scholarly journals Mutation or deletion of the Saccharomyces cerevisiae RAT3/NUP133 gene causes temperature-dependent nuclear accumulation of poly(A)+ RNA and constitutive clustering of nuclear pore complexes.

1995 ◽  
Vol 6 (4) ◽  
pp. 401-417 ◽  
Author(s):  
O Li ◽  
C V Heath ◽  
D C Amberg ◽  
T C Dockendorff ◽  
C S Copeland ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Messenger Rna ◽  
Nuclear Periphery ◽  
Nuclear Accumulation ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Permissive Temperature ◽  
Temperature Dependent ◽  
Pore Complexes ◽  
Mutant Cells

To identify genes whose products play potential roles in the nucleocytoplasmic export of messenger RNA, we isolated temperature-sensitive strains of Saccharomyces cerevisiae and examined them by fluorescent in situ hybridization. With the use of a digoxigen-tagged oligo-(dT)50 probe, we identified those that showed nuclear accumulation of poly(A)+ RNA when cells were shifted to the nonpermissive temperature. We describe here the properties of yeast strains bearing the rat3-1 mutation (RAT-ribonucleic acid trafficking) and the cloning of the RAT3 gene. When cultured at the permissive temperature of 23 degrees C, fewer than 10% of cells carrying the rat3-1 allele showed nuclear accumulation of poly(A)+ RNA, whereas approximately 70% showed nuclear accumulation of poly(A)+ RNA, whereas approximately 70% showed nuclear accumulation of poly(A)+ RNA after a shift to 37 degrees C for 4 h. In wild-type cells, nuclear pore complexes (NPCs) are distributed relatively evenly around the nuclear envelope. Both indirect immunofluorescence analysis and electron microscopy of rat3-1 cells indicated that NPCs were clustered into one or a few regions of the NE in mutant cells. Similar NPC clustering was seen in mutant cells cultured at temperatures between 15 degrees C and 37 degrees C. The RAT3 gene encodes an 1157-amino acid protein without similarity to other known proteins. It is essential for growth only at 37 degrees C. Cells carrying a disruption of the RAT3 gene were very similar to cells carrying the original rat3-1 mutation; they showed temperature-dependent nuclear accumulation of poly(A)+ RNA and exhibited constitutive clustering of NPCs. Epitope tagging of Rat3p demonstrated that it is located at the nuclear periphery and co-localizes with nuclear pore proteins recognized by the RL1 monoclonal antibody. We refer to this nucleoporin as Rat3p/Nup133p.

scholarly journals Nuclear pore complex clustering and nuclear accumulation of poly(A)+ RNA associated with mutation of the Saccharomyces cerevisiae RAT2/NUP120 gene.

1995 ◽  
Vol 131 (6) ◽  
pp. 1677-1697 ◽  
Author(s):  
C V Heath ◽  
C S Copeland ◽  
D C Amberg ◽  
V Del Priore ◽  
M Snyder ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nuclear Envelope ◽  
Messenger Rna ◽  
Nuclear Periphery ◽  
Nuclear Accumulation ◽  
Growth Defect ◽  
Nuclear Pore ◽  
Permissive Temperature ◽  
Mutant Cells ◽  
In Cells

To identify genes involved in the export of messenger RNA from the nucleus to the cytoplasm, we used an in situ hybridization assay to screen temperature-sensitive strains of Saccharomyces cerevisiae. This identified those which accumulated poly(A)+ RNA in their nuclei when shifted to the non-permissive temperature of 37 degrees C. We describe here the properties of yeast strains carrying mutations in the RAT2 gene (RAT - ribonucleic acid trafficking) and the cloning of the RAT2 gene. Only a low percentage of cells carrying the rat2-1 allele showed nuclear accumulation of poly(A)+ RNA when cultured at 15 degrees or 23 degrees C, but within 4 h of a shift to the nonpermissive temperature of 37 degrees C, poly(A)+ RNA accumulated within the nuclei of approximately 80% of cells. No defect was seen in the nuclear import of a reporter protein bearing a nuclear localization signal. Nuclear pore complexes (NPCs) are distributed relatively evenly around the nuclear envelope in wild-type cells. In cells carrying either the rat2-1 or rat2-2 allele, NPCs were clustered together into one or a few regions of the nuclear envelope. This clustering was a constitutive property of mutant cells. NPCs remained clustered in crude nuclei isolated from mutant cells, indicating that these clusters are not able to redistribute around the nuclear envelope when nuclei are separated from cytoplasmic components. Electron microscopy revealed that these clusters were frequently found in a protuberance of the nuclear envelope and were often located close to the spindle pole body. The RAT2 gene encodes a 120-kD protein without similarity to other known proteins. It was essential for growth only at 37 degrees C, but the growth defect at high temperature could be suppressed by growth of mutant cells in the presence of high osmolarity media containing 1.0 M sorbitol or 0.9 M NaCl. The phenotypes seen in cells carrying a disruption of the RAT2 gene were very similar to those seen with the rat2-1 and rat2-2 alleles. Epitope tagging was used to show that Rat2p is located at the nuclear periphery and co-localizes with yeast NPC proteins recognized by the RL1 monoclonal antibody. The rat2-1 allele was synthetically lethal with both the rat3-1/nup133-1 and rat7-1/nup159-1 alleles. These results indicate that the product of this gene is a nucleoporin which we refer to as Rat2p/Nup120p.

scholarly journals Saccharomyces cerevisiae Ndc1p Is a Shared Component of Nuclear Pore Complexes and Spindle Pole Bodies

1998 ◽  
Vol 143 (7) ◽  
pp. 1789-1800 ◽  
Author(s):  
Heidi J. Chial ◽  
Michael P. Rout ◽  
Thomas H. Giddings ◽  
Mark Winey
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Immunofluorescence Microscopy ◽  
Spindle Pole ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Indirect Immunofluorescence Microscopy ◽  
Spindle Pole Bodies ◽  
Shared Component ◽  
Pore Complexes ◽  
Mutant Cells

We report a novel connection between nuclear pore complexes (NPCs) and spindle pole bodies (SPBs) revealed by our studies of the Saccharomyces cerevisiae NDC1 gene. Although both NPCs and SPBs are embedded in the nuclear envelope (NE) in yeast, their known functions are quite distinct. Previous work demonstrated that NDC1 function is required for proper SPB duplication (Winey, M., M.A. Hoyt, C. Chan, L. Goetsch, D. Botstein, and B. Byers. 1993. J. Cell Biol. 122:743–751). Here, we show that Ndc1p is a membrane protein of the NE that localizes to both NPCs and SPBs. Indirect immunofluorescence microscopy shows that Ndc1p displays punctate, nuclear peripheral localization that colocalizes with a known NPC component, Nup49p. Additionally, distinct spots of Ndc1p localization colocalize with a known SPB component, Spc42p. Immunoelectron microscopy shows that Ndc1p localizes to the regions of NPCs and SPBs that interact with the NE. The NPCs in ndc1-1 mutant cells appear to function normally at the nonpermissive temperature. Finally, we have found that a deletion of POM152, which encodes an abundant but nonessential nucleoporin, suppresses the SPB duplication defect associated with a mutation in the NDC1 gene. We show that Ndc1p is a shared component of NPCs and SPBs and propose a shared function in the assembly of these organelles into the NE.

scholarly journals A conditional allele of the novel repeat-containing yeast nucleoporin RAT7/NUP159 causes both rapid cessation of mRNA export and reversible clustering of nuclear pore complexes.

1995 ◽  
Vol 129 (4) ◽  
pp. 939-955 ◽  
Author(s):  
L C Gorsch ◽  
T C Dockendorff ◽  
C N Cole
Keyword(s):  
Messenger Rna ◽  
Mrna Export ◽  
Temperature Shift ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Temperature Sensitive ◽  
Nuclear Pore Complexes ◽  
Reporter Protein ◽  
Direct Role ◽  
Pore Complexes

In a screen for Saccharomyces cerevisiae genes required for nucleocytoplasmic transport of messenger RNA, we identified the RAT7 gene (ribonucleic acid trafficking), which encodes an essential protein of 1,460 amino acids. Rat7p is located at the nuclear rim in a punctate pattern characteristic of nucleoporins. Furthermore, the central third of Rat7p contains 22 XXFG and three XFXFG degenerate repeats that are similar to signature GLFG and XFXFG repeats present in a majority of yeast and some mammalian nucleoporins sequenced to date. Shift of a strain bearing the temperature-sensitive rat7-1 allele from 23 degrees C to 37 degrees C resulted in rapid (within 15 minutes) cessation of mRNA export, but did not cause concomitant cytoplasmic accumulation of a reporter protein bearing a nuclear localization signal. This suggests that Rat7p may play a direct role in nucleocytoplasmic export of RNA. Immunofluorescence and thin section electron microscopy revealed that in rat7-1 cells grown at 23 degrees C, the majority of nuclear pore complexes (NPCs) were clustered on one side of the nucleus. No ultrastructural abnormalities of the nuclear envelope were seen. Interestingly, shifting rat7-1 cells to 37 degrees C for 1 h caused the NPCs to disperse, restoring near wild-type NPC distribution. After this temperature shift, the mutant Rat7p was no longer detectable by immunofluorescence.

scholarly journals The dynamic three-dimensional organization of the diploid yeast genome

2016 ◽  
Author(s):  
Seungsoo Kim ◽  
Ivan Liachko ◽  
Donna G Brickner ◽  
Kate Cook ◽  
William S Noble ◽  
...  
Keyword(s):  
Nuclear Periphery ◽  
Three Dimensional ◽  
Culture Conditions ◽  
Yeast Genome ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Chromosome Conformation ◽  
Homolog Pairing ◽  
Pore Complexes ◽  
Eukaryotic Genomes

AbstractThe budding yeast Saccharomyces cerevisiae is a long-standing model for the three-dimensional organization of eukaryotic genomes. Even in this well-studied model, it is unclear how homolog pairing in diploids and environment-induced gene relocalization influence overall genome organization. Here, we performed high-throughput chromosome conformation capture on diverged Saccharomyces hybrid diploids to obtain the first global view of chromosome conformation in diploid yeasts. After controlling for the Rabl-like orientation, we observe significant homolog proximity that increased in saturated culture conditions. Surprisingly, we observe a localized increase in homologous interactions between the HAS1 alleles specifically under galactose induction and saturated growth, mediated by association with nuclear pore complexes at the nuclear periphery. Together, these results reveal that the diploid yeast genome has a dynamic and complex 3D organization.

Relationships at the nuclear envelope: lamins and nuclear pore complexes in animals and plants

2010 ◽  
Vol 38 (3) ◽  
pp. 829-831 ◽  
Author(s):  
Jindriska Fiserova ◽  
Martin W. Goldberg
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Periphery ◽  
Nuclear Lamina ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Nucleocytoplasmic Trafficking ◽  
Animal Cells ◽  
Cellular Processes ◽  
Associated Proteins ◽  
Pore Complexes

The nuclear envelope comprises a distinct compartment at the nuclear periphery that provides a platform for communication between the nucleus and cytoplasm. Signal transfer can proceed by multiple means. Primarily, this is by nucleocytoplasmic trafficking facilitated by NPCs (nuclear pore complexes). Recently, it has been indicated that signals can be transmitted from the cytoskeleton to the intranuclear structures via interlinking transmembrane proteins. In animal cells, the nuclear lamina tightly underlies the inner nuclear membrane and thus represents the protein structure located at the furthest boundary of the nucleus. It enables communication between the nucleus and the cytoplasm via its interactions with chromatin-binding proteins, transmembrane and membrane-associated proteins. Of particular interest is the interaction of the nuclear lamina with NPCs. As both structures fulfil essential roles in close proximity at the nuclear periphery, their interactions have a large impact on cellular processes resulting in affects on tissue differentiation and development. The present review concentrates on the structural and functional lamina–NPC relationship in animal cells and its potential implications to plants.

Sequential recruitment of NPC proteins to the nuclear periphery at the end of mitosis

1999 ◽  
Vol 112 (13) ◽  
pp. 2253-2264 ◽  
Author(s):  
K. Bodoor ◽  
S. Shaikh ◽  
D. Salina ◽  
W.H. Raharjo ◽  
R. Bastos ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Time Course ◽  
Nuclear Periphery ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Confocal Immunofluorescence Microscopy ◽  
Pore Complexes ◽  
Nuclear Membrane Protein

Nuclear pore complexes (NPCs) are extremely elaborate structures that mediate the bidirectional movement of macromolecules between the nucleus and cytoplasm. With a mass of about 125 MDa, NPCs are thought to be composed of 50 or more distinct protein subunits, each present in multiple copies. During mitosis in higher cells the nuclear envelope is disassembled and its components, including NPC subunits, are dispersed throughout the mitotic cytoplasm. At the end of mitosis, all of these components are reutilized. Using both conventional and digital confocal immunofluorescence microscopy we have been able to define a time course of post-mitotic assembly for a group of NPC components (CAN/Nup214, Nup153, POM121, p62 and Tpr) relative to the integral nuclear membrane protein LAP2 and the NPC membrane glycoprotein gp210. Nup153, a component of the nuclear basket, associates with chromatin towards the end of anaphase, in parallel with the inner nuclear membrane protein, LAP2. However, immunogold labeling suggests that the initial Nup153 chromatin association is membrane-independent. Assembly of the remaining proteins follows that of the nuclear membranes and occurs in the sequence POM121, p62, CAN/Nup214 and gp210/Tpr. Since p62 remains as a complex with three other NPC proteins (p58, 54, 45) during mitosis and CAN/Nup214 maintains a similar interaction with its partner, Nup84, the relative timing of assembly of these additional four proteins may also be inferred. These observations suggest that there is a sequential association of NPC proteins with chromosomes during nuclear envelope reformation and the recruitment of at least eight of these precedes that of gp210. These findings support a model in which it is POM121 rather than gp210 that defines initial membrane-associated NPC assembly intermediates.

Peripheral Localization and Regulation of the Murine β-Globin Locus.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4070-4070
Author(s):  
Tobias Ragoczy ◽  
Agnes Telling ◽  
Rachel Byron ◽  
M.A. Bender ◽  
Mark Groudine
Keyword(s):  
Gene Expression ◽  
Globin Gene ◽  
Nuclear Periphery ◽  
Embryonic Stem ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Erythroid Progenitor ◽  
Interphase Cell ◽  
Pore Complexes ◽  
Globin Gene Expression

Abstract The interphase cell nucleus is structurally and functionally compartmentalized, making the subnuclear position of genes an important determinant of their activity. During cellular differentiation, as tissue-specific expression programs unfold, dynamic reorganization of the nucleus positions sets of genes in active or repressive compartments. The nuclear periphery has emerged as an unusually complex compartment in this process. While it is marked by facultative heterochromatin and has been considered primarily as a repressive compartment, recent work suggests that active genes may also associate with the periphery. Moreover, the nuclear envelope is riddled with nuclear pore complexes, the gateways for molecular exchange between the cytoplasm and the nucleus, resulting in substantial traffic through this compartment. Using murine erythropoiesis as a model system, our recent three dimensional analysis of the positioning of the β-globin locus revealed that, when inactive in undifferentiated embryonic stem cells and erythroid progenitor cells, the locus is positioned predominantly at the nuclear periphery and appears to contact the lamina. This association is lost with progressing erythroid maturation, and the locus is repositioned towards the nuclear interior concomitant with increasing β-globin gene expression. Importantly, however, β-major globin expression begins at the nuclear periphery prior to relocalization, suggesting that associations of the locus at the periphery may shift from repressive to activating complexes. We are investigating the interactions of the β-globin locus with the nuclear periphery by two approaches: Using enhanced imaging permitted by Cryo-ImmunoFISH, we are analyzing the position of the locus relative to specific components of the nuclear lamina and nuclear pore complexes at distinct differentiation stages. In addition, we are probing the physical interactions of the locus with the periphery by biochemical means. To this end we are using ChIP-chip to identify lamina associated proteins binding the β-globin locus and to determine what sequence elements within the locus mediate these interactions. Ultimately these experiments will shed further light on mechanisms regulating β-globin gene expression during erythropoiesis and how stage-specific nuclear localization contributes to this process.

scholarly journals The FG-repeat asymmetry of the nuclear pore complex is dispensable for bulk nucleocytoplasmic transport in vivo

2004 ◽  
Vol 167 (4) ◽  
pp. 583-590 ◽  
Author(s):  
Bryan Zeitler ◽  
Karsten Weis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nuclear Pore Complex ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Nucleocytoplasmic Trafficking ◽  
Transport Pathways ◽  
Pore Complexes ◽  
Biased Distribution

Nucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore complexes (NPCs). Translocation through the NPC is exquisitely selective and is mediated by interactions between soluble transport carriers and insoluble NPC proteins that contain phenylalanine-glycine (FG) repeats. Although most FG nucleoporins (Nups) are organized symmetrically about the planar axis of the nuclear envelope, very few localize exclusively to one side of the NPC. We constructed Saccharomyces cerevisiae mutants with asymmetric FG repeats either deleted or swapped to generate NPCs with inverted FG asymmetry. The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1. Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups. These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

scholarly journals Chromatin targeting of nuclear pore proteins induces chromatin decondensation

2019 ◽  
Vol 218 (9) ◽  
pp. 2945-2961 ◽  
Author(s):  
Terra M. Kuhn ◽  
Pau Pascual-Garcia ◽  
Alejandro Gozalo ◽  
Shawn C. Little ◽  
Maya Capelson
Keyword(s):  
Target Gene ◽  
Nuclear Periphery ◽  
Nuclear Pore ◽  
Open Chromatin ◽  
Nuclear Pore Complexes ◽  
Chromatin Binding ◽  
Chromatin Decondensation ◽  
Chromatin Remodeling Complex ◽  
Biochemical Interaction ◽  
Pore Complexes

Nuclear pore complexes have emerged in recent years as chromatin-binding nuclear scaffolds, able to influence target gene expression. However, how nucleoporins (Nups) exert this control remains poorly understood. Here we show that ectopically tethering Drosophila Nups, especially Sec13, to chromatin is sufficient to induce chromatin decondensation. This decondensation is mediated through chromatin-remodeling complex PBAP, as PBAP is both robustly recruited by Sec13 and required for Sec13-induced decondensation. This phenomenon is not correlated with localization of the target locus to the nuclear periphery, but is correlated with robust recruitment of Nup Elys. Furthermore, we identified a biochemical interaction between endogenous Sec13 and Elys with PBAP, and a role for endogenous Elys in global as well as gene-specific chromatin decompaction. Together, these findings reveal a functional role and mechanism for specific nuclear pore components in promoting an open chromatin state.

Sign in / Sign up

Export Citation Format

Share Document