DOCSLIB.ORG

Nup2 and a Newly Discovered Nuclear Pore Complex Protein, Nupa, Function at Mitotic Chromatin Controlled by the Nima Kinase

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

NUP2 AND A NEWLY DISCOVERED NUCLEAR PORE COMPLEX PROTEIN, NUPA, FUNCTION AT MITOTIC CHROMATIN CONTROLLED BY THE NIMA KINASE DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Sarine W. Markossian Graduate Program in Molecular Genetics The Ohio State University 2011 Dissertation Committee: Stephen A. Osmani, Ph.D., Advisor Anita K. Hopper, Ph.D. Iris Meier, Ph.D. Harold A. Fisk, Ph.D. Copyright by Sarine W. Markossian 2011 ABSTRACT In Aspergillus nidulans the mitotic NIMA kinase is required for the transition from G2 into mitosis. How NIMA regulates this process is not well understood. The nucleoporin Nup2 is of interest because it is essential and has a unique and conserved ability to transfer from nuclear pore complexes (NPCs) to chromatin specifically at mitosis. This unique transition suggests that Nup2 might play mitotic roles downstream of NIMA conserved from fungi to humans. We have determined that NIMA promotes Nup2 phosphorylation and relocation from NPCs onto chromatin. Affinity purifications followed by Mass spectroscopy identified a novel nucleoporin, NupA, which copurified with Nup2. NupA transfers from NPCs to mitotic chromatin and is also essential. Deletion analysis and heterokaryon rescue revealed that deletion of either does not cause major defects in nuclear transport but causes mitotic abnormalities that engage the spindle assembly checkpoint (SAC). We have also defined the essential function of NupA which is to tether Nup2 to both the NPC and mitotic chromatin where we propose Nup2 has a role in promoting efficient generation and function of the mitotic apparatus. In addition, Nup2 and NupA have dynamic locations within mitotic chromatin during different phases of mitosis. They are associated throughout chromatin at early prophase but move to locate at the periphery of the chromatin during the transition from anaphase to telophase. This movement at anaphase is likely functionally significant as deletion of either Nup2 or NupA causes defects in mitotic exit into G1. Hence, Nup2 and NupA may have dual roles in mitosis, the first being during early mitosis, defects in which activates the SAC, and the latter in mitotic exit and early G1. Because Nup2 co- purifies with ii importins α and , we propose that both roles might be linked to proper localizations and functions of importins α and during metaphase and mitotic exit. These studies collectively demonstrate that Nup2 and the newly discovered NupA have important functions during mitosis that are controlled by NIMA. iii To my mother, Iskouhi Shamikian, for providing me the opportunity of higher education and teaching me to believe that everything is possible through hard work. iv ACKNOWLEDGEMENTS I am deeply indebted to my advisor Dr. Stephen A. Osmani for his continual guidance and constant support throughout the years of my graduate studies. I thoroughly enjoyed his mentorship and his friendship. I would also like to thank my current and previous committee members, Dr. Anita K. Hopper, Dr. Iris Meier, Dr. Harold A. Fisk, and Dr. Berl R. Oakley for their valuable inputs into my work and all the encouragement during my graduate studies. In addition, I would like to thank all current and former members of the Osmani lab for their friendship, the assistance provided with experiments, the rewarding discussions about my work, and the continuous and critical feedback they have provided throughout the years. I would like to particularly thank Aysha H. Osmani, Subbulakshmi Suresh and Jessica S. El-Hallal because this work would not have been possible without their fruitful collaborations. I would also like to thank all the Molecular Genetics family, particularly my classmates for all the fun times, their friendship, and the extreme encouragement and support they provided throughout this experience. Finally, I would like to thank my parents, my sister, and all the rest of my extended family, including friends that I was lucky enough to encounter throughout the years in Lebanon and in Ohio. It is through their constant and unconditional love, faith, support and encouragement that this achievement was possible. v VITA 2001………………………………………B.S. Biology, American University of Beirut 2003………………………………………M.S. Biology, American University of Beirut 2004 to present …………………………..Graduate Research Associate, Department of Molecular Genetics, The Ohio State University PUBLICATIONS Liu HL, Osmani AH, Ukil L, Son S, Markossian S, Shen KF, Govindaraghavan M, Varadaraj A, Hashmi SB, De Souza CP, Osmani SA. 2010. Single step affinity purification for fungal proteomics. Eukaryot Cell 9:831-3. Kreydiyyeh S.I., Markossian S., Hodeify R.F. 2006. PGE2 exerts dose-dependent opposite effects on net water and chloride absorption from the rat colon. Prostaglandins and Other Lipid Mediators 79:43-52. vi Kreydiyyeh S.I., Markossian S. 2006. Tumor necrosis factor alpha down-regulates the Na+-K+ ATPase and the Na+-K+2Cl- cotransporter in the kidney cortex and medulla. Cytokine 33:138-44. Markossian S., Kreydiyyeh S.I. 2005. TNF-alpha down-regulates the Na+-K+ ATPase and the Na+-K+-2Cl-cotransporter in the rat colon via PGE2. Cytokine 30:319-27. Markossian S.W. 2003. Effect of TNF-α on electrolyte transport across epithelia : possible mechanism of action. M.S. Thesis. Beirut: American University of Beirut. Kreydiyyeh S.I., Usta J., Knio K., Markossian S., Dagher S. 2003. Aniseed oil increases glucose absorption and reduces urine output in the rat. Life Sciences 74:663-73. FIELDS OF STUDY Major Field: Molecular Genetics vii TABLE OF CONTENTS ABSTRACT ........................................................................................................................ ii ACKNOWLEDGEMENTS ................................................................................................ v VITA .................................................................................................................................. vi PUBLICATIONS ............................................................................................................... vi FIELDS OF STUDY......................................................................................................... vii TABLE OF CONTENTS ................................................................................................. viii LIST OF TABLES ........................................................................................................... xiv LIST OF FIGURES .......................................................................................................... xv LIST OF ABBREVIATIONS ........................................................................................ xviii CHAPTER 1 ....................................................................................................................... 1 INTRODUCTION .............................................................................................................. 1 1.1 Project Goal .................................................................................................................. 1 1.2 Aspergillus nidulans as a model system to study mitosis ............................................. 2 1.2.1 Biology ................................................................................................................... 2 1.2.2 Genome assembly and annotation .......................................................................... 4 1.2.3 Advantages ............................................................................................................. 5 1.3 The Nucleus .................................................................................................................. 6 1.3.1 Overview ................................................................................................................ 6 1.3.2 Differences in nuclear structure between lower and higher eukaryotes ................. 7 1.3.3 The nuclear envelope (NE) .................................................................................... 7 1.3.4 The Nuclear Pore Complex .................................................................................... 9 1.3.4.1 Overview ......................................................................................................... 9 1.3.4.2 Isolation of Nups ........................................................................................... 10 1.3.4.3 NPC biogenesis into an intact NE ................................................................. 10 1.3.5 Nucleoporin 2 (Nup2) .......................................................................................... 11 1.3.5.1 Protein structure ............................................................................................. 11 viii 1.3.5.2 Isolation and characterization ........................................................................ 12 1.3.6 Nucleo-cytoplasmic transport .............................................................................. 14 1.3.6.1 Overview ....................................................................................................... 14 1.3.6.2. Role of Nup2 in transport ............................................................................. 15 1.3.7 Nuclear chromatin ................................................................................................ 16 1.3.7.1 Overview ......................................................................................................
Recommended publications
  • Transcriptional Regulation Constrains the Organization of Genes on Eukaryotic Chromosomes

    Transcriptional Regulation Constrains the Organization of Genes on Eukaryotic Chromosomes

    Transcriptional regulation constrains the organization of genes on eukaryotic chromosomes Sarath Chandra Janga*†, Julio Collado-Vides‡, and M. Madan Babu*† *Laboratory of Molecular Biology, Medical Research Council, Hills Road, Cambridge CB2 0QH, United Kingdom; and ‡Programa de Genomica Computacional, Centro de Ciencias Genomicas, Universidad Nacional Autonoma de Mexico, Apartado Postal 565-A, Av Universidad, Cuernavaca, Morelos, 62100 Mexico D.F., Mexico Edited by Aaron Klug, Medical Research Council, Cambridge, United Kingdom, and approved August 21, 2008 (received for review July 1, 2008) Genetic material in eukaryotes is tightly packaged in a hierarchical organization of genes across the different eukaryotic chromosomes. manner into multiple linear chromosomes within the nucleus. This becomes particularly interesting in the light of a recent work Although it is known that eukaryotic transcriptional regulation is that demonstrated that tuning the expression level of a single gene complex and requires an intricate coordination of several molec- could provide an enormous fitness advantage to an individual in a ular events both in space and time, whether the complexity of this population of cells (12). Thus, one could extrapolate that optimi- process constrains genome organization is still unknown. Here, we zation of transcriptional regulation on a global scale, such as the present evidence for the existence of a higher-order organization efficient expression of relevant genes under specific conditions, of genes across and within chromosomes that is constrained by would have significant advantage on the fitness of an individual in transcriptional regulation. In particular, we reveal that the target a genetically heterogeneous population. genes (TGs) of transcription factors (TFs) for the yeast, Saccharo- Although several studies have reported that genes with similar myces cerevisiae, are encoded in a highly ordered manner both expression patterns cluster on the genome and that gene order is across and within the 16 chromosomes.
  • Chapter 1: Introduction

    Chapter 1: Introduction

    The role of lamin A and emerin in mediating genome organisation A thesis for the degree of Doctor of Philosophy by Lauren Sarah Godwin School of Health Sciences and Social Care Brunel University July 2010 Abstract The nuclear matrix (NM) is proposed to be a permanent network of core filaments underlying thicker fibres, present regardless of transcriptional activity. It is found to be both RNA and protein rich; indeed, numerous important nuclear proteins are components of the structure. In addition to mediating the organisation of entire chromosomes, the NM has also been demonstrated to tether telomeres via their TTAGGG repeats. In order to examine telomeric interactions with the NM, a technique known as the DNA halo preparation has been employed. Regions of DNA that are tightly attached to the structure are found within a so-called residual nucleus, while those sequences forming lesser associations produce a halo of DNA. Coupled with various FISH methodologies, this technique allowed the anchorage of genomic regions by the NM, to be analysed. In normal fibroblasts, the majority of chromosomes and telomeres were extensively anchored to the NM. Such interactions did not vary significantly in proliferating and senescent nuclei. However, a decrease in NM-associated telomeres was detected in quiescence. Since lamin A is an integral component of the NM, it seemed pertinent to examine chromosome and telomere NM-anchorage in Hutchinson-Gilford Progeria Syndrome (HGPS) fibroblasts, which contain mutant forms of lamin A. Indeed, genome tethering by the NM was perturbed in HGPS. In immortalised HGPS fibroblasts, this disrupted anchorage appeared to be rescued; the implications of this finding will be discussed.
  • Nuclear Pore Proteins and the Control of Genome Functions

    Nuclear Pore Proteins and the Control of Genome Functions

    Downloaded from genesdev.cshlp.org on September 30, 2021 - Published by Cold Spring Harbor Laboratory Press REVIEW Nuclear pore proteins and the control of genome functions Arkaitz Ibarra and Martin W. Hetzer Molecular and Cell Biology Laboratory, Salk Institute for Biological Studies, La Jolla, California 92037, USA Nuclear pore complexes (NPCs) are composed of several Cytoplasmic filaments are mainly formed by Nup358/ copies of ~30 different proteins called nucleoporins (Nups). RanBP2, Nup214, and Nup88, while the nuclear basket is NPCs penetrate the nuclear envelope (NE) and regulate the composed of Nup153 and Tpr (Fig. 1; for Nup othologs, see nucleocytoplasmic trafficking of macromolecules. Beyond Rothballer and Kutay 2012). this vital role, NPC components influence genome func- The selective access of regulatory factors into the tions in a transport-independent manner. Nups play an nucleus and export of specific RNA molecules mediated evolutionarily conserved role in gene expression regulation by the NPC is required for the accurate progression of most that, in metazoans, extends into the nuclear interior. major cellular processes. However, our perception of Additionally, in proliferative cells, Nups play a crucial role the NPC components is rapidly evolving, as accumulating in genome integrity maintenance and mitotic progression. evidence indicates that they can also directly impact Here we discuss genome-related functions of Nups and DNA metabolism by genome-related functions (Liang their impact on essential DNA metabolism processes such and Hetzer 2011). Among these, one of the most remark- as transcription, chromosome duplication, and segregation. able and well-conserved roles of Nups is to associate with specific target genes to regulate their transcriptional activity (Casolari et al.
  • Module IV Nucleus

    Module IV Nucleus

    Module IV Nucleus Structure and functions of interphase nucleus, Nuclear membrane, pore complex, structure and functions of nucleolus Chromosomes – Structure; Heterochromatin, Euchromatin, Nucleosomes, Nucleus is the most important part of the cell situated in the cytoplasm. All the cellular activities are controlled by it. Nucleus is a directing and organizing unit without which the cell could not exist. It was discovered by Robert Brown (1831) in flowering plants and is now recognized as the structure that contains the hereditary material of the cell. The study of nucleus or karyosome constitutes karyology. The location of nucleus varies in the cell depending upon the species. Usually it is situated in the centre of the cell surrounded on all sides by cytoplasm. In green algae, Acetabularia, it shows various positions, though mainly present in the basal part of cell. Generally the nuclei are scattered in the cytoplasm. Morphology: 1. Shape: The shape of nucleus is variable according to cell type. It is generally spheroid but ellipsoid or flattened nuclei may also occur in certain cells. The nuclear margins are generally smooth but they may be lobulated bearing small infoldings of nuclear membrane as in leucocytes. In certain white blood corpuscles the nucleus is dumbbell-shaped and exhibits variation during life history stages. In human neutrophil, it is trilobed. 2. Number: Mostly cell contains a single nucleus, known as mononucleate cell. Cells containing two nuclei are known as binucleate cells (e.g., Paramecium), and cells of cartilage and liver. Sometimes more than two nuclei (3 to 100 nuclei) are present in a single cell.
  • Molecular Genetics of Microcephaly Primary Hereditary: an Overview

    Molecular Genetics of Microcephaly Primary Hereditary: an Overview

    brain sciences Review Molecular Genetics of Microcephaly Primary Hereditary: An Overview Nikistratos Siskos † , Electra Stylianopoulou †, Georgios Skavdis and Maria E. Grigoriou * Department of Molecular Biology & Genetics, Democritus University of Thrace, 68100 Alexandroupolis, Greece; [email protected] (N.S.); [email protected] (E.S.); [email protected] (G.S.) * Correspondence: [email protected] † Equal contribution. Abstract: MicroCephaly Primary Hereditary (MCPH) is a rare congenital neurodevelopmental disorder characterized by a significant reduction of the occipitofrontal head circumference and mild to moderate mental disability. Patients have small brains, though with overall normal architecture; therefore, studying MCPH can reveal not only the pathological mechanisms leading to this condition, but also the mechanisms operating during normal development. MCPH is genetically heterogeneous, with 27 genes listed so far in the Online Mendelian Inheritance in Man (OMIM) database. In this review, we discuss the role of MCPH proteins and delineate the molecular mechanisms and common pathways in which they participate. Keywords: microcephaly; MCPH; MCPH1–MCPH27; molecular genetics; cell cycle 1. Introduction Citation: Siskos, N.; Stylianopoulou, Microcephaly, from the Greek word µικρoκεϕαλi´α (mikrokephalia), meaning small E.; Skavdis, G.; Grigoriou, M.E. head, is a term used to describe a cranium with reduction of the occipitofrontal head circum- Molecular Genetics of Microcephaly ference equal, or more that teo standard deviations
  • Nuclear Pore Complexes and Nucleocytoplasmic Exchange

    Nuclear Pore Complexes and Nucleocytoplasmic Exchange

    Pore Relations: Nuclear Pore Complexes and Nucleocytoplasmic Exchange Michael P. Rout and John D. Aitchison Laboratory of Cellular and Structural Biology The Rockefeller University, 1230 York Ave, New York, NY 10021 USA [email protected] 212 327 8135 Department of Cell Biology University of Alberta Edmonton, Alberta T6G 2H7 Canada [email protected] 780 492 6062 1 Introduction One of the main characteristics distinguishing eukaryotes from prokaryotes is that eukaryotes compartmentalize many life processes within membrane bound organelles. The most obvious of these is the nucleus, bounded by a double-membraned nuclear envelope (NE). The NE thus acts as a barrier separating the nucleoplasm from the cytoplasm. An efficient, regulated and continuous exchange system between the nucleoplasm and cytoplasm is therefore necessary to maintain the structures of the nucleus and the communication between the genetic material and the rest of the cell. The sole mediators of this exchange are the nuclear pore complexes (NPCs), large proteinaceous assemblies embedded within reflexed pores of the NE membranes (Davis, 1995). While small molecules (such as nucleotides, water and ions) can freely diffuse across the NPCs, macromolecules such as proteins and ribonucleoprotein (RNP) particles are actively transported in a highly regulated and selective manner. Transport through the NPC requires specific soluble factors which recognize transport substrates in either the nucleoplasm or cytoplasm and mediate their transport by docking them to specific components of the NPC (Mattaj and Englmeier, 1998). In order to understand how transport works, we must first catalog the soluble transport factors and NPC components, and then study the details of how they interact.
  • Targeting the Dosage Compensation Complex to the Male X

    Targeting the Dosage Compensation Complex to the Male X

    PDF hosted at the Radboud Repository of the Radboud University Nijmegen The following full text is a publisher's version. For additional information about this publication click this link. http://hdl.handle.net/2066/65589 Please be advised that this information was generated on 2021-09-24 and may be subject to change. Targeting the dosage compensation complex to the male X chromosome of Drosophila melanogaster Targeting the dosage compensation complex to the male X chromosome of Drosophila melanogaster Gene Expression programme European Molecular Biology Laboratory (EMBL) Heidelberg, Germany And Department of Molecular Biology Nijmegen Centre for Molecular Life Sciences (NCMLS) Radboud University Nijmegen, The Netherlands Published by the Radboud University Nijmegen Printed by Ponsen & Looijen bv, Wageningen ISBN/EAN: 978-90-9023194-5 Cover: Artistic interpretation of a Drosophila polytene chromosome squash Targeting the dosage compensation complex to the male X chromosome in Drosophila melanogaster Een wetenschappelijke proeve op het gebied van de Natuurwetenschappen, Wiskunde en Informatica Proefschrift ter verkrijging van de graad van doctor aan de Radboud Universiteit Nijmegen op gezag van de rector magnificus prof. mr. S.CJ.J. Kortmann volgens besluit van het College van Decanen in het openbaar te verdedigen op vrijdag 20 juni 2008 om 10.30 uur precies door Jop Haico Kind geboren op 28 december 1978 te Lelystad Promotor: prof. dr. H.G. Stunnenberg Copromotor: dr. A. Akhtar, European Moleculair Biology Laboratory Heidelberg, Duitsland
  • Functional Studies of Nuclear Envelope-Associated Proteins in Saccharomyces Cerevisiae

    Functional Studies of Nuclear Envelope-Associated Proteins in Saccharomyces Cerevisiae

    Functional studies of nuclear envelope-associated proteins in Saccharomyces cerevisiae Ida Olsson Stockholm University © Ida Olsson, Stockholm 2008 ISBN 978-91-7155-666-0, pp 1-58 Typesetting: Intellecta Docusys Printed in Sweden by Universitetsservice US-AB, Stockholm 2008 Distributor: Department of Biochemistry and Biophysics, Stockholm University To Carl with love ABSTRACT Proteins of the nuclear envelope play important roles in a variety of cellular processes e.g. transport of proteins between the nucleus and cytoplasm, co- ordination of nuclear and cytoplasmic events, anchoring of chromatin to the nuclear periphery and regulation of transcription. Defects in proteins of the nuclear envelope and the nuclear pore complexes have been related to a number of human diseases. To understand the cellular functions in which nuclear envelope proteins participate it is crucial to map the functions of these proteins. The present study was done in order to characterize the role of three different proteins in functions related to the nuclear envelope in the yeast Saccharo- myces cerevisiae. The arginine methyltransferase Rmt2 was demonstrated to associate with proteins of the nuclear pore complexes and to influence nu- clear export. In addition, Rmt2 was found to interact with the Lsm4 protein involved in RNA degradation, splicing and ribosome biosynthesis. These results provide support for a role of Rmt2 at the nuclear periphery and poten- tially in nuclear transport and RNA processing. The integral membrane pro- tein Cwh43 was localized to the inner nuclear membrane and was also found at the nucleolus. A nuclear function for Cwh43 was demonstrated by its abil- ity to bind DNA in vitro.
  • Condensins Exert Force on Chromatin-Nuclear Envelope Tethers to Mediate Nucleoplasmic Reticulum Formation in Drosophila Melanogaster

    Condensins Exert Force on Chromatin-Nuclear Envelope Tethers to Mediate Nucleoplasmic Reticulum Formation in Drosophila Melanogaster

    INVESTIGATION Condensins Exert Force on Chromatin-Nuclear Envelope Tethers to Mediate Nucleoplasmic Reticulum Formation in Drosophila melanogaster Julianna Bozler,* Huy Q. Nguyen,* Gregory C. Rogers,† and Giovanni Bosco*,1 *Geisel School of Medicine at Dartmouth, Hanover, New Hampshire 03755, and †Department of Cellular and Molecular Medicine, University of Arizona Cancer Center, University of Arizona, Tucson, Arizona 85724 ABSTRACT Although the nuclear envelope is known primarily for its role as a boundary between the KEYWORDS nucleus and cytoplasm in eukaryotes, it plays a vital and dynamic role in many cellular processes. Studies of nuclear nuclear structure have revealed tissue-specific changes in nuclear envelope architecture, suggesting that its architecture three-dimensional structure contributes to its functionality. Despite the importance of the nuclear envelope, chromatin force the factors that regulate and maintain nuclear envelope shape remain largely unexplored. The nuclear nucleus envelope makes extensive and dynamic interactions with the underlying chromatin. Given this inexorable chromatin link between chromatin and the nuclear envelope, it is possible that local and global chromatin organization compaction reciprocally impact nuclear envelope form and function. In this study, we use Drosophila salivary glands to nuclear envelope show that the three-dimensional structure of the nuclear envelope can be altered with condensin II- mediated chromatin condensation. Both naturally occurring and engineered chromatin-envelope interac- tions are sufficient to allow chromatin compaction forces to drive distortions of the nuclear envelope. Weakening of the nuclear lamina further enhanced envelope remodeling, suggesting that envelope struc- ture is capable of counterbalancing chromatin compaction forces. Our experiments reveal that the nucle- oplasmic reticulum is born of the nuclear envelope and remains dynamic in that they can be reabsorbed into the nuclear envelope.
  • O-Fucosylated Glycoproteins Form Assemblies in Close Proximity to the Nuclear Pore Complexes of Toxoplasma Gondii

    O-Fucosylated Glycoproteins Form Assemblies in Close Proximity to the Nuclear Pore Complexes of Toxoplasma Gondii

    O-fucosylated glycoproteins form assemblies in close proximity to the nuclear pore complexes of Toxoplasma gondii Giulia Bandinia, John R. Hasericka,b, Edwin Motaria,b,1, Dinkorma T. Ouologuemc, Sebastian Louridod, David S. Roosc, Catherine E. Costellob, Phillips W. Robbinsa,2, and John Samuelsona,2 aDepartment of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118; bDepartment of Biochemistry, Boston University School of Medicine, Boston, MA 02118; cDepartment of Biology, University of Pennsylvania, Philadelphia, PA 19104; and dWhitehead Institute for Biomedical Research, Cambridge, MA 02142 Contributed by Phillips W. Robbins, August 19, 2016 (sent for review June 8, 2016; reviewed by Richard Cummings and John A. Hanover) Toxoplasma gondii is an intracellular parasite that causes dissem- (FG-Nups) and a putative Nup54 can be predicted by primary inated infections in fetuses and immunocompromised individuals. sequence homology (20). Although gene regulation is important for parasite differentiation Here we report the discovery of numerous assemblies of and pathogenesis, little is known about protein organization in O-fucosylated proteins that associate with the nuclear membrane the nucleus. Here we show that the fucose-binding Aleuria aurantia near the NPCs. These results improve our understanding of the ar- lectin (AAL) binds to numerous punctate structures in the nuclei of chitecture of the T. gondii nuclear periphery and highlight O-fuco- tachyzoites, bradyzoites, and sporozoites but not oocysts. AAL also sylation as a PTM involved in assemblies associated with the NPC. binds to Hammondia and Neospora nuclei but not to more distantly Results related apicomplexans. Analyses of the AAL-enriched fraction indi- cate that AAL binds O-linked fucose added to Ser/Thr residues pre- The Fucose-Binding Aleuria aurantia Lectin Labels the Nuclei of T.
  • Perspectives

    Perspectives

    FOCUS ON MECHANOTRANSDUCTION PERSPECTIVES network that can promote coordinated OPINION changes in cell, cytoskeletal and nuclear struc- ture in response to mechanical distortion14 Mechanotransduction at a (FIG. 1a). (Herein, the term hard-wired refers to cytoskeletal structures that are stable enough distance: mechanically coupling the as interconnected units to resist mechanical stresses and thereby maintain shape stabil- ity, even though they undergo continuous extracellular matrix with the nucleus dynamic remodelling at the molecular level.) This model takes into account the observa- Ning Wang, Jessica D. Tytell and Donald E. Ingber tion that individual cytoskeletal filaments can bear significant tensile and compressive loads Abstract | Research in cellular mechanotransduction often focuses on how in living cells because their structural integrity extracellular physical forces are converted into chemical signals at the cell surface. is maintained for longer than the turnover However, mechanical forces that are exerted on surface-adhesion receptors, such time of individual protein monomers15–17. as integrins and cadherins, are also channelled along cytoskeletal filaments and Key to the cellular tensegrity model is concentrated at distant sites in the cytoplasm and nucleus. Here, we explore the the idea that overall cell-shape stability and long-distance force transfer are governed by molecular mechanisms by which forces might act at a distance to induce the level of isometric tension, or ‘prestress’, mechanochemical conversion in the nucleus and alter gene activities. in the cytoskeleton that is generated through the establishment of a force balance between Mechanical forces influence the growth and For example, endothelial cells sense fluid opposing structural elements (that is, micro- shape of virtually every tissue and organ in shear through a cell–cell junctional com- tubules, contractile microfilaments and our bodies.
  • The Nucleolus As a Multiphase Liquid Condensate

    The Nucleolus As a Multiphase Liquid Condensate

    REVIEWS The nucleolus as a multiphase liquid condensate Denis L. J. Lafontaine 1 ✉ , Joshua A. Riback 2, Rümeyza Bascetin 1 and Clifford P. Brangwynne 2,3 ✉ Abstract | The nucleolus is the most prominent nuclear body and serves a fundamentally important biological role as a site of ribonucleoprotein particle assembly, primarily dedicated to ribosome biogenesis. Despite being one of the first intracellular structures visualized historically, the biophysical rules governing its assembly and function are only starting to become clear. Recent studies have provided increasing support for the concept that the nucleolus represents a multilayered biomolecular condensate, whose formation by liquid–liquid phase separation (LLPS) facilitates the initial steps of ribosome biogenesis and other functions. Here, we review these biophysical insights in the context of the molecular and cell biology of the nucleolus. We discuss how nucleolar function is linked to its organization as a multiphase condensate and how dysregulation of this organization could provide insights into still poorly understood aspects of nucleolus-associated diseases, including cancer, ribosomopathies and neurodegeneration as well as ageing. We suggest that the LLPS model provides the starting point for a unifying quantitative framework for the assembly, structural maintenance and function of the nucleolus, with implications for gene regulation and ribonucleoprotein particle assembly throughout the nucleus. The LLPS concept is also likely useful in designing new therapeutic strategies to target nucleolar dysfunction. Protein trans-acting factors Among numerous microscopically visible nuclear sub- at the inner core where rRNA transcription occurs and Proteins important for structures, the nucleolus is the most prominent and proceeding towards the periphery (Fig.