Nuclear Pore Complex Protein Mediated Nuclear Localization of Dicer Protein in Human Cells

Tpr is a 270-kD coiled-coil protein localized to intranuclear filaments of the nuclear pore complex (NPC). The mechanism by which Tpr contributes to the structure and function of the nuclear pore is currently unknown. To gain insight into Tpr function, we expressed the full-length protein and several subdomains in mammalian cell lines and examined their effects on nuclear pore function. Through this analysis, we identified an NH2-terminal domain that was sufficient for association with the nucleoplasmic aspect of the NPC. In addition, we unexpectedly found that the acidic COOH terminus was efficiently transported into the nuclear interior, an event that was apparently mediated by a putative nuclear localization sequence. Ectopic expression of the full-length Tpr caused a dramatic accumulation of poly(A)+ RNA within the nucleus. Similar results were observed with domains that localized to the NPC and the nuclear interior. In contrast, expression of these proteins did not appear to affect nuclear import. These data are consistent with a model in which Tpr is tethered to intranuclear filaments of the NPC by its coiled coil domain leaving the acidic COOH terminus free to interact with soluble transport factors and mediate export of macromolecules from the nucleus.

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Simple kinetic relationships and nonspecific competition govern nuclear import rates in vivo

The Journal of Cell Biology ◽

10.1083/jcb.200608141 ◽

2006 ◽

Vol 175 (4) ◽

pp. 579-593 ◽

Cited By ~ 100

Author(s):

Benjamin L. Timney ◽

Jaclyn Tetenbaum-Novatt ◽

Diana S. Agate ◽

Rosemary Williams ◽

Wenzhu Zhang ◽

...

Keyword(s):

Nuclear Localization ◽

Nuclear Pore Complex ◽

Nuclear Import ◽

Yeast Cells ◽

Limiting Factor ◽

Nuclear Pore ◽

Nuclear Localization Signals ◽

The Poor ◽

Cytoplasmic Proteins

Many cargoes destined for nuclear import carry nuclear localization signals that are recognized by karyopherins (Kaps). We present methods to quantitate import rates and measure Kap and cargo concentrations in single yeast cells in vivo, providing new insights into import kinetics. By systematically manipulating the amounts, types, and affinities of Kaps and cargos, we show that import rates in vivo are simply governed by the concentrations of Kaps and their cargo and the affinity between them. These rates fit to a straightforward pump–leak model for the import process. Unexpectedly, we deduced that the main limiting factor for import is the poor ability of Kaps and cargos to find each other in the cytoplasm in a background of overwhelming nonspecific competition, rather than other more obvious candidates such as the nuclear pore complex and Ran. It is likely that most of every import round is taken up by Kaps and nuclear localization signals sampling other cytoplasmic proteins as they locate each other in the cytoplasm.

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From Signalosome to Enhanceosome: The Translocation of CD40 from Membrane to Nucleus Involves Multiple Mechanisms

Blood ◽

10.1182/blood.v112.11.3781.3781 ◽

2008 ◽

Vol 112 (11) ◽

pp. 3781-3781

Author(s):

Hai-Jun Zhou ◽

Archie Tamayo ◽

Lan Pham ◽

Yen-Chiu Lin-Lee ◽

Richard J. Ford

Keyword(s):

Endoplasmic Reticulum ◽

B Cell ◽

B Lymphocytes ◽

Nuclear Localization ◽

Cell Nucleus ◽

Nuclear Pore Complex ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Nuclear Pore ◽

Lymphoma Cells

Abstract CD40 plays important roles in the proliferation, survival and differentiation of lymphocytes. Constitutively active CD40 recruits TRAFs and IKKs within the lipid rafts to form a signalosome that mediates pivotal downstream proliferation and survival mechanisms involving NF-kB. Recently, we have reported that nuclear localization of CD40, through its interaction with c-Rel, promotes growth, cell cycle progression and survival in large B cell lymphoma. Our studies have opened a new paradigm in the functional role of CD40 in non-Hodgkin lymphomas of B cell origin (NHL-B). However, the mechanism about how CD40 enters nuclear still remains elusive. In this study, we show that CD40 ligation enhances its nuclear accumulation with activation of c-Rel in both normal B-lymphocytes and B cell lymphoma cells with cell fractionation assay and con-focal microscopy. Over-expression of c-Rel in B cell lymphoma cells drives CD40 into cell nucleus. We hypothesize that the route CD40 enters nucleus may involve endosome-endoplasmic reticulum-nuclear pore complex. Indeed, further studies show CD40 co-localizes with endosome marker-EEA1 and endoplasmic reticulum marker-Sec61. Furthermore, our co-immunoprecipitation assay has demonstrated CD40 interacts with Sec61. CD40 also co-localizes and immuno-precipitates with nuclear pore complex (NPC) proteins-NUP62 in normal B-lymphocytes and B lymphoma cells, which suggests NPC proteins may facilitate the nuclear translocation of CD40 protein. Overall, our study suggests that translocation of CD40 into cell nucleus involves multiple pathways. Blocking nuclear localization may modulate the function of CD40 in lymphoma cells; which could provide a new-targeted therapeutic approach for lymphoma therapy.

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ADP-ribose polymer depletion leads to nuclear Ctcf re-localization and chromatin rearrangement1

Biochemical Journal ◽

10.1042/bj20121429 ◽

2013 ◽

Vol 449 (3) ◽

pp. 623-630 ◽

Cited By ~ 18

Author(s):

Tiziana Guastafierro ◽

Angela Catizone ◽

Roberta Calabrese ◽

Michele Zampieri ◽

Oliviano Martella ◽

...

Keyword(s):

Nuclear Localization ◽

Nuclear Pore Complex ◽

De Novo ◽

Nuclear Organization ◽

Ctcf Binding ◽

Nuclear Pore ◽

Active Chromatin ◽

Lamin B1 ◽

Binding Factor ◽

Parp Activity

Ctcf (CCCTC-binding factor) directly induces Parp [poly(ADP-ribose) polymerase] 1 activity and its PARylation [poly(ADPribosyl)ation] in the absence of DNA damage. Ctcf, in turn, is a substrate for this post-synthetic modification and as such it is covalently and non-covalently modified by PARs (ADP-ribose polymers). Moreover, PARylation is able to protect certain DNA regions bound by Ctcf from DNA methylation. We recently reported that de novo methylation of Ctcf target sequences due to overexpression of Parg [poly(ADP-ribose)glycohydrolase] induces loss of Ctcf binding. Considering this, we investigate to what extent PARP activity is able to affect nuclear distribution of Ctcf in the present study. Notably, Ctcf lost its diffuse nuclear localization following PAR (ADP-ribose polymer) depletion and accumulated at the periphery of the nucleus where it was linked with nuclear pore complex proteins remaining external to the perinuclear Lamin B1 ring. We demonstrated that PAR depletion-dependent perinuclear localization of Ctcf was due to its blockage from entering the nucleus. Besides Ctcf nuclear delocalization, the outcome of PAR depletion led to changes in chromatin architecture. Immunofluorescence analyses indicated DNA redistribution, a generalized genomic hypermethylation and an increase of inactive compared with active chromatin marks in Parg-overexpressing or Ctcf-silenced cells. Together these results underline the importance of the cross-talk between Parp1 and Ctcf in the maintenance of nuclear organization.

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The Nuclear Pore Complex Protein Elys Is Required for Genome Stability in Mouse Intestinal Epithelial Progenitor Cells

Gastroenterology ◽

10.1053/j.gastro.2011.01.048 ◽

2011 ◽

Vol 140 (5) ◽

pp. 1547-1555.e10 ◽

Cited By ~ 24

Author(s):

Nan Gao ◽

Gangarao Davuluri ◽

Weilong Gong ◽

Christoph Seiler ◽

Kristin Lorent ◽

...

Keyword(s):

Progenitor Cells ◽

Nuclear Pore Complex ◽

Genome Stability ◽

Nuclear Pore ◽

Intestinal Epithelial ◽

Complex Protein ◽

Epithelial Progenitor Cells

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A Small Ubiquitin-Related Polypeptide Involved in Targeting RanGAP1 to Nuclear Pore Complex Protein RanBP2

Cell ◽

10.1016/s0092-8674(00)81862-0 ◽

1997 ◽

Vol 88 (1) ◽

pp. 97-107 ◽

Cited By ~ 832

Author(s):

Rohit Mahajan ◽

Christian Delphin ◽

Tinglu Guan ◽

Larry Gerace ◽

Frauke Melchior

Keyword(s):

Nuclear Pore Complex ◽

Nuclear Pore ◽

Complex Protein

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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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