Origin of the nuclear proteome on the basis of pre-existing nuclear localization signals in prokaryotic proteins

AbstractThe origin of the selective nuclear protein import machinery, which consists of nuclear pore complexes and adaptor molecules interacting with the nuclear localization signals (NLSs) of cargo molecules, was one of the most important events in the evolution of the eukaryotic cell. How the proteins were selected for import into the forming nuclei remains an open question. Here, we demonstrate that functional NLSs may be integrated inside nucleotide-binding domains of both eukaryotic and prokaryotic proteins and may co-evolve with these domains. We propose that the pre-existence of NLSs inside prokaryotic proteins dictated, at least partially, the nuclear proteome composition.

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Evolutionary development of redundant nuclear localization signals in the mRNA export factor NXF1

Molecular Biology of the Cell ◽

10.1091/mbc.e11-03-0222 ◽

2011 ◽

Vol 22 (23) ◽

pp. 4657-4668 ◽

Cited By ~ 15

Author(s):

Zi Chao Zhang ◽

Neal Satterly ◽

Beatriz M. A. Fontoura ◽

Yuh Min Chook

Keyword(s):

Nuclear Localization ◽

Nuclear Import ◽

Mrna Export ◽

Evolutionary Development ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Nuclear Localization Signals ◽

Basic Segment ◽

Importin Α ◽

Pore Complexes

In human cells, the mRNA export factor NXF1 resides in the nucleoplasm and at nuclear pore complexes. Karyopherin β2 or transportin recognizes a proline–tyrosine nuclear localization signal (PY-NLS) in the N-terminal tail of NXF1 and imports it into the nucleus. Here biochemical and cellular studies to understand the energetic organization of the NXF1 PY-NLS reveal unexpected redundancy in the nuclear import pathways used by NXF1. Human NXF1 can be imported via importin β, karyopherin β2, importin 4, importin 11, and importin α. Two NLS epitopes within the N-terminal tail, an N-terminal basic segment and a C-terminal R-X2-5-P-Y motif, provide the majority of binding energy for all five karyopherins. Mutation of both NLS epitopes abolishes binding to the karyopherins, mislocalized NXF1 to the cytoplasm, and significantly compromised its mRNA export function. The understanding of how different karyopherins recognize human NXF1, the examination of NXF1 sequences from divergent eukaryotes, and the interactions of NXF1 homologues with various karyopherins reveals the evolutionary development of redundant NLSs in NXF1 of higher eukaryotes. Redundancy of nuclear import pathways for NXF1 increases progressively from fungi to nematodes and insects to chordates, potentially paralleling the increasing complexity in mRNA export regulation and the evolution of new nuclear functions for NXF1.

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Nuclear localization signals for four distinct karyopherin-β nuclear import systems

Biochemical Journal ◽

10.1042/bj20150368 ◽

2015 ◽

Vol 468 (3) ◽

pp. 353-362 ◽

Cited By ~ 74

Author(s):

Michael Soniat ◽

Yuh Min Chook

Keyword(s):

Nuclear Localization ◽

Nuclear Export ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Paper Briefly ◽

Nuclear Localization Signals ◽

Importin Α ◽

Pore Complexes ◽

First Time ◽

Transport Of Macromolecules

The Karyopherin-β family of proteins mediates nuclear transport of macromolecules. Nuclear versus cytoplasmic localization of proteins is often suggested by the presence of NLSs (nuclear localization signals) or NESs (nuclear export signals). Import-Karyopherin-βs or Importins bind to NLSs in their protein cargos to transport them through nuclear pore complexes into the nucleus. Until recently, only two classes of NLS had been biochemically and structurally characterized: the classical NLS, which is recognized by the Importin-α/β heterodimer and the PY-NLS (proline–tyrosine NLS), which is recognized by Karyopherin-β2 or Transportin-1. Structures of two other Karyopherin-βs, Kap121 and Transportin-SR2, in complex with their respective cargos were reported for the first time recently, revealing two new distinct classes of NLSs. The present paper briefly describes the classical NLS, reviews recent literature on the PY-NLS and provides in-depth reviews of the two newly discovered classes of NLSs that bind Kap121p and Transportin-SR respectively.

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Targeting and function in mRNA export of nuclear pore complex protein Nup153.

The Journal of Cell Biology ◽

10.1083/jcb.134.5.1141 ◽

1996 ◽

Vol 134 (5) ◽

pp. 1141-1156 ◽

Cited By ~ 136

Author(s):

R Bastos ◽

A Lin ◽

M Enarson ◽

B Burke

Keyword(s):

Nuclear Pore Complex ◽

Protein Import ◽

Nucleocytoplasmic Transport ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Terminal Domain ◽

General Decline ◽

Complex Protein ◽

Pore Complexes ◽

And Function

Nup153 is a large (153 kD) O-linked glyco-protein which is a component of the basket structure located on the nucleoplasmic face of nuclear pore complexes. This protein exhibits a tripartite structure consisting of a zinc finger domain flanked by large (60-70 kD) NH2- and COOH-terminal domains. When full-length human Nup153 is expressed in BHK cells, it accumulates appropriately at the nucleoplasmic face of the nuclear envelope. Targeting information for Nup153 resides in the NH2-terminal domain since this region of the molecule can direct an ordinarily cytoplasmic protein, pyruvate kinase, to the nuclear face of the nuclear pore complex. Overexpression of Nup153 results in the dramatic accumulation of nuclear poly (A)+ RNA, suggesting an inhibition of RNA export from the nucleus. This is not due to a general decline in nucleocytoplasmic transport or to occlusion or loss of nuclear pore complexes since nuclear protein import is unaffected. While overexpression of certain Nup153 constructs was found to result in the formation of unusual intranuclear membrane arrays, this structural phenotype could not be correlated with the effects on poly (A)+ RNA distribution. The RNA trafficking defect was, however, dependent upon the Nup153 COOH-terminal domain which contains most of the XFXFG repeats. It is proposed that this region of Nup153, lying within the distal ring of the nuclear basket, represents a docking site for mRNA molecules exiting the nucleus.

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Dissection of the NUP107 nuclear pore subcomplex reveals a novel interaction with spindle assembly checkpoint protein MAD1 in Caenorhabditis elegans

Molecular Biology of the Cell ◽

10.1091/mbc.e11-11-0927 ◽

2012 ◽

Vol 23 (5) ◽

pp. 930-944 ◽

Cited By ~ 31

Author(s):

Eduardo Ródenas ◽

Cristina González-Aguilera ◽

Cristina Ayuso ◽

Peter Askjaer

Keyword(s):

Caenorhabditis Elegans ◽

Nuclear Envelope ◽

Molecular Mechanisms ◽

Spindle Assembly Checkpoint ◽

Protein Import ◽

Spindle Assembly ◽

Nucleocytoplasmic Transport ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Pore Complexes

Nuclear pore complexes consist of several subcomplexes. The NUP107 complex is important for nucleocytoplasmic transport, nuclear envelope assembly, and kinetochore function. However, the underlying molecular mechanisms and the roles of individual complex members remain elusive. We report the first description of a genetic disruption of NUP107 in a metazoan. Caenorhabditis elegans NUP107/npp-5 mutants display temperature-dependent lethality. Surprisingly, NPP-5 is dispensable for incorporation of most nucleoporins into nuclear pores and for nuclear protein import. In contrast, NPP-5 is essential for proper kinetochore localization of NUP133/NPP-15, another NUP107 complex member, whereas recruitment of NUP96/NPP-10C and ELYS/MEL-28 is NPP-5 independent. We found that kinetochore protein NUF2/HIM-10 and Aurora B/AIR-2 kinase are less abundant on mitotic chromatin upon NPP-5 depletion. npp-5 mutants are hypersensitive to anoxia, suggesting that the spindle assembly checkpoint (SAC) is compromised. Indeed, NPP-5 interacts genetically and physically with SAC protein MAD1/MDF-1, whose nuclear envelope accumulation requires NPP-5. Thus our results strengthen the emerging connection between nuclear pore proteins and chromosome segregation.

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Dynamic Rearrangement of Nucleoporins during Fungal “Open” Mitosis

Molecular Biology of the Cell ◽

10.1091/mbc.e07-02-0130 ◽

2008 ◽

Vol 19 (3) ◽

pp. 1230-1240 ◽

Cited By ~ 37

Author(s):

Ulrike Theisen ◽

Anne Straube ◽

Gero Steinberg

Keyword(s):

Nuclear Envelope ◽

Protein Import ◽

Complex Dynamics ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Nuclear Pores ◽

Basic Principles ◽

Early Anaphase ◽

Pore Complexes ◽

Corn Smut

Mitosis in animals starts with the disassembly of the nuclear pore complexes and the breakdown of the nuclear envelope. In contrast to many fungi, the corn smut fungus Ustilago maydis also removes the nuclear envelope. Here, we report on the dynamic behavior of the nucleoporins Nup214, Pom152, Nup133, and Nup107 in this “open” fungal mitosis. In prophase, the nuclear pore complexes disassembled and Nup214 and Pom152 dispersed in the cytoplasm and in the endoplasmic reticulum, respectively. Nup107 and Nup133 initially spread throughout the cytoplasm, but in metaphase and early anaphase occurred on the chromosomes. In anaphase, the Nup107-subcomplex redistributed to the edge of the chromosome masses, where the new envelope was reconstituted. Subsequently, Nup214 and Pom152 are recruited to the nuclear pores and protein import starts. Recruitment of nucleoporins and protein import reached a steady state in G2 phase. Formation of the nuclear envelope and assembly of nuclear pores occurred in the absence of microtubules or F-actin, but not if both were disrupted. Thus, the basic principles of nuclear pore complex dynamics seem to be conserved in organisms displaying open mitosis.

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Caenorhabditis elegans Nucleoporins Nup93 and Nup205 Determine the Limit of Nuclear Pore Complex Size Exclusion In Vivo

Molecular Biology of the Cell ◽

10.1091/mbc.e03-04-0237 ◽

2003 ◽

Vol 14 (12) ◽

pp. 5104-5115 ◽

Cited By ~ 124

Author(s):

Vincent Galy ◽

Iain W. Mattaj ◽

Peter Askjaer

Keyword(s):

Caenorhabditis Elegans ◽

Nuclear Envelope ◽

Protein Import ◽

Chromatin Condensation ◽

Size Exclusion ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

C Elegans ◽

Pore Complexes

Nuclear pore complexes (NPCs) span the nuclear envelope and mediate communication between the nucleus and the cytoplasm. To obtain insight into the structure and function of NPCs of multicellular organisms, we have initiated an extensive analysis of Caenorhabditis elegans nucleoporins. Of 20 assigned C. elegans nucleoporin genes, 17 were found to be essential for embryonic development either alone or in combination. In several cases, depletion of nucleoporins by RNAi caused severe defects in nuclear appearance. More specifically, the C. elegans homologs of vertebrate Nup93 and Nup205 were each found to be required for normal NPC distribution in the nuclear envelope in vivo. Depletion of Nup93 or Nup205 caused a failure in nuclear exclusion of nonnuclear macromolecules of ∼70 kDa without preventing active nuclear protein import or the assembly of the nuclear envelope. The defects in NPC exclusion were accompanied by abnormal chromatin condensation and early embryonic arrest. Thus, the contribution to NPC structure of Nup93 and Nup205 is essential for establishment of normal NPC function and for cell viability.

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Importins and Exportins Regulating Allergic Immune Responses

Mediators of Inflammation ◽

10.1155/2014/476357 ◽

2014 ◽

Vol 2014 ◽

pp. 1-14 ◽

Cited By ~ 18

Author(s):

Ankita Aggarwal ◽

Devendra K. Agrawal

Keyword(s):

Transcription Factors ◽

Nuclear Export ◽

Molecular Mechanisms ◽

Allergic Diseases ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Immune Modulators ◽

Nuclear Localization Signals ◽

Intracellular Signals ◽

Pore Complexes

Nucleocytoplasmic shuttling of macromolecules is a well-controlled process involving importins and exportins. These karyopherins recognize and bind to receptor-mediated intracellular signals through specific signal sequences that are present on cargo proteins and transport into and out of the nucleus through nuclear pore complexes. Nuclear localization signals (NLS) present on cargo molecules to be imported while nuclear export signals (NES) on the molecules to be exported are recognized by importins and exportins, respectively. The classical NLS are found on many transcription factors and molecules that are involved in the pathogenesis of allergic diseases. In addition, several immune modulators, including corticosteroids and vitamin D, elicit their cellular responses by regulating the expression and activity of importin molecules. In this review article, we provide a comprehensive list of importin and exportin molecules and their specific cargo that shuttled between cytoplasm and the nucleus. We also critically review the role and regulation of specific importin and exportin involved in the transport of activated transcription factors in allergic diseases, the underlying molecular mechanisms, and the potential target sites for developing better therapeutic approaches.

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Nuclear mRNA export requires specific FG nucleoporins for translocation through the nuclear pore complex

The Journal of Cell Biology ◽

10.1083/jcb.200704174 ◽

2007 ◽

Vol 178 (7) ◽

pp. 1121-1132 ◽

Cited By ~ 84

Author(s):

Laura J. Terry ◽

Susan R. Wente

Keyword(s):

Messenger Rna ◽

Large Scale ◽

Protein Import ◽

Global Analysis ◽

Mrna Export ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Versus Protein ◽

Transport Receptors ◽

Pore Complexes

Trafficking of nucleic acids and large proteins through nuclear pore complexes (NPCs) requires interactions with NPC proteins that harbor FG (phenylalanine-glycine) repeat domains. Specialized transport receptors that recognize cargo and bind FG domains facilitate these interactions. Whether different transport receptors utilize preferential FG domains in intact NPCs is not fully resolved. In this study, we use a large-scale deletion strategy in Saccharomyces cerevisiae to generate a new set of more minimal pore (mmp) mutants that lack specific FG domains. A comparison of messenger RNA (mRNA) export versus protein import reveals unique subsets of mmp mutants with functional defects in specific transport receptors. Thus, multiple functionally independent NPC translocation routes exist for different transport receptors. Our global analysis of the FG domain requirements in mRNA export also finds a requirement for two NPC substructures—one on the nuclear NPC face and one in the NPC central core. These results pinpoint distinct steps in the mRNA export mechanism that regulate NPC translocation efficiency.

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Nup214 Is Required for CRM1-Dependent Nuclear Protein Export In Vivo

Molecular and Cellular Biology ◽

10.1128/mcb.00342-06 ◽

2006 ◽

Vol 26 (18) ◽

pp. 6772-6785 ◽

Cited By ~ 101

Author(s):

Saskia Hutten ◽

Ralph H. Kehlenbach

Keyword(s):

Protein Import ◽

Nuclear Protein ◽

Biochemical Properties ◽

Protein Export ◽

Nuclear Pore ◽

Minor Effect ◽

Nuclear Pore Complexes ◽

High Affinity ◽

Pore Complexes

ABSTRACT Nucleoporins mediate transport of macromolecules across the nuclear pore complex, yet the function of many individual nucleoporins is largely unresolved. To address this question, we depleted cells of the cytoplasmic nucleoporins Nup214/CAN and Nup358/RanBP2 by RNA interference. Depletion of Nup214 resulted in codepletion of its binding partner, Nup88. Nuclear pore complexes assembled in the absence of Nup214/Nup88 or Nup358 were fully functional in nuclear protein import, whereas nuclear mRNA export was slightly impaired. Depletion of Nup358 had only a minor effect on nuclear protein export. In contrast, depletion of Nup214/Nup88 led to strongly reduced CRM1-mediated export of the shuttling transcription factor NFAT as well as a human immunodeficiency virus-Rev derivative. A specific role of Nup214 in protein export is furthered by the biochemical properties of a high-affinity complex containing Nup214, CRM1, RanGTP, and an export cargo. Our results show that the Nup214/Nup88 complex is required for efficient CRM1-mediated transport, supporting a model involving a high-affinity binding site for CRM1 at Nup214 in the terminal steps of export.

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Functional domains in nuclear import factor p97 for binding the nuclear localization sequence receptor and the nuclear pore.

Molecular Biology of the Cell ◽

10.1091/mbc.8.6.945 ◽

1997 ◽

Vol 8 (6) ◽

pp. 945-956 ◽

Cited By ~ 55

Author(s):

N C Chi ◽

S A Adam

Keyword(s):

Nuclear Localization ◽

Receptor Binding ◽

Protein Import ◽

Binding Domain ◽

Nuclear Localization Sequence ◽

Functional Domains ◽

Nuclear Pore ◽

Binding Domains ◽

Localization Sequence ◽

Ran Gtp

The interaction of the nuclear protein import factor p97 with the nuclear localization sequence (NLS) receptor, the nuclear pore complex, and Ran/TC4 is important for coordinating the events of protein import to the nucleus. We have mapped the binding domains on p97 for the NLS receptor and the nuclear pore. The NLS receptor-binding domain of p97 maps to the C-terminal 60% of the protein between residues 356 and 876. The pore complex-binding domain of p97 maps to residues 152-352. The pore complex-binding domain overlaps the Ran-GTP- and Ran-GDP-binding domains on p97, but only Ran-GTP competes for docking in permeabilized cells. The N-ethylmaleimide sensitivity of the p97 for docking was investigated and found to be due to inhibition of p97 binding to the pore complex and to the NLS receptor. Site-directed mutagenesis of conserved cysteine residues in the pore- and receptor-binding domains identified two cysteines, C223 and C228, that were required for p97 to bind the nuclear pore. Inhibition studies on docking and accumulation of a NLS protein provided additional evidence that the domains identified biochemically are the functional domains involved in protein import. Together, these results suggest that Ran-GTP dissociates the receptor complex and prevents p97 binding to the pore by inducing a conformational change in the structure of p97 rather than simple competition for binding sites.

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