physical chemistry of cytoplasm and its influence on cell function: an update , 2013, Molecular biology of the cell.

[65]  S. Zimmerman,et al.  Shape and compaction of Escherichia coli nucleoids. , 2006, Journal of structural biology.

[66]  T. Misteli,et al.  High mobility of proteins in the mammalian cell nucleus , 2000, Nature.

[67]  G R Welch,et al.  Transient-time analysis of substrate-channelling in interacting enzyme systems. , 1989, The Biochemical journal.

[68]  Jimin Pei,et al.  Cell-free Formation of RNA Granules: Bound RNAs Identify Features and Components of Cellular Assemblies , 2012, Cell.

[69]  David S. Williams,et al.  Stabilization and enhanced reactivity of actinorhodin polyketide synthase minimal complex in polymer-nucleotide coacervate droplets. , 2012, Chemical communications.

[70]  M. Tirrell,et al.  Phase behaviour and complex coacervation of aqueous polypeptide solutions , 2012 .

[71]  Jun Soo Kim,et al.  Crowding-induced phase separation of Lennard-Jones particles: implications to nuclear structures in a biological cell. , 2012, The journal of physical chemistry. B.

[72]  R. Lipowsky,et al.  Lipid membranes in contact with aqueous phases of polymer solutions , 2012 .

[73]  D. Brooks,et al.  Phase separation in cytoplasm, due to macromolecular crowding, is the basis for microcompartmentation , 1995, FEBS letters.

[74]  Mohona Sarkar,et al.  Macromolecular crowding and protein stability. , 2012, Journal of the American Chemical Society.

[75]  F. Szoka,et al.  The influence of polymer structure on the interactions of cationic polymers with DNA and morphology of the resulting complexes , 1997, Gene Therapy.

[76]  J. Clegg,et al.  From protoplasmic theory to cellular systems biology: a 150-year reflection. , 2010, American journal of physiology. Cell physiology.

[77]  M. Tirrell,et al.  Polyelectrolyte Molecular Weight and Salt Effects on the Phase Behavior and Coacervation of Aqueous Solutions of Poly(acrylic acid) Sodium Salt and Poly(allylamine) Hydrochloride , 2013 .

[78]  J. Arons,et al.  Thermodynamics of phase separation in aqueous solutions of polymers , 1993 .

[79]  C. Pikaard,et al.  Subnuclear partitioning of rRNA genes between the nucleolus and nucleoplasm reflects alternative epiallelic states. , 2013, Genes & development.

[80]  Adrian H Elcock,et al.  Models of macromolecular crowding effects and the need for quantitative comparisons with experiment. , 2010, Current opinion in structural biology.

[81]  Anthony K. L. Leung,et al.  Nucleolar proteome dynamics , 2005, Nature.

[82]  A. Minton,et al.  Analysis of non-ideal behavior in concentrated hemoglobin solutions. , 1977, Journal of molecular biology.

[83]  Rajni Hatti-Kaul,et al.  Aqueous two-phase systems : methods and protocols , 2000 .

[84]  David S. Williams,et al.  Polymer/nucleotide droplets as bio-inspired functional micro-compartments , 2012 .

[85]  Maxwell Z. Wilson,et al.  Beyond the cytoskeleton: mesoscale assemblies and their function in spatial organization. , 2013, Current opinion in microbiology.

[86]  Reinhard Lipowsky,et al.  Concentration dependence of the interfacial tension for aqueous two-phase polymer solutions of dextran and polyethylene glycol. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[87]  J. Pelta,et al.  DNA Aggregation Induced by Polyamines and Cobalthexamine (*) , 1996, The Journal of Biological Chemistry.

[88]  K. Praveen,et al.  Nuclear bodies: random aggregates of sticky proteins or crucibles of macromolecular assembly? , 2009, Developmental cell.

[89]  Christine D. Keating,et al.  Aqueous Phase Separation as a Possible Route to Compartmentalization of Biological Molecules , 2012, Accounts of chemical research.

[90]  M. Tirrell,et al.  Interfacial energy of polypeptide complex coacervates measured via capillary adhesion. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[91]  H. Sugiyama,et al.  Mapping a nucleolar targeting sequence of an RNA binding nucleolar protein, Nop25. , 2006, Experimental cell research.

[92]  C. Keating,et al.  Microcompartmentation in artificial cells: pH-induced conformational changes alter protein localization. , 2010, Langmuir : the ACS journal of surfaces and colloids.

[93]  G. Benedek,et al.  Phase separation in aqueous solutions of lens gamma-crystallins: special role of gamma s. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[94]  ENZYMATIC SYNTHESIS OF POLYADENYLIC ACID IN COACERVATE DROPS , 1963 .

[95]  E. B. Wilson THE STRUCTURE OF PROTOPLASM. , 1899, Science.

[96]  P. Vekilov Phase diagrams and kinetics of phase transitions in protein solutions , 2012, Journal of physics. Condensed matter : an Institute of Physics journal.

[97]  M. Dundr,et al.  Nuclear bodies: multifunctional companions of the genome. , 2012, Current opinion in cell biology.

[98]  Huan‐Xiang Zhou,et al.  Macromolecular crowding and confinement: biochemical, biophysical, and potential physiological consequences. , 2008, Annual review of biophysics.

[99]  Philip C Bevilacqua,et al.  RNA catalysis through compartmentalization. , 2012, Nature chemistry.

[100]  Reinhard Lipowsky,et al.  Membrane nanotubes induced by aqueous phase separation and stabilized by spontaneous curvature , 2011, Proceedings of the National Academy of Sciences.

[101]  Diane J. Burgess,et al.  Microelectrophoretic studies of gelatin and acacia for the prediction of complex coacervation , 1984 .

[102]  R. Ellis Macromolecular crowding : obvious but underappreciated , 2022 .

[103]  E. Kaler,et al.  Protein phase behavior in aqueous solutions: crystallization, liquid-liquid phase separation, gels, and aggregates. , 2008, Biophysical journal.

[104]  B. Zaslavsky,et al.  Aqueous Two-Phase Partitioning: Physical Chemistry and Bioanalytical Applications , 1994 .

[105]  Jason R. Swedlow,et al.  In Vivo Analysis of Cajal Body Movement, Separation, and Joining in Live Human Cells , 2000, The Journal of cell biology.

[106]  E. Dickinson,et al.  Interfacial structuring in a phase-separating mixed biopolymer solution containing colloidal particles. , 2009, Langmuir : the ACS journal of surfaces and colloids.

[107]  Jimin Pei,et al.  Cell-free Formation of RNA Granules: Low Complexity Sequence Domains Form Dynamic Fibers within Hydrogels , 2012, Cell.

[108]  Pedro Gutemberg de Alcântara,et al.  A Hydration Shell‐Based Thermodynamic Model for Aqueous Two‐Phase Systems , 2008 .

[109]  Marco Rito-Palomares,et al.  Aqueous two-phase affinity partitioning systems: current applications and trends. , 2012, Journal of chromatography. A.

[110]  Judit Ovádi,et al.  On the origin of intracellular compartmentation and organized metabolic systems , 2004, Molecular and Cellular Biochemistry.

[111]  M. Nomura,et al.  Mutational Analysis of the Structure and Localization of the Nucleolus in the Yeast Saccharomyces cerevisiae , 1998, The Journal of cell biology.

[112]  G. Setterfield,et al.  Effects of low salt concentration on structural organization and template activity of chromatin in chicken erythrocyte nuclei. , 1971, Experimental cell research.

[113]  Christine D. Keating,et al.  Positioning lipid membrane domains in giant vesicles by micro-organization of aqueous cytoplasm mimic. , 2008, Journal of the American Chemical Society.

[114]  C. Keating,et al.  Aqueous phase separation in giant vesicles. , 2002, Journal of the American Chemical Society.

[115]  A. Hyman,et al.  Germline P Granules Are Liquid Droplets That Localize by Controlled Dissolution/Condensation , 2009, Science.