Purification of NSP1 reveals complex formation with ‘GLFG’ nucleoporins and a novel nuclear pore protein NIC96.

Grandi, Paola; Doye, Valérie; Hurt, Ed

doi:10.1002/j.1460-2075.1993.tb05975.x

Cited by 178 publications

(240 citation statements)

References 46 publications

(50 reference statements)

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“…A second group of yeast nucleoporins that includes Nup49p, NuplOOp, Nupll6p (Wente et al, 1992;Wimmer et al, 1992;Wente and Blobel, 1994), and Nupl45p (Fabre et al, 1994;Wente and Blobel, 1994) have repeats of GLFG in their N-terminal and central domains. Nic96p (Grandi et al, 1993), a major component of the yeast NPC by mass (Rout and Wente, 1994), lacks repeats, whereas Srplp contains eight degenerate repeats that are related to repeats seen in plakoglobin, ,B-catenin, and the Drosophila armadillo protein (Yano et al, 1992(Yano et al, , 1994. A tenth yeast protein, POM152p, is thought to be a membrane protein (Wozniak et al, 1994), is not essential, and is also a very abundant component of the yeast NPC (Rout and Blobel, 1993).…”

Section: Discussionmentioning

confidence: 99%

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.

Heath

Amberg

et al. 1995

MBoC

119

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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°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 after a shift to 37°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°C and 37°C. The RAT3 gene encodes an 1157-amino acid protein without similarity to other known proteins. It is essential for growth only at 37°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/Nupl33p.

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Section: Discussionmentioning

confidence: 99%

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.

Heath

Amberg

et al. 1995

MBoC

119

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“…Be cause the GARl-ProtA and ProtA-NOPl fusion proteins supported cell growth at wild-type levels (data not shown), we concluded that they are fully functional. As a control, a yeast strain expressing a protein A-tagged nuclear pore protein, NSPl (Grandi et al 1993), was used. Cell extracts were prepared from each strain, and snoRNAs associated with the tagged proteins were immunoprecipitated with IgG-Sepharose.…”

Section: Yeast Aca Snornas Are Complexed With Garl But Not With Fibhlmentioning

confidence: 99%

“…Yeast strains expressing NSPl (Grandi et al 1993) son et al 1987) was PCR-amplified using sequence-specific primers (oligonucleotides 11 and 12) that introduced BamHL restriction sites at both the 5' and 3' termini. The amplified fragment was cut by BamHl and inserted into the BamHl site of pJPG67 (Girard et al 1994).…”

Section: Immunoprecipita Tionmentioning

confidence: 99%

The family of box ACA small nucleolar RNAs is defined by an evolutionarily conserved secondary structure and ubiquitous sequence elements essential for RNA accumulation.

Ganot¹,

Caizergues‐Ferrer²,

Kiss³

1997

Genes Dev.

317

387

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Eukaryotic cells contain a large number of small nucleolar RNAs (snoRNAs).A major family of snoRNAs features a consensus ACA motif positioned 3 nucleotides from the 3' end of the RNA. In this study we have characterized nine novel human ACA snoRNAs (U64-U72). Structural probing of U64 RNA followed by systematic computer modeling of all known box ACA snoRNAs revealed that this class of snoRNAs is defined by a phylogenetically conserved secondary structure. The ACA snoRNAs fold into two hairpin structures connected by a single-stranded hinge region and followed by a short 3' tail. In eukaryotic cells, maturation of rRNAs takes place in the nucleolus. The 18S, 5.8S, and 25/28S rRNAs are syn thesized as a large precursor RNA (pre-rRNA) that car ries long external and internal spacer sequences. Shortly after transcription, many nucleotides in the coding re gions of pre-rRNA are methylated at the 2'-0-hydroxyl position and numerous U residues are converted into pseudouridines (Maden 1990;Eichler and Craig 1995). The covalently modified pre-rRNA is than processed into mature rRNAs through a series of endo-and exonucleolytic cleavages (Eichler and Craig 1995; Venema and Tollervey 1995;Sollner-Webb et al. 1996).The nucleolus contains a large number of snoRNAs. More than 80 distinct small nucleolar RNA (snoRNA) sequences have been identified in vertebrate and yeast

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“…Several nucleoporin subcomplexes have been identified (Grandi et al, 1993;Grandi et al, 1995a;Grandi et al, 1995b;Nehrbass et al, 1996;Zabel et al, 1996;Siniossoglou et al, 1996;Iovine et al, 1997;Bailer et al, 1998;Belgareh et al, 1998;Marelli et al, 1998;Lutzmann et al, 2002). Different nucleoporins have been localised to distinct substructures of both the vertebrate and yeast NPCs (reviewed in Stoffler et al, 1999;Doye and Hurt, 1997;Rout et al, 2000).…”

Section: Introductionmentioning

confidence: 99%

Yeast nuclear pore complexes have a cytoplasmic ring and internal filaments

Киселева

Allen

Rutherford

et al. 2004

Journal of Structural Biology

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The full-text may be used and/or reproduced, and given to third parties in any format or medium, without prior permission or charge, for personal research or study, educational, or not-for-pro t purposes provided that:• a full bibliographic reference is made to the original source • a link is made to the metadata record in DRO • the full-text is not changed in any way The full-text must not be sold in any format or medium without the formal permission of the copyright holders.Please consult the full DRO policy for further details. AbstractThe nuclear pore complex (NPC) controls transport of macromolecules across the nuclear envelope. It is large and complex but appears to consist of only ~30 different proteins despite its mass of >60 MDa. Vertebrate NPC structure has been analyzed by several methods giving a comprehensive architectural model. Despite our knowledge of yeast nucleoporins, structural data is more limited and suggests the basic organization is similar to vertebrates, but may lack some peripheral and other components. Using field emission scanning electron microscopy to probe NPC structure we found that the yeast, like higher eukaryotic, NPCs contain similar peripheral components. We can detect cytoplasmic rings and evidence of nucleoplasmic rings in yeasts. A filamentous basket is present on the nucleoplasmic face and evidence for cytoplasmic filaments is shown. We observe a central structure, possibly the transporter, that which may be linked to the cytoplasmic ring by internal filaments. Immuno-gold labeling suggests that Nup159p may be attached to the cytoplasmic ring, whereas Nup116p may be associated, partly, with the cytoplasmic filaments. Analysis of a Nup57p mutant suggested a role in maintaining the stability of cytoplasmic components of the NPC. We conclude that peripheral NPC components appear similar in yeasts compared to higher organisms and present a revised model for yeast NPC structural composition.4

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Purification of NSP1 reveals complex formation with ‘GLFG’ nucleoporins and a novel nuclear pore protein NIC96.

Cited by 178 publications

References 46 publications

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.

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.

The family of box ACA small nucleolar RNAs is defined by an evolutionarily conserved secondary structure and ubiquitous sequence elements essential for RNA accumulation.

Yeast nuclear pore complexes have a cytoplasmic ring and internal filaments

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