scholarly journals Cortical Recruitment and Nuclear–Cytoplasmic Shuttling of Scd5p, a Protein Phosphatase-1-targeting Protein Involved in Actin Organization and Endocytosis

2006 ◽  
Vol 17 (1) ◽  
pp. 251-262 ◽  
Author(s):  
Ji Suk Chang ◽  
Kenneth Henry ◽  
María Isabel Geli ◽  
Sandra K. Lemmon
Keyword(s):  
Amino Acids ◽  
Protein Phosphatase ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Protein Phosphatase 1 ◽  
Dependent Manner ◽  
Repeat Region ◽  
Cortical Actin ◽  
Cortical Function ◽  
Actin Organization

Scd5p regulates endocytosis and cortical actin organization as a targeting subunit for the Ser/Thr protein phosphatase-1 (PP1) in yeast. To identify localization signals in Scd5p required for cell surface recruitment, visualization of GFP-tagged Scd5 truncations and deletions was performed. Scd5p contains a PP1 binding site, a 3-repeat region of 20 amino acids (3R), and a 9-repeat region of 12 amino acids (9R). We found that the 9R is critical for cortical localization of Scd5p, but cortical recruitment is not essential for Scd5p's function in actin organization and endocytosis. We propose that Scd5p can target PP1 to endocytic factors in the cytoplasm that have been disassembled and/or inactivated by phosphorylation. We also found that Scd5p undergoes nuclear-cytoplasmic shuttling in a Crm1p-dependent manner. Scd5p-ΔCT lacking the 9R region and its nuclear export signal (NES) accumulates in the nucleus, causing cortical actin and endocytic defects. Cytoplasmic localization and function of Scd5p-ΔCT is restored by NES addition. However, removal of Scd5p's nuclear localization signal prevents nuclear entry, but endocytosis and actin organization remain relatively normal. These results indicate that nuclear-cytoplasmic shuttling is not required for regulation of Scd5p's cortical function and suggest that Scd5p has an independent nuclear function.

scholarly journals OsWRKY62 and OsWRKY76 interact with Importin α1s for Negative Regulation of Defensive Responses in Nucleus

2021 ◽  
Author(s):  
Xiaohui Xu ◽  
Han Wang ◽  
Jiqin Liu ◽  
Shuying Han ◽  
Miaomiao Lin ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nuclear Localization ◽  
Nuclear Export ◽  
Nuclear Translocation ◽  
Nuclear Export Signal ◽  
Defense Responses ◽  
Nuclear Localization Signals ◽  
Defensive Responses ◽  
Export Signal ◽  
Positively Charged

Abstract Background: OsWRKY62 and OsWRKY76, two close members of WRKY transcription factors, function together as transcriptional repressors. OsWRKY62 is predominantly localized in the cytosol. What are the regulatory factors for OsWRKY62 nuclear translocation?Results: In this study, we characterized they interacted with rice importin, OsIMα1a and OsIMα1b, for nuclear translocation. Chimeric OsWRKY62.1-GFP, which is predominantly localized in the cytoplasm, was translocated to the nucleus of Nicotiana benthamiana leaf cells in the presence of OsIMα1a or OsIMαDIBB1a lacking the auto-inhibitory importin β-binding domain. OsIMαDIBB1a interacted with the WRKY domain of OsWRKY62.1, which has specific bipartite positively charged concatenated amino acids functioning as a nuclear localization signal. Similarly, we found that OsIMαDIBB1a interacted with the AvrPib effector of rice blast fungus Magnaporthe oryzae, which contains a scattered distribution of positively charged amino acids. Furthermore, we identified a nuclear export signal in OsWRKY62.1 that inhibited nuclear transportation. Overexpression of OsIMα1a or OsIMα1b enhanced resistance to M. oryzae, whereas knockout mutants decreased resistance to the pathogen. However, overexpressing both OsIMα1a and OsWRKY62.1 were slightly more susceptible to M. oryzae than OsWRKY62.1 alone. Ectopic overexpression of OsWRKY62.1 with an extra nuclear export signal compromised the enhanced susceptibility of OsWRKY62.1 to M. oryzae.Conclusion: These results indicated that OsWRKY62 localization is a consequence of competition binding between rice importins and exportins. OsWRKY62, OsWRKY76, and AvrPib effector translocate to nucleus in association with importin α1s through new types of nuclear localization signals for negatively regulating defense responses.

scholarly journals Identification of the Nuclear Export Signal and STAT-Binding Domains of the Nipah Virus V Protein Reveals Mechanisms Underlying Interferon Evasion

2004 ◽  
Vol 78 (10) ◽  
pp. 5358-5367 ◽  
Author(s):  
Jason J. Rodriguez ◽  
Cristian D. Cruz ◽  
Curt M. Horvath
Keyword(s):  
Amino Acids ◽  
Protein Interactions ◽  
Nuclear Export ◽  
Nipah Virus ◽  
Nuclear Export Signal ◽  
Hendra Virus ◽  
Nuclear Accumulation ◽  
V Protein ◽  
Binding Domains ◽  
Export Signal

ABSTRACT The V proteins of Nipah virus and Hendra virus have been demonstrated to bind to cellular STAT1 and STAT2 proteins to form high-molecular-weight complexes that inhibit interferon (IFN)-induced antiviral transcription by preventing STAT nuclear accumulation. Analysis of the Nipah virus V protein has revealed a region between amino acids 174 and 192 that functions as a CRM1-dependent nuclear export signal (NES). This peptide is sufficient to complement an export-defective human immunodeficiency virus Rev protein, and deletion and substitution mutagenesis revealed that this peptide is necessary for both V protein shuttling and cytoplasmic retention of STAT1 and STAT2 proteins. However, the NES is not required for V-dependent IFN signaling inhibition. IFN signaling is blocked primarily by interaction between Nipah virus V residues 100 to 160 and STAT1 residues 509 to 712. Interaction with STAT2 requires a larger Nipah virus V segment between amino acids 100 and 300, but deletion of residues 230 to 237 greatly reduced STAT2 coprecipitation. Further, V protein interactions with cellular STAT1 is a prerequisite for STAT2 binding, and sequential immunoprecipitations demonstrate that V, STAT1, and STAT2 can form a tripartite complex. These findings characterize essential regions for Henipavirus V proteins that represent potential targets for therapeutic intervention.

scholarly journals A Regulated Nucleocytoplasmic Shuttle Contributes to Bright's Function as a Transcriptional Activator of Immunoglobulin Genes

2006 ◽  
Vol 26 (6) ◽  
pp. 2187-2201 ◽  
Author(s):  
Dongkyoon Kim ◽  
Philip W. Tucker
Keyword(s):  
Cell Cycle ◽  
Growth Factor ◽  
Nuclear Matrix ◽  
Nuclear Export ◽  
Cellular Localization ◽  
Cell Cycle Progression ◽  
Chromosome Region ◽  
Nuclear Export Signal ◽  
Variable Region ◽  
Dependent Manner

ABSTRACT Bright/ARID3a has been implicated in mitogen- and growth factor-induced up-regulation of immunoglobulin heavy-chain (IgH) genes and in E2F1-dependent G1/S cell cycle progression. For IgH transactivation, Bright binds to nuclear matrix association regions upstream of certain variable region promoters and flanking the IgH intronic enhancer. While Bright protein was previously shown to reside within the nuclear matrix, we show here that a significant amount of Bright resides in the cytoplasm of normal and transformed B cells. Leptomycin B, chromosome region maintenance 1 (CRM1) overexpression, and heterokaryon experiments indicate that Bright actively shuttles between the nucleus and the cytoplasm in a CRM1-dependent manner. We mapped the functional nuclear localization signal to the N-terminal region of REKLES, a domain conserved within ARID3 paralogues. Residues within the C terminus of REKLES contain its nuclear export signal, whose regulation is primarily responsible for Bright shuttling. Growth factor depletion and cell synchronization experiments indicated that Bright shuttling during S phase of the cell cycle leads to an increase in its nuclear abundance. Finally, we show that shuttle-incompetent Bright point mutants, even if sequestered within the nucleus, are incapable of transactivating an IgH reporter gene. Therefore, regulation of Bright's cellular localization appears to be required for its function.

scholarly journals RanGTP-Binding Protein NXT1 Facilitates Nuclear Export of Different Classes of RNA In Vitro

2000 ◽  
Vol 20 (13) ◽  
pp. 4562-4571 ◽  
Author(s):  
Batool Ossareh-Nazari ◽  
Christèle Maison ◽  
Ben E. Black ◽  
Lyne Lévesque ◽  
Bryce M. Paschal ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Small Gtpase ◽  
Dependent Manner ◽  
Vitro Assay ◽  
U1 Snrna ◽  
Rna Export ◽  
Gtpase Ran ◽  
Export Signal

ABSTRACT To better characterize the mechanisms responsible for RNA export from the nucleus, we developed an in vitro assay based on the use of permeabilized HeLa cells. This new assay supports nuclear export of U1 snRNA, tRNA, and mRNA in an energy- and Xenopusextract-dependent manner. U1 snRNA export requires a 5′ monomethylated cap structure, the nuclear export signal receptor CRM1, and the small GTPase Ran. In contrast, mRNA export does not require the participation of CRM1. We show here that NXT1, an NTF2-related protein that binds directly to RanGTP, strongly stimulates export of U1 snRNA, tRNA, and mRNA. The ability of NXT1 to promote export is dependent on its capacity to bind RanGTP. These results support the emerging view that NXT1 is a general export factor, functioning on both CRM1-dependent and CRM1-independent pathways of RNA export.

scholarly journals The Mobile FG Nucleoporin Nup98 Is a Cofactor for Crm1-dependent Protein Export

2010 ◽  
Vol 21 (11) ◽  
pp. 1885-1896 ◽  
Author(s):  
Masahiro Oka ◽  
Munehiro Asally ◽  
Yoshinari Yasuda ◽  
Yutaka Ogawa ◽  
Taro Tachibana ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Nuclear Protein ◽  
Mutational Analysis ◽  
Specific Inhibitor ◽  
Protein Export ◽  
Dependent Manner ◽  
Repeat Region ◽  
Leptomycin B ◽  
Dependent Protein ◽  
Nuclear Export Receptor

Nup98 is a mobile nucleoporin that forms distinct dots in the nucleus, and, although a role for Nup98 in nuclear transport has been suggested, its precise function remains unclear. Here, we show that Nup98 plays an important role in Crm1-mediated nuclear protein export. Nuclear, but not cytoplasmic, dots of EGFP-tagged Nup98 disappeared rapidly after cell treatment with leptomycin B, a specific inhibitor of the nuclear export receptor, Crm1. Mutational analysis demonstrated that Nup98 physically and functionally interacts with Crm1 in a RanGTP-dependent manner through its N-terminal phenylalanine-glycine (FG) repeat region. Moreover, the activity of the Nup98-Crm1 complex was modulated by RanBP3, a known cofactor for Crm1-mediated nuclear export. Finally, cytoplasmic microinjection of anti-Nup98 inhibited the Crm1-dependent nuclear export of proteins, concomitant with the accumulation of anti-Nup98 in the nucleus. These results clearly demonstrate that Nup98 functions as a novel shuttling cofactor for Crm1-mediated nuclear export in conjunction with RanBP3.

scholarly journals The microRNA cluster miR-183/96/182 contributes to long-term memory in a protein phosphatase 1-dependent manner

2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Bisrat T. Woldemichael ◽  
Ali Jawaid ◽  
Eloïse A. Kremer ◽  
Niharika Gaur ◽  
Jacek Krol ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Protein Phosphatase 1 ◽  
Long Term Memory ◽  
Dependent Manner ◽  
Term Memory ◽  
Microrna Cluster

scholarly journals IPP5, a novel protein inhibitor of protein phosphatase 1, promotes G1/S progression in a Thr-40-dependent manner.

2015 ◽  
Vol 290 (10) ◽  
pp. 6004-6004
Author(s):  
Xiaojian Wang ◽  
Bin Liu ◽  
Nan Li ◽  
Hongzhe Li ◽  
Jianming Qiu ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Protein Phosphatase 1 ◽  
Dependent Manner ◽  
Protein Inhibitor ◽  
Novel Protein

scholarly journals Characterization of the Nuclear Localization and Nuclear Export Signals of Bovine Herpesvirus 1 VP22

2005 ◽  
Vol 79 (18) ◽  
pp. 11864-11872 ◽  
Author(s):  
Chunfu Zheng ◽  
Robert Brownlie ◽  
Lorne A. Babiuk ◽  
Sylvia van Drunen Littel-van den Hurk
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Nuclear Localization ◽  
Nuclear Export ◽  
Yellow Fluorescent Protein ◽  
Nuclear Export Signal ◽  
Bovine Herpesvirus ◽  
Nuclear Targeting ◽  
Bovine Herpesvirus 1 ◽  
Herpesvirus 1

ABSTRACT The bovine herpesvirus 1 (BHV-1) tegument protein VP22 is predominantly localized in the nucleus after viral infection. To analyze subcellular localization in the absence of other viral proteins, a plasmid expressing BHV-1 VP22 fused to enhanced yellow fluorescent protein (EYFP) was constructed. The transient expression of VP22 fused to EYFP in COS-7 cells confirmed the predominant nuclear localization of VP22. Analysis of the amino acid sequence of VP22 revealed that it does not have a classical nuclear localization signal (NLS). However, by constructing a series of deletion derivatives, we mapped the nuclear targeting domain of BHV-1 VP22 to amino acids (aa) 121 to 139. Furthermore, a 4-aa motif, 130PRPR133, was able to direct EYFP and an EYFP dimer (dEYFP) or trimer (tEYFP) predominantly into the nucleus, whereas a deletion or mutation of this arginine-rich motif abrogated the nuclear localization property of VP22. Thus, 130PRPR133 is a functional nonclassical NLS. Since we observed that the C-terminal 68 aa of VP22 mediated the cytoplasmic localization of EYFP, an analysis was performed on these C-terminal amino acid sequences, and a leucine-rich motif, 204LDRMLKSAAIRIL216, was detected. Replacement of the leucines in this putative nuclear export signal (NES) with neutral amino acids resulted in an exclusive nuclear localization of VP22. Furthermore, this motif was able to localize EYFP and dEYFP in the cytoplasm, and the nuclear export function of this NES could be blocked by leptomycin B. This demonstrates that this leucine-rich motif is a functional NES. These data represent the first identification of a functional NLS and NES in a herpesvirus VP22 homologue.

scholarly journals Characterization of Staufen 1 ribonucleoprotein complexes

2004 ◽  
Vol 384 (2) ◽  
pp. 239-246 ◽  
Author(s):  
Cornelia BRENDEL ◽  
Monika REHBEIN ◽  
Hans-Jürgen KREIENKAMP ◽  
Friedrich BUCK ◽  
Dietmar RICHTER ◽  
...  
Keyword(s):  
Protein Phosphatase ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Protein Phosphatase 1 ◽  
Cell Fractionation ◽  
Dependent Manner ◽  
Ribonucleoprotein Complexes ◽  
Ribonucleoprotein Particles ◽  
Purification Protocol

In Drosophila oocytes and neuroblasts, the double-stranded RNA binding protein Staufen assembles into ribonucleoprotein particles, which mediate cytoplasmic mRNA trafficking and translation. Two different mammalian orthologues also appear to reside in distinct RNA-containing particles. To date, relatively little is known about the molecular composition of Staufen-containing ribonucleoprotein complexes. Here, we have used a novel one-step affinity purification protocol to identify components of Staufen 1-containing particles. Whereas the nucleocytoplasmic RNA-binding protein nucleolin is linked to Staufen in an RNA-dependent manner, the association of protein phosphatase 1, the microtubule-dependent motor protein kinesin and several components of the large and small ribosomal subunits with Staufen ribonucleoprotein complexes is RNA-independent. Notably, all these components do not co-purify with a second RNA-binding protein, hnRNPK (heterogeneous ribonucleoprotein K), demonstrating the high specificity of the purification protocol. Furthermore, pull-down and immunoprecipitation experiments suggest a direct interaction between Staufen 1 and the ribosomal protein P0 in vitro as well as in cells. In cell fractionation and sucrose gradient assays, Staufen co-fractionates with intact ribosomes and polysomes, but not with the isolated 40 S ribosomal subunit. Taken together, these findings imply that, in the cytoplasm of mammalian cells, an association with the ribosomal P-stalk protein P0 recruits Staufen 1 into ribosome-containing ribonucleoprotein particles, which also contain kinesin, protein phosphatase 1 and nucleolin.

scholarly journals NESdb: a database of NES-containing CRM1 cargoes

2012 ◽  
Vol 23 (18) ◽  
pp. 3673-3676 ◽  
Author(s):  
Darui Xu ◽  
Nick V. Grishin ◽  
Yuh Min Chook
Keyword(s):  
Amino Acids ◽  
Nonprofit Organizations ◽  
Experimental Evidence ◽  
Cell Nucleus ◽  
Nuclear Export ◽  
Nuclear Export Signal ◽  
Hydrophobic Residues ◽  
Cargo Protein ◽  
Targeting Signal ◽  
Export Signal

The leucine-rich nuclear export signal (NES) is the only known class of targeting signal that directs macromolecules out of the cell nucleus. NESs are short stretches of 8–15 amino acids with regularly spaced hydrophobic residues that bind the export karyopherin CRM1. NES-containing proteins are involved in numerous cellular and disease processes. We compiled a database named NESdb that contains 221 NES-containing CRM1 cargoes that were manually curated from the published literature. Each NESdb entry is annotated with information about sequence and structure of both the NES and the cargo protein, as well as information about experimental evidence of NES-mapping and CRM1-mediated nuclear export. NESdb will be updated regularly and will serve as an important resource for nuclear export signals. NESdb is freely available to nonprofit organizations at http://prodata.swmed.edu/LRNes .

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