DOCSLIB.ORG

Evaluating Phase Separation in Live Cells: Diagnosis, Caveats, and Functional Consequences

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Evaluating Phase Separation in Live Cells: Diagnosis, Caveats, and Functional Consequences Downloaded from genesdev.cshlp.org on October 1, 2021 - Published by Cold Spring Harbor Laboratory Press PERSPECTIVE Evaluating phase separation in live cells: diagnosis, caveats, and functional consequences David T. McSwiggen,1,2 Mustafa Mir,1 Xavier Darzacq,1,2 and Robert Tjian1,3 1Department of Molecular and Cell Biology, University of California Berkeley, California 94720, USA; 2California Institute of Regenerative Medicine Center of Excellence, University of California Berkeley, California 94720, USA; 3Howard Hughes Medical Institute, University of California Berkeley, California 94720, USA The idea that liquid–liquid phase separation (LLPS) may sure that these assemblies occur, and furthermore, that be a general mechanism by which molecules in the com- they do so on timescales relevant to their biological func- plex cellular milieu may self-organize has generated much tion. Prototypical examples of cellular organization are excitement and fervor in the cell biology community. the membrane-bound organelles, but it has long been ap- While this concept is not new, its rise to preeminence preciated that many compartments exist in the cell with- has resulted in renewed interest in the mechanisms that out an enclosing membrane (Montgomery 1898; Wilson shape and drive diverse cellular self-assembly processes 1899). from gene expression to cell division to stress responses. In the past decade, a fresh perspective on membraneless In vitro biochemical data have been instrumental in deriv- compartments—now often referred to as biomolecular ing some of the fundamental principles and molecular condensates (Banani et al. 2017)—has led to a resurgence grammar by which biological molecules may phase sepa- in the idea that a majority of these compartments may ex- rate, and the molecular basis of these interactions. Defin- ist as separate liquid phases (Courchaine et al. 2016). itive evidence is lacking as to whether the same principles There has been a renaissance in understanding how liq- apply in the physiological environment inside living cells. uid–liquid phase separation (LLPS) might function in com- In this Perspective, we analyze the evidence supporting partment formation and maintenance (Hyman et al. 2014; phase separation in vivo across multiple cellular process- Banani et al. 2017). Perhaps the most often cited example es. We find that the evidence for in vivo LLPS is often phe- is the nucleolus, where a convergence of studies examin- nomenological and inadequate to discriminate between ing its liquid-like behavior (Brangwynne et al. 2011), sup- phase separation and other possible mechanisms. More- ported with biochemical (Feric et al. 2016; Mitrea et al. over, the causal relationship and functional consequences 2016) and in vivo experiments (Berry et al. 2015; Weber of LLPS in vivo are even more elusive. We underscore the and Brangwynne 2015), collectively support a model importance of performing quantitative measurements on where the nucleolus behaves as a separate liquid phase proteins in their endogenous state and physiological abun- within the nucleus. Inspired by these and other early dance, as well as make recommendations for experiments examples of compartments with liquid-like properties that may yield more conclusive results. (Brangwynne et al. 2009), there has been a surge of publi- cations revisiting the formation of well-known cellular compartments through the lens of LLPS. Far from being Fundamentally, a cell is a collection of molecules com- the peculiarity it once was, phase separation now has partmentalized in a manner to modulate biochemical re- become, for many, the default explanation to rationalize actions that support diverse cellular activities. The the remarkable way in which a cell achieves various types challenges faced by a cell in managing these biochemical of compartmentalization, prompting significant debate processes scales with organismal complexity. In eukary- within the scientific community (Mir et al. 2019). otes, where some cellular tasks can require the coordinat- Much of the debate around LLPS condensates arises ed activity of tens to hundreds of individual molecular because it is unclear how strong the evidence for in vivo components, elaborate mechanisms have evolved to en- LLPS is, particularly when LLPS is invoked so broadly across many cellular contexts. The current focus on LLPS as a mechanism may come at the expense of under- [Keywords: fluorescence recovery after photobleaching; condensate; standing alternative mechanisms by which a high local liquid–liquid phase separation; phase separation] Corresponding authors: [email protected], [email protected] Article published online ahead of print. Article and publication date are online at http://www.genesdev.org/cgi/doi/10.1101/gad.331520.119. Free- © 2019 McSwiggen et al. This article, published in Genes & Develop- ly available online through the Genes & Development Open Access ment, is available under a Creative Commons License (Attribution 4.0 In- option. ternational), as described at http://creativecommons.org/licenses/by/4.0/. GENES & DEVELOPMENT 33:1619–1634 Published by Cold Spring Harbor Laboratory Press; ISSN 0890-9369/19; www.genesdev.org 1619 Downloaded from genesdev.cshlp.org on October 1, 2021 - Published by Cold Spring Harbor Laboratory Press McSwiggen et al. concentration of factors can be achieved in the absence of volumes occupied by the two phases (Fig. 1). A simplistic a membrane. For example, while nucleoli exhibit many example of this is the nucleation and growth of water properties consistent with LLPS, the formation of nucleoli droplets on a cold glass. Accumulating evidence suggests and many other nuclear bodies have previously been ex- the potential for LLPS to occur widely with biological plained by alternative mechanisms (Mao et al. 2011a,b; macromolecules as well, and it has been shown that cer- Shevtsov and Dundr 2011). In a recent study we found tain classes of proteins—as well as RNA and other biolog- that Herpes Simplex Virus replication compartments ical polymers—readily undergo LLPS in vitro (Jain and derive their ability to concentrate cellular factors through Vale 2017; Wang et al. 2018). transient nonspecific binding to the viral DNA in a man- The topic of phase separation in biology has been exten- ner distinct from liquid–liquid phase separation (McSwig- sively reviewed elsewhere, and the reader is encouraged to gen et al. 2019). Despite this mechanistic distinction, refer to these reviews for a more thorough explanation of these replication compartments display many of the hall- the forces that drive liquid–liquid demixing (Hyman marks that are often deemed sufficient to claim that such et al. 2014; Brangwynne et al. 2015; Banani et al. 2017; a compartment is formed via LLPS (McSwiggen et al. Boeynaems et al. 2018). Much of what we know now has 2019). foundations in early works on polymer physics (Overbeek Our data on replication compartments, as well as other and Voorn 1957) and has been advanced by efforts to recent studies from our group (Mir et al. 2017, 2018; improve crystallographic methods for which phase separa- Chong et al. 2018) demonstrate that there are multiple tion was used as a means of increasing a protein’s concen- routes to establish regions with high local concentrations tration without it crashing out of solution (Lomakin et al. of specific factors inside the cell. These studies prompted 1996; Asherie 2004; Vekilov 2010). Other types of phase us to critically reexamine the current evidence for LLPS in transition have also been proposed to occur in cells. vivo. The appeal for invoking phase separation is under- For example, it has been proposed that some proteins standable, as it presents a way to rationalize—and at least may transition into gel-like structures (Kato et al. 2012; superficially explain—certain behaviors of cellular com- Kwon et al. 2013) or liquid-crystalline structures (Rog partments. However, in light of various recent studies et al. 2017), again drawing models from lessons learned and upon further analysis, we find that the evidence for in polymer physics and materials science for inspiration. LLPS occurring in the cell is often far from conclusive. Physical models exist to explain liquid demixing (Loma- This is not to imply that LLPS cannot function in biolog- kin et al. 1996; Velasco et al. 1998), and for purified com- ical contexts, but rather to highlight how the tests com- ponents like proteins or nucleic acids, there exist rigorous monly used in probing LLPS are insufficient to rule out standards by which one may determine whether a given other mechanistic interpretations. system is undergoing liquid–liquid demixing. Modulating In this Perspective, we summarize the evidence used to the concentration of a polymer, the ionic strength of the diagnose liquid–liquid phase separation in vivo. Recently, buffer, the temperature of the system, and intra- or inter- others have similarly urged caution in overinterpreting in polymer interactions can all quantifiably change the pro- vivo experiments to test LLPS (Alberti et al. 2019), but the pensity of the polymer to demix (Lomakin et al. 1996; issues in this field run deeper than the authors discuss. Velasco et al. 1998; Vekilov 2010; Brangwynne 2013). Fol- This Perspective is, to our knowledge, the first to system- lowing this model, beautiful in vitro experiments have atically and holistically consider the evidence presented been performed demonstrating the ability of LLPS sys- by this emerging field. We first provide a summary of tems to exhibit exclusivity (Nott et al. 2015; Banani the state of evidence for LLPS condensates across multiple et al. 2016; Feric et al. 2016); to form and dissolve on the contexts, and address important considerations for this basis of post-translational modifications (Li et al. 2012; evidence. Second, we address the evidence for the func- Lu et al. 2018) and to exhibit changes in viscosity and to tional consequences of LLPS in the underlying biological “ripen” or harden over time (Patel et al. 2015; Wegmann processes being studied. Finally, we urge the application et al.
Load more

Recommended publications
  • Biogenesis of Nuclear Bodies
    Downloaded from http://cshperspectives.cshlp.org/ on September 30, 2021 - Published by Cold Spring Harbor Laboratory Press Biogenesis of Nuclear Bodies Miroslav Dundr1 and Tom Misteli2 1Department of Cell Biology, Rosalind Franklin University of Medicine and Science, North Chicago, Ilinois 60064 2National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892 Correspondence: [email protected]; [email protected] The nucleus is unique amongst cellular organelles in that it contains a myriad of discrete suborganelles. These nuclear bodies are morphologically and molecularly distinct entities, and they host specific nuclear processes. Although the mode of biogenesis appears to differ widely between individual nuclear bodies, several common design principles are emerging, particularly, the ability of nuclear bodies to form de novo, a role of RNA as a struc- tural element and self-organization as a mode of formation. The controlled biogenesis of nuclear bodies is essential for faithful maintenance of nuclear architecture during the cell cycle and is an important part of cellular responses to intra- and extracellular events. he mammalian cell nucleus contains a mul- seems to act indirectly by regulating the local Ttitude of discrete suborganelles, referred to concentration of its components in the nucleo- as nuclear bodies or nuclear compartments plasm. (reviewed in Dundr and Misteli 2001; Spector In many ways, nuclear bodies are similar 2001; Lamond and Spector 2003; Handwerger to conventional cellular organelles in the cy- and Gall 2006; Zhao et al. 2009). These bodies toplasm. Like cytoplasmic organelles, they con- are an essential part of the nuclear landscape tain a specific set of resident proteins, which as they compartmentalize the nuclear space defines each structure molecularly.
    [Show full text]
  • Nucleus Size and DNA Accessibility Are Linked to the Regulation Of
    Grosch et al. BMC Biology (2020) 18:42 https://doi.org/10.1186/s12915-020-00770-y RESEARCH ARTICLE Open Access Nucleus size and DNA accessibility are linked to the regulation of paraspeckle formation in cellular differentiation Markus Grosch1, Sebastian Ittermann1, Ejona Rusha2, Tobias Greisle1, Chaido Ori1,3, Dong-Jiunn Jeffery Truong4, Adam C. O’Neill1, Anna Pertek2, Gil Gregor Westmeyer4 and Micha Drukker1,2* Abstract Background: Many long noncoding RNAs (lncRNAs) have been implicated in general and cell type-specific molecular regulation. Here, we asked what underlies the fundamental basis for the seemingly random appearance of nuclear lncRNA condensates in cells, and we sought compounds that can promote the disintegration of lncRNA condensates in vivo. Results: As a basis for comparing lncRNAs and cellular properties among different cell types, we screened lncRNAs in human pluripotent stem cells (hPSCs) that were differentiated to an atlas of cell lineages. We found that paraspeckles, which form by aggregation of the lncRNA NEAT1, are scaled by the size of the nucleus, and that small DNA-binding molecules promote the disintegration of paraspeckles and other lncRNA condensates. Furthermore, we found that paraspeckles regulate the differentiation of hPSCs. Conclusions: Positive correlation between the size of the nucleus and the number of paraspeckles exist in numerous types of human cells. The tethering and structure of paraspeckles, as well as other lncRNAs, to the genome can be disrupted by small molecules that intercalate in DNA. The structure-function relationship of lncRNAs that regulates stem cell differentiation is likely to be determined by the dynamics of nucleus size and binding site accessibility.
    [Show full text]
  • Supplementary Table S4. FGA Co-Expressed Gene List in LUAD
    Supplementary Table S4. FGA co-expressed gene list in LUAD tumors Symbol R Locus Description FGG 0.919 4q28 fibrinogen gamma chain FGL1 0.635 8p22 fibrinogen-like 1 SLC7A2 0.536 8p22 solute carrier family 7 (cationic amino acid transporter, y+ system), member 2 DUSP4 0.521 8p12-p11 dual specificity phosphatase 4 HAL 0.51 12q22-q24.1histidine ammonia-lyase PDE4D 0.499 5q12 phosphodiesterase 4D, cAMP-specific FURIN 0.497 15q26.1 furin (paired basic amino acid cleaving enzyme) CPS1 0.49 2q35 carbamoyl-phosphate synthase 1, mitochondrial TESC 0.478 12q24.22 tescalcin INHA 0.465 2q35 inhibin, alpha S100P 0.461 4p16 S100 calcium binding protein P VPS37A 0.447 8p22 vacuolar protein sorting 37 homolog A (S. cerevisiae) SLC16A14 0.447 2q36.3 solute carrier family 16, member 14 PPARGC1A 0.443 4p15.1 peroxisome proliferator-activated receptor gamma, coactivator 1 alpha SIK1 0.435 21q22.3 salt-inducible kinase 1 IRS2 0.434 13q34 insulin receptor substrate 2 RND1 0.433 12q12 Rho family GTPase 1 HGD 0.433 3q13.33 homogentisate 1,2-dioxygenase PTP4A1 0.432 6q12 protein tyrosine phosphatase type IVA, member 1 C8orf4 0.428 8p11.2 chromosome 8 open reading frame 4 DDC 0.427 7p12.2 dopa decarboxylase (aromatic L-amino acid decarboxylase) TACC2 0.427 10q26 transforming, acidic coiled-coil containing protein 2 MUC13 0.422 3q21.2 mucin 13, cell surface associated C5 0.412 9q33-q34 complement component 5 NR4A2 0.412 2q22-q23 nuclear receptor subfamily 4, group A, member 2 EYS 0.411 6q12 eyes shut homolog (Drosophila) GPX2 0.406 14q24.1 glutathione peroxidase
    [Show full text]
  • The Sub-Nuclear Localization of RNA-Binding Proteins in KSHV-Infected Cells
    cells Article The Sub-Nuclear Localization of RNA-Binding Proteins in KSHV-Infected Cells Ella Alkalay, Chen Gam Ze Letova Refael, Irit Shoval, Noa Kinor, Ronit Sarid and Yaron Shav-Tal * The Mina & Everard Goodman Faculty of Life Sciences and The Institute of Nanotechnology and Advanced Materials, Bar-Ilan University, Ramat Gan 5290002, Israel; [email protected] (E.A.); [email protected] (C.G.Z.L.R.); [email protected] (I.S.); [email protected] (N.K.); [email protected] (R.S.) * Correspondence: [email protected] Received: 14 August 2020; Accepted: 21 August 2020; Published: 25 August 2020 Abstract: RNA-binding proteins, particularly splicing factors, localize to sub-nuclear domains termed nuclear speckles. During certain viral infections, as the nucleus fills up with replicating virus compartments, host cell chromatin distribution changes, ending up condensed at the nuclear periphery. In this study we wished to determine the fate of nucleoplasmic RNA-binding proteins and nuclear speckles during the lytic cycle of the Kaposi’s sarcoma associated herpesvirus (KSHV). We found that nuclear speckles became fewer and dramatically larger, localizing at the nuclear periphery, adjacent to the marginalized chromatin. Enlarged nuclear speckles contained splicing factors, whereas other proteins were nucleoplasmically dispersed. Polyadenylated RNA, typically found in nuclear speckles under regular conditions, was also found in foci separated from nuclear speckles in infected cells. Poly(A) foci did not contain lncRNAs known to colocalize with nuclear speckles but contained the poly(A)-binding protein PABPN1. Examination of the localization of spliced viral RNAs revealed that some spliced transcripts could be detected within the nuclear speckles.
    [Show full text]
  • Lncrna NEAT1 in Paraspeckles: a Structural Scaffold for Cellular DNA Damage Response Systems?
    non-coding RNA Review LncRNA NEAT1 in Paraspeckles: A Structural Scaffold for Cellular DNA Damage Response Systems? Elisa Taiana 1,2,* , Domenica Ronchetti 1,2 , Katia Todoerti 2, Lucia Nobili 1 , Pierfrancesco Tassone 3, Nicola Amodio 3 and Antonino Neri 1,2,* 1 Department of Oncology and Hemato-oncology, University of Milan, 20122 Milan, Italy; [email protected] (D.R.); [email protected] (L.N.) 2 Hematology, Fondazione Cà Granda IRCCS Policlinico, 20122 Milan, Italy; [email protected] 3 Department of Experimental and Clinical Medicine, Magna Graecia University of Catanzaro, 88100 Catanzaro, Italy; [email protected] (P.T.); [email protected] (N.A.) * Correspondence: [email protected] (E.T.); [email protected] (A.N.); Tel.: +39-02-5032-0420 (E.T. & A.N.) Received: 5 June 2020; Accepted: 28 June 2020; Published: 1 July 2020 Abstract: Nuclear paraspeckle assembly transcript 1 (NEAT1) is a long non-coding RNA (lncRNA) reported to be frequently deregulated in various types of cancers and neurodegenerative processes. NEAT1 is an indispensable structural component of paraspeckles (PSs), which are dynamic and membraneless nuclear bodies that affect different cellular functions, including stress response. Furthermore, increasing evidence supports the crucial role of NEAT1 and essential structural proteins of PSs (PSPs) in the regulation of the DNA damage repair (DDR) system. This review aims to provide an overview of the current knowledge on the involvement of NEAT1 and PSPs in DDR, which might strengthen the rationale underlying future NEAT1-based therapeutic options in tumor and neurodegenerative diseases. Keywords: lncRNA; NEAT1; paraspeckle; DNA damage repair; cancer; neurodegenerative disease 1.
    [Show full text]
  • The VE-Cadherin/Amotl2 Mechanosensory Pathway Suppresses Aortic InAmmation and the Formation of Abdominal Aortic Aneurysms
    The VE-cadherin/AmotL2 mechanosensory pathway suppresses aortic inammation and the formation of abdominal aortic aneurysms Yuanyuan Zhang Karolinska Institute Evelyn Hutterer Karolinska Institute Sara Hultin Karolinska Institute Otto Bergman Karolinska Institute Maria Forteza Karolinska Institute Zorana Andonovic Karolinska Institute Daniel Ketelhuth Karolinska University Hospital, Stockholm, Sweden Joy Roy Karolinska Institute Per Eriksson Karolinska Institute Lars Holmgren ( [email protected] ) Karolinska Institute Article Keywords: arterial endothelial cells (ECs), vascular disease, abdominal aortic aneurysms Posted Date: June 15th, 2021 DOI: https://doi.org/10.21203/rs.3.rs-600069/v1 License: This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License The VE-cadherin/AmotL2 mechanosensory pathway suppresses aortic inflammation and the formation of abdominal aortic aneurysms Yuanyuan Zhang1, Evelyn Hutterer1, Sara Hultin1, Otto Bergman2, Maria J. Forteza2, Zorana Andonovic1, Daniel F.J. Ketelhuth2,3, Joy Roy4, Per Eriksson2 and Lars Holmgren1*. 1Department of Oncology-Pathology, BioClinicum, Karolinska Institutet, Stockholm, Sweden. 2Department of Medicine Solna, BioClinicum, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden. 3Department of Cardiovascular and Renal Research, Institutet of Molecular Medicine, Univ. of Southern Denmark, Odense, Denmark 4Department of Molecular Medicine and Surgery, Karolinska Institutet, Karolinska University Hospital, Stockholm,
    [Show full text]
  • Paraspeckles
    Downloaded from http://cshperspectives.cshlp.org/ on October 2, 2021 - Published by Cold Spring Harbor Laboratory Press Paraspeckles Archa H. Fox1 and Angus I. Lamond2 1Western Australian Institute for Medical Research and Centre For Medical Research, University of Western Australia, Crawley 6009 Western Australia, Australia 2Wellcome Trust Centre for Gene Regulation & Expression, College of Life Sciences, University of Dundee DUNDEE DD1 5EH UK Correspondence: [email protected] Paraspeckles are a relatively new class of subnuclear bodies found in the interchromatin space of mammalian cells. They are RNA-protein structures formed by the interaction between a long nonprotein-coding RNA species, NEAT1/Men 1/b, and members of the DBHS (Drosophila Behavior Human Splicing) family of proteins: P54NRB/NONO, PSPC1, and PSF/SFPQ. Paraspeckles are critical to the control of gene expression through the nuclear retention of RNA containing double-stranded RNA regions that have been subject to adenosine-to-inosine editing. Through this mechanism paraspeckles and their components may ultimately have a role in controlling gene expression during many cellular processes including differentiation, viral infection, and stress responses. DISCOVERY OF PARASPECKLES human nucleoli, 271 proteins were identified, he cell nucleus is a large and complex cellu- 30% of which were novel (Andersen et al. Tlar organelle with an intricate internal or- 2002). A follow up analysis on one of these ganization that is still not fully characterized. newly identified novel proteins, showed that it One feature of nuclear organization is the was not enriched in nucleoli, but instead was presence of distinct subnuclear bodies, each of found diffusely distributed within the nucleo- which contain specific nuclear proteins and plasm as well as concentrated in 5–20 sub- nucleic acids (Platani and Lamond 2004).
    [Show full text]
  • Snapshot: Cellular Bodies David L
    SnapShot: Cellular Bodies David L. Spector Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724, USA Number/ Typical Size Marker Body Name Description Image Cell and Shape Protein Involved in snRNP and snoRNP biogenesis and 0.1–2.0 µm; Cajal Body 0–6 Coilin posttranscriptional modification of newly assembled round spliceosomal snRNAs. 20S core Contains ubiquitin conjugates, the proteolytically active 0.2–1.2 µm; catalytic Clastosome 0–3 20S core and 19S regulatory complexes of the 26S irregular component of proteasome, and protein substrates of the proteasome. proteasome Contains several factors involved in 3′ cleavage of mRNAs. 0.2–1.0 µm; Cleavage Body 1–4 CstF 64 kDa ?20% contain newly synthesized RNA. Some cleavage round bodies localize adjacent to Cajal and PML bodies. Nuclear Contains proteins for pre-mRNA processing. Involved in Speckle or 0.8–1.8 µm; SC35, 25–50 the storage, assembly, and/or modification of pre-mRNA Interchromatin irregular SF2/ASF splicing factors. Granule Cluster Induced by heat shock response. Associates with Nuclear Stress 0.3–3.0 µm; satellite III repeats on human chromosome 9q12 and 2–10 HSF1 Body irregular other pericentromeric regions; recruits various RNA- binding proteins. Contains several transcription factors (Oct1/PTF) and 1.0–1.5 µm; OPT Domain 1–3 PTF RNA transcripts; predominant in late G1 cells. Often round Nuclear Bodies localizes close to nucleolus. 0.5 µm; Contains several RNA-binding proteins and nuclear- Paraspeckle 10–20 p54nrb, PSP1 round retained CTN-RNA. Cap on surface of nucleolus; found mainly in transformed Perinucleolar 0.3–1.0 µm; 1–4 hnRNPI (PTB) cells.
    [Show full text]
  • Molecular Anatomy of the Architectural NEAT1 Noncoding
    Molecular anatomy of the architectural NEAT1 noncoding RNA : The domains, interactors, and biogenesis pathway Title required to build phase-separated nuclear paraspeckles Author(s) Hirose, Tetsuro; Yamazaki, Tomohiro; Nakagawa, Shinichi Wiley interdisciplinary reviews, RNA, 10(6), e1545 Citation https://doi.org/10.1002/wrna.1545 Issue Date 2019-11 Doc URL http://hdl.handle.net/2115/79649 This is the peer reviewed version of the following article: Tetsuro Hirose Tomohiro Yamazaki Shinichi Nakagawa. (2019) Molecular anatomy of the architectural NEAT1 noncoding RNA: The domains, interactors, and biogenesis Rights pathway required to build phase-separated nuclear paraspeckles. Wiley interdisciplinary reviews. RNA: 10(6): e1545., which has been published in final form at https://doi.org/10.1002/wrna.1545. This article may be used for non- commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. Type article (author version) File Information Wiley Interdiscip Rev RNA_10(6)_e1545.pdf Instructions for use Hokkaido University Collection of Scholarly and Academic Papers : HUSCAP Article Title: Molecular anatomy of the architectural NEAT1 noncoding RNA: the domains, interactors, and biogenesis pathway required to build phase-separated nuclear paraspeckles Article Type: Authors: First author Tetsuro Hirose*, ORCID iD: https://orcid.org/0000-0003-1068-5464, Affiliation: Institute for Genetic Medicine, Hokkaido University, E-mail address: [email protected], no conflict of interest Second author Tomohiro Yamazaki, ORCID iD: https://orcid.org/0000-0003-0411-3590, Affiliation: Institute for Genetic Medicine, Hokkaido University, E-mail address: [email protected], no conflict of interest Third author Shinichi Nakagawa, ORCID iD: https://orcid.org/0000-0002-6806-7493, Faculty of Pharmaceutical Sciences, Hokkaido University, E-mail address: [email protected], no conflict of interest Abstract Long noncoding RNAs (lncRNAs) are extremely diverse and have various significant physiological functions.
    [Show full text]
  • Glucocorticoid Receptor Condensates Link DNA-Dependent Receptor 2 Dimerization and Transcriptional Transactivation 3 4 5 Filipp Frank1*, Xu Liu1, Eric A
    bioRxiv preprint doi: https://doi.org/10.1101/2020.11.10.376327; this version posted November 10, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 1 Glucocorticoid receptor condensates link DNA-dependent receptor 2 dimerization and transcriptional transactivation 3 4 5 Filipp Frank1*, Xu Liu1, Eric A. Ortlund1* 6 1Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA 7 8 *Corresponding author: 9 Filipp Frank, 10 [email protected], 11 Emory University, 12 Department of Biochemistry, 13 1510 Clifton Rd. NE, Atlanta, GA, 30322, USA. 14 15 Eric A. Ortlund, 16 [email protected], 17 Emory University, 18 Department of Biochemistry, 19 1510 Clifton Rd. NE, Atlanta, GA, 30322, USA. bioRxiv preprint doi: https://doi.org/10.1101/2020.11.10.376327; this version posted November 10, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 20 Abstract 21 The glucocorticoid receptor (GR) is a ligand-regulated transcription factor (TF) that controls the 22 tissue- and gene-specific transactivation and transrepression of thousands of target genes. 23 Distinct GR DNA binding sequences with activating or repressive activities have been identified, 24 but how they modulate transcription in opposite ways is not known. We show that GR forms 25 phase-separated condensates that specifically concentrate known co-regulators via their 26 intrinsically disordered regions (IDRs) in vitro. A combination of dynamic, multivalent (between 27 IDRs) and specific, stable interactions (between LxxLL motifs and the GR ligand binding domain) 28 control the degree of recruitment.
    [Show full text]
  • Co-Condensation of Proteins with Single- and Double-Stranded
    bioRxiv preprint doi: https://doi.org/10.1101/2021.03.17.435834; this version posted March 17, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 Co-condensation of proteins with single- and double- 2 stranded DNA 3 4 Authors 5 Roman Renger1,2, Jose A. Morin1,3, Regis Lemaitre1, Martine Ruer-Gruss1, Frank Jülicher4,7, 6 Andreas Hermann2,5,6, Stephan W. Grill1,3,7,* 7 8 Affiliations 9 1 Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 10 Dresden, Germany. 11 2 German Center for Neurodegenerative Diseases (DZNE), Dresden, Germany 12 3 Biotechnologisches Zentrum, Technische Universität Dresden, Tatzberg 47/49, Dresden, 13 Germany. 14 4 Max Planck Institute for the Physics of Complex Systems, Nöthnitzer Straße 38, Dresden, 15 Germany 16 5 Translational Neurodegeneration Section “Albrecht-Kossel”, Department of Neurology, and 17 Center for Transdisciplinary Neurosciences, University Medical Center Rostock, University of 18 Rostock, Rostock, Germany 19 6 German Center for Neurodegenerative Diseases (DZNE), Rostock/Greifswald, Rostock, 20 Germany 21 7 Cluster of Excellence Physics of Life, Technische Universität Dresden, Dresden, Germany 22 23 24 * Correspondence to: [email protected] 25 26 Summary 27 Biomolecular condensates provide distinct compartments that can localize and organize 28 biochemistry inside cells. Recent evidence suggests that condensate formation is prevalent in 29 the cell nucleus and associated with a variety of important processes ranging from 30 transcriptional regulation to DNA repair.
    [Show full text]
  • Paraspeckles: Possible Nuclear Hubs by the RNA for the RNA
    BioMol Concepts, Vol. 3 (2012), pp. 415–428 • Copyright © by Walter de Gruyter • Berlin • Boston. DOI 10.1515/bmc-2012-0017 Review Paraspeckles: possible nuclear hubs by the RNA for the RNA Tetsuro Hirose 1, * and Shinichi Nakagawa 2 Introduction 1 Biomedicinal Information Research Center , National Institute of Advanced Industrial Science and Technology, The eukaryotic cell nucleus is highly compartmentalized. 2-4-7 Aomi, Koutou 135-0064, Tokyo , Japan More than 10 membraneless subnuclear organelles have 2 RNA Biology Laboratory , RIKEN Advanced Research been identifi ed (1, 2) . These so-called nuclear bodies exist Institute, 2-1 Hirosawa, Wako 351-0198 , Japan in the interchromosomal space, where they are enriched in multiple nuclear regulatory factors, such as transcription and * Corresponding author RNA-processing factors. These factors are thought to serve e-mail: [email protected] as specialized hubs for various nuclear events, including transcriptional regulation and RNA processing (3, 4) . Some nuclear bodies serve as sites for the biogenesis of macromo- Abstract lecular machineries, such as ribosomes and spliceosomes. Multiple cancer cell types show striking alterations in their The mammalian cell nucleus is a highly compartmental- nuclear body organization, including changes in the numbers, ized system in which multiple subnuclear structures, called shapes and sizes of certain nuclear bodies (5) . The structural nuclear bodies, exist in the nucleoplasmic spaces. Some of complexity and dynamics of nuclear bodies have been impli- the nuclear bodies contain specifi c long non-coding RNAs cated in the regulation of complex gene expression pathways (ncRNAs) as their components, and may serve as sites for in mammalian cells.
    [Show full text]