scholarly journals Nup358/RanBP2 Attaches to the Nuclear Pore Complex via Association with Nup88 and Nup214/CAN and Plays a Supporting Role in CRM1-Mediated Nuclear Protein Export

2004 ◽  
Vol 24 (6) ◽  
pp. 2373-2384 ◽  
Author(s):  
Rafael Bernad ◽  
Hella van der Velde ◽  
Maarten Fornerod ◽  
Helen Pickersgill
Keyword(s):  
Nuclear Export ◽  
Nuclear Protein ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Strong Reduction ◽  
Cytoplasmic Face ◽  
Pore Complexes ◽  
Export Signal

ABSTRACT Nuclear pore complexes (NPCs) traverse the nuclear envelope (NE), providing a channel through which nucleocytoplasmic transport occurs. Nup358/RanBP2, Nup214/CAN, and Nup88 are components of the cytoplasmic face of the NPC. Here we show that Nup88 localizes midway between Nup358 and Nup214 and physically interacts with them. RNA interference of either Nup88 or Nup214 in human cells caused a strong reduction of Nup358 at the NE. Nup88 and Nup214 showed an interdependence at the NPC and were not affected by the absence of Nup358. These data indicate that Nup88 and Nup214 mediate the attachment of Nup358 to the NPC. We show that localization of the export receptor CRM1 at the cytoplasmic face of the NE is Nup358 dependent and represents its empty state. Also, removal of Nup358 causes a distinct reduction in nuclear export signal-dependent nuclear export. We propose that Nup358 provides both a platform for rapid disassembly of CRM1 export complexes and a binding site for empty CRM1 recycling into the nucleus.

scholarly journals The nuclear export factor Xpo1p targets Mad1p to kinetochores in yeast

2009 ◽  
Vol 184 (1) ◽  
pp. 21-29 ◽  
Author(s):  
Robert J. Scott ◽  
Lucas V. Cairo ◽  
David W. Van de Vosse ◽  
Richard W. Wozniak
Keyword(s):  
Signaling Pathway ◽  
Nuclear Export ◽  
Spindle Assembly Checkpoint ◽  
Nuclear Export Signal ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Guanosine Triphosphate ◽  
Intranuclear Transport ◽  
Pore Complexes ◽  
Export Signal

Nuclear pore complexes (NPCs) mediate all nucleocytoplasmic traffic and provide docking sites for the spindle assembly checkpoint (SAC) protein Mad1p. Upon SAC activation, Mad1p is recruited onto kinetochores and rapidly cycles between NPCs and kinetochores. We examined the mechanism of Mad1p movement onto kinetochores and show that it is controlled by two components of the nuclear transport machinery, the exportin Xpo1p and Ran–guanosine triphosphate (GTP). Mad1p contains a nuclear export signal (NES) that is recognized by Xpo1p. The NES, Xpo1p, and RanGTP are all required for Mad1p recruitment onto kinetochores in checkpoint-activated cells. Consistent with this function, Xpo1p also accumulates on kinetochores after SAC activation. We have also shown that Xpo1p and RanGTP are required for the dynamic cycling of Mad1p between NPCs and kinetochores in checkpoint-arrested cells. These results reveal an important function for Xpo1p in mediating intranuclear transport events and identify a signaling pathway between kinetochores and NPCs.

scholarly journals SUMO-1 Modification and Its Role in Targeting the Ran GTPase-activating Protein, RanGAP1, to the Nuclear Pore Complex

1998 ◽  
Vol 140 (3) ◽  
pp. 499-509 ◽  
Author(s):  
Michael J. Matunis ◽  
Jian Wu ◽  
Günter Blobel
Keyword(s):  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Gtpase Activating Protein ◽  
Terminal Domain ◽  
Localization Signal

RanGAP1 is the GTPase-activating protein for Ran, a small ras-like GTPase involved in regulating nucleocytoplasmic transport. In vertebrates, RanGAP1 is present in two forms: one that is cytoplasmic, and another that is concentrated at the cytoplasmic fibers of nuclear pore complexes (NPCs). The NPC-associated form of RanGAP1 is covalently modified by the small ubiquitin-like protein, SUMO-1, and we have recently proposed that SUMO-1 modification functions to target RanGAP1 to the NPC. Here, we identify the domain of RanGAP1 that specifies SUMO-1 modification and demonstrate that mutations in this domain that inhibit modification also inhibit targeting to the NPC. Targeting of a heterologous protein to the NPC depended on determinants specifying SUMO-1 modification and also on additional determinants in the COOH-terminal domain of RanGAP1. SUMO-1 modification and these additional determinants were found to specify interaction between the COOH-terminal domain of RanGAP1 and a region of the nucleoporin, Nup358, between Ran-binding domains three and four. Together, these findings indicate that SUMO-1 modification targets RanGAP1 to the NPC by exposing, or creating, a Nup358 binding site in the COOH-terminal domain of RanGAP1. Surprisingly, the COOH-terminal domain of RanGAP1 was also found to harbor a nuclear localization signal. This nuclear localization signal, and the presence of nine leucine-rich nuclear export signal motifs, suggests that RanGAP1 may shuttle between the nucleus and the cytoplasm.

scholarly journals Altered RNA processing and export lead to retention of mRNAs near transcription sites and nuclear pore complexes or within the nucleolus

2016 ◽  
Vol 27 (17) ◽  
pp. 2742-2756 ◽  
Author(s):  
Biplab Paul ◽  
Ben Montpetit
Keyword(s):  
Single Molecule ◽  
Rna Processing ◽  
Nuclear Export ◽  
Essential Gene ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Eukaryotic Gene ◽  
Transcription Sites ◽  
Pore Complexes

Many protein factors are required for mRNA biogenesis and nuclear export, which are central to the eukaryotic gene expression program. It is unclear, however, whether all factors have been identified. Here we report on a screen of >1000 essential gene mutants in Saccharomyces cerevisiae for defects in mRNA processing and export, identifying 26 mutants with defects in this process. Single-molecule FISH data showed that the majority of these mutants accumulated mRNA within specific regions of the nucleus, which included 1) mRNAs within the nucleolus when nucleocytoplasmic transport, rRNA biogenesis, or RNA processing and surveillance was disrupted, 2) the buildup of mRNAs near transcription sites in 3′-end processing and chromosome segregation mutants, and 3) transcripts being enriched near nuclear pore complexes when components of the mRNA export machinery were mutated. These data show that alterations to various nuclear processes lead to the retention of mRNAs at discrete locations within the nucleus.

scholarly journals Exportin Crm1 is repurposed as a docking protein to generate microtubule organizing centers at the nuclear pore

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Xun X Bao ◽  
Christos Spanos ◽  
Tomoko Kojidani ◽  
Eric M Lynch ◽  
Juri Rappsilber ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Cell Types ◽  
Bound State ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Cytoplasmic Protein ◽  
Microtubule Organization ◽  
Microtubule Organizing Centers ◽  
Pore Complexes

Non-centrosomal microtubule organizing centers (MTOCs) are important for microtubule organization in many cell types. In fission yeast Schizosaccharomyces pombe, the protein Mto1, together with partner protein Mto2 (Mto1/2 complex), recruits the γ-tubulin complex to multiple non-centrosomal MTOCs, including the nuclear envelope (NE). Here, we develop a comparative-interactome mass spectrometry approach to determine how Mto1 localizes to the NE. Surprisingly, we find that Mto1, a constitutively cytoplasmic protein, docks at nuclear pore complexes (NPCs), via interaction with exportin Crm1 and cytoplasmic FG-nucleoporin Nup146. Although Mto1 is not a nuclear export cargo, it binds Crm1 via a nuclear export signal-like sequence, and docking requires both Ran in the GTP-bound state and Nup146 FG repeats. In addition to determining the mechanism of MTOC formation at the NE, our results reveal a novel role for Crm1 and the nuclear export machinery in the stable docking of a cytoplasmic protein complex at NPCs.

scholarly journals The SUMO-specific isopeptidase SENP2 associates dynamically with nuclear pore complexes through interactions with karyopherins and the Nup107-160 nucleoporin subcomplex

2011 ◽  
Vol 22 (24) ◽  
pp. 4868-4882 ◽  
Author(s):  
Jacqueline Goeres ◽  
Pak-Kei Chan ◽  
Debaditya Mukhopadhyay ◽  
Hong Zhang ◽  
Brian Raught ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Mammalian Cells ◽  
Nuclear Export Signal ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Localization Signal ◽  
Import And Export ◽  
Pore Complexes ◽  
Eukaryotic Organisms ◽  
Export Receptors

The association of small, ubiquitin-related modifier–specific isopeptidases (also known as sentrin-specific proteases, or SENPs) with nuclear pore complexes (NPCs) is conserved in eukaryotic organisms ranging from yeast to mammals. However, the functional significance of this association remains poorly understood, particularly in mammalian cells. In this study, we have characterized the molecular basis for interactions between SENP2 and NPCs in human cells. Using fluorescence recovery after photobleaching, we demonstrate that SENP2, although concentrated at the nuclear basket, is dynamically associated with NPCs. This association is mediated by multiple targeting elements within the N-terminus of SENP2 that function cooperatively to mediate NPC localization. One of these elements consists of a high-affinity nuclear localization signal that mediates indirect tethering to FG-repeat–containing nucleoporins through karyopherins. A second element mediates interactions with the Nup107-160 nucleoporin subcomplex. A third element consists of a nuclear export signal. Collectively, our findings reveal that SENP2 is tethered to NPCs through a complex interplay of interactions with nuclear import and export receptors and nucleoporins. Disruption of these interactions enhances SENP2 substrate accessibility, suggesting an important regulatory node in the SUMO pathway.

scholarly journals Active nuclear pore complexes in Chironomus: visualization of transporter configurations related to mRNP export

1998 ◽  
Vol 111 (2) ◽  
pp. 223-236 ◽  
Author(s):  
E. Kiseleva ◽  
M.W. Goldberg ◽  
T.D. Allen ◽  
C.W. Akey
Keyword(s):  
Nuclear Export ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Internal Ring ◽  
Balbiani Ring ◽  
Transmission Electron ◽  
The Individual ◽  
Pore Complexes ◽  
Small Channels

The Nuclear Pore Complex (NPC) regulates nucleocytoplasmic transport by providing small channels for passive diffusion and multiple docking surfaces that lead to a central translocation channel for active transport. In this study we have investigated by high resolution scanning and transmission electron microscopy the dynamics of NPC structure in salivary gland nuclei from Chironomus during Balbiani ring (BR) mRNP translocation, and present evidence of rearrangement of the transporter related to mRNP export. Analysis of the individual NPC components verified a strong evolutionary conservation of NPC structure between vertebrates and invertebrates. The transporter is an integral part of the NPC and is composed of a central short double cylinder that is retained within the inner spoke ring, and two peripheral globular assemblies which are tethered to the cytoplasmic and nucleoplasmic coaxial rings by eight conserved internal ring filaments. Distinct stages of BR mRNP nuclear export through the individual NPC components were directly visualized and placed in a linear transport sequence. The BR mRNP first binds to the NPC basket, which forms an expanded distal basket ring. In this communication we present stages of BR mRNP transport through the nucleoplasmic, central and cytoplasmic transporter subunits, which change their conformation during mRNP translocation, and the emergence of mRNP into the cytoplasm. We propose that the reorganization of the basket may be driven, in part, by an active translocation process at the transporter. Furthermore, the images provide dramatic evidence that the transporter functions as a central translocation channel with transiently open discrete gates in its globular assemblies. A model of NPC transporter reorganization accompanied with mRNP translocation is discussed.

scholarly journals p53 SUMOylation promotes its nuclear export by facilitating its release from the nuclear export receptor CRM1

2013 ◽  
Vol 24 (17) ◽  
pp. 2739-2752 ◽  
Author(s):  
Aleixo Santiago ◽  
Dawei Li ◽  
Lisa Y. Zhao ◽  
Adam Godsey ◽  
Daiqing Liao
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Underlying Mechanism ◽  
Binding Pocket ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Subunit Dissociation ◽  
Binding Groove ◽  
Pore Complexes ◽  
Nuclear Export Receptor

Chromosomal region maintenance 1 (CRM1) mediates p53 nuclear export. Although p53 SUMOylation promotes its nuclear export, the underlying mechanism is unclear. Here we show that tethering of a small, ubiquitin-like modifier (SUMO) moiety to p53 markedly increases its cytoplasmic localization. SUMO attachment to p53 does not affect its oligomerization, suggesting that subunit dissociation required for exposing p53’s nuclear export signal (NES) is unnecessary for p53 nuclear export. Surprisingly, SUMO-mediated p53 nuclear export depends on the SUMO-interacting motif (SIM)-binding pocket of SUMO-1. The CRM1 C-terminal domain lacking the NES-binding groove interacts with tetrameric p53, and the proper folding of the p53 core domain, rather than the presence of the N- or C-terminal tails, appears to be important for p53–CRM1 interaction. The CRM1 Huntington, EF3, a subunit of PP2A, and TOR1 9 (HEAT9) loop, which regulates GTP-binding nuclear protein Ran binding and cargo release, contains a prototypical SIM. Remarkably, disruption of this SIM in conjunction with a mutated SIM-binding groove of SUMO-1 markedly enhances the binding of CRM1 to p53-SUMO-1 and their accumulation in the nuclear pore complexes (NPCs), as well as their persistent association in the cytoplasm. We propose that SUMOylation of a CRM1 cargo such as p53 at the NPCs unlocks the HEAT9 loop of CRM1 to facilitate the disassembly of the transporting complex and cargo release to the cytoplasm.

scholarly journals Exportin Crm1 is repurposed as a docking protein to generate microtubule organizing centers at the nuclear pore

2017 ◽  
Author(s):  
Xun X. Bao ◽  
Christos Spanos ◽  
Tomoko Kojidani ◽  
Eric M. Lynch ◽  
Juri Rappsilber ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Cell Types ◽  
Bound State ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Cytoplasmic Protein ◽  
Microtubule Organization ◽  
Microtubule Organizing Centers ◽  
Pore Complexes

ABSTRACTNon-centrosomal microtubule organizing centers (MTOCs) are important for microtubule organization in many cell types. In fission yeast Schizosaccharomyces pombe, the protein Mto1, together with partner protein Mto2 (Mto1/2 complex), recruits the γ-tubulin complex to multiple non-centrosomal MTOCs, including the nuclear envelope (NE). Here, we develop a comparative-interactome mass spectrometry approach to determine how Mto1 localizes to the NE. Surprisingly, we find that Mto1, a constitutively cytoplasmic protein, docks at nuclear pore complexes (NPCs), via interaction with exportin Crm1 and cytoplasmic FG-nucleoporin Nup146. Although Mto1 is not a nuclear export cargo, it binds Crm1 via a nuclear export signal-like sequence, and docking requires both Ran in the GTP-bound state and Nup146 FG repeats. In addition to determining the mechanism of MTOC formation at the NE, our results reveal a novel role for Crm1 and the nuclear export machinery in the stable docking of a cytoplasmic protein complex at NPCs.

scholarly journals SUMO Modification of Heterogeneous Nuclear Ribonucleoproteins

2004 ◽  
Vol 24 (9) ◽  
pp. 3623-3632 ◽  
Author(s):  
Maria T. Vassileva ◽  
Michael J. Matunis
Keyword(s):  
Nuclear Export ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Sumo Modification ◽  
M Proteins ◽  
Hnrnp C ◽  
Heterogeneous Nuclear Ribonucleoproteins ◽  
Pore Complexes ◽  
And Function

ABSTRACT Small ubiquitin-related modifiers (SUMOs) are proteins that are posttranslationally conjugated to other cellular proteins, particularly those that localize and function in the nucleus. Enzymes regulating SUMO modification localize in part to nuclear pore complexes (NPCs), indicating that modification of some proteins may occur as they are translocated between the nucleus and the cytoplasm. Substrates that are regulated by SUMO modification at NPCs, however, have not been previously identified. Among the most abundant cargos transported through NPCs are the heterogeneous nuclear ribonucleoproteins (hnRNPs). HnRNPs are involved in various aspects of mRNA biogenesis, including regulation of pre-mRNA splicing and nuclear export. Here, we demonstrate that two subsets of hnRNPs, the hnRNP C and M proteins, are substrates for SUMO modification. We demonstrate that the hnRNP C proteins are modified by SUMO at a single lysine residue, K237, and that SUMO modification at this site decreases their binding to nucleic acids. We also show that Nup358, a SUMO E3 ligase associated with the cytoplasmic fibrils of NPCs, enhances the SUMO modification of the hnRNP C and M proteins. Based on our findings, we propose that SUMO modification of the hnRNP C and M proteins may occur at NPCs and facilitate the nucleocytoplasmic transport of mRNAs.

scholarly journals Intracellular Localization of Blattella germanica Densovirus (BgDV1) Capsid Proteins

Viruses ◽  
2018 ◽  
Vol 10 (7) ◽  
pp. 370 ◽  
Author(s):  
Evgeny Kozlov ◽  
Elena Martynova ◽  
Vladimir Popenko ◽  
Coby Schal ◽  
Dmitry Mukha
Keyword(s):  
Nuclear Export ◽  
Intracellular Localization ◽  
Nuclear Export Signal ◽  
Blattella Germanica ◽  
Capsid Proteins ◽  
Progeny Virus ◽  
Nuclear Pore ◽  
Genome Replication ◽  
Nuclear Pore Complexes ◽  
Pore Complexes

Densovirus genome replication and capsid assembly take place in the nucleus of the infected cells. However, the mechanisms underlying such processes as the delivery of virus proteins to the nucleus and the export of progeny virus from the nucleus remain elusive. It is evident that nuclear transport signals should be involved in these processes. We performed an in silico search for the putative nuclear localization signal (NLS) and nuclear export signal (NES) motifs in the capsid proteins of the Blattella germanica Densovirus 1 (BgDV1) densovirus. A high probability NLS motif was found in the common C-terminal of capsid proteins together with a NES motif in the unique N-terminal of VP2. We also performed a global search for the nuclear traffic signals in the densoviruses belonging to five Densovirinae genera, which revealed high diversity in the patterns of NLSs and NESs. Using a heterologous system, the HeLa mammalian cell line expressing GFP-fused BgDV1 capsid proteins, we demonstrated that both signals are functionally active. We suggest that the NLS shared by all three BgDV1 capsid proteins drives the trafficking of the newly-synthesized proteins into the nucleus, while the NES may play a role in the export of the newly-assembled BgDV1 particles into the cytoplasm through nuclear pore complexes.

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