Dynamics of Nuclear Pore Distribution in Nucleoporin Mutant Yeast Cells

Belgareh, Naïma; Doye, Valérie

doi:10.1083/jcb.136.4.747

Cited by 133 publications

(112 citation statements)

References 63 publications

(121 reference statements)

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“…pASZ11-ADE2-PUS1-GFP, encoding PUS1-GFP (Hellmuth et al, 1998), and pASZ11-ADE2-NUP49-GFP, encoding NUP49-GFP (Belgareh and Doye, 1997), were gifts from Ed Hurt (University of Heidelberg, Heidelberg, Germany). These plasmids were modified by PCR-mediated replacement of the ADE2 gene with the URA3 gene to yield plasmids pPUS1-URA and pNUP49-URA, respectively.…”

Section: Strains and Plasmidsmentioning

confidence: 99%

Yeast Nuclear Envelope Subdomains with Distinct Abilities to Resist Membrane Expansion

Campbell

Lorenz

Witkin

et al. 2006

MBoC

117

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Little is known about what dictates the round shape of the yeast Saccharomyces cerevisiae nucleus. In spo7⌬ mutants, the nucleus is misshapen, exhibiting a single protrusion. The Spo7 protein is part of a phosphatase complex that represses phospholipid biosynthesis. Here, we report that the nuclear protrusion of spo7⌬ mutants colocalizes with the nucleolus, whereas the nuclear compartment containing the bulk of the DNA is unaffected. Using strains in which the nucleolus is not intimately associated with the nuclear envelope, we show that the single nuclear protrusion of spo7⌬ mutants is not a result of nucleolar expansion, but rather a property of the nuclear membrane. We found that in spo7⌬ mutants the peripheral endoplasmic reticulum (ER) membrane was also expanded. Because the nuclear membrane and the ER are contiguous, this finding indicates that in spo7⌬ mutants all ER membranes, with the exception of the membrane surrounding the bulk of the DNA, undergo expansion. Our results suggest that the nuclear envelope has distinct domains that differ in their ability to resist membrane expansion in response to increased phospholipid biosynthesis. We further propose that in budding yeast there is a mechanism, or structure, that restricts nuclear membrane expansion around the bulk of the DNA. INTRODUCTIONThe nucleus has a distinct organization, characterized by the presence of internal subcompartments. Moreover, in most, but not all, cell types the nucleus adopts a round shape. Understanding how this organization and shape are achieved is of major biological and medical interest. Certain cell types, such as those found in blood lineages (e.g., neutrophils, monocytes, and eosinophils), undergo dramatic nuclear shape changes as they differentiate (Gartner et al., 1990). Cancerous states are often associated with changes in nuclear morphology, most frequently nucleolar enlargement, changes in nuclear shape, or a combination of the two (Zink et al., 2004). Although these changes provide useful diagnostic markers of cancer progression, their mechanistic basis is still poorly understood. Other diseases are also associated with changes in nuclear shape and organization. For example, Hutchinson-Gilford progeria syndrome, which leads to premature aging, is caused by a specific mutation in the gene encoding A-type lamins and is associated with nuclear shape changes . Interestingly, progeria-like changes in nuclear shape are part of the normal aging process of nonneuronal cells in Caenorhabditis elegans (Haithcock et al., 2005). Genetic defects in the nuclear lamina are also associated with several types of muscular dystrophy . Nuclear lamins and their associated proteins provide both a rigid structure that helps shape the nuclear membrane and a platform onto which protein complexes and chromatin can bind (Holaska et al., 2002). Although much has been learned in recent years about the function of nuclear lamins, many significant questions remain. For example, it is not known how diseaseassociated changes in nuclear shape affect...

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Section: Strains and Plasmidsmentioning

confidence: 99%

Yeast Nuclear Envelope Subdomains with Distinct Abilities to Resist Membrane Expansion

Campbell

Lorenz

Witkin

et al. 2006

MBoC

117

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“…Transformed strains were grown in minimal YNB medium containing 0.67% of yeast nitrogen base supplemented with the appropriate amino acids and 2% glucose. The following expression plasmids were used in this study: Nop1p-green fluorescent protein (GFP) (Verheggen et al, 2001), Tgs1p-GFP , Nup49p-GFP (Belgareh and Doye, 1997), and Gar1p-GFP (Trumtel et al, 2000).…”

Section: Strains and Plasmidsmentioning

confidence: 99%

Inactivation of Cleavage Factor I Components Rna14p and Rna15p Induces Sequestration of Small Nucleolar Ribonucleoproteins at Discrete Sites in the Nucleus

Carneiro

Carvalho

Braga

et al. 2008

MBoC

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Small nucleolar RNAs (snoRNAs) associate with specific proteins forming small nucleolar ribonucleoprotein (snoRNP) particles, which are essential for ribosome biogenesis. The snoRNAs are transcribed, processed, and assembled in snoRNPs in the nucleoplasm. Mature particles are then transported to the nucleolus. In yeast, 3-end maturation of snoRNAs involves the activity of Rnt1p endonuclease and cleavage factor IA (CFIA). We report that after inhibition of CFIA components Rna14p and Rna15p, the snoRNP proteins Nop1p, Nop58p, and Gar1p delocalize from the nucleolus and accumulate in discrete nucleoplasmic foci. The U14 snoRNA, but not U3 snoRNA, similarly redistributes from the nucleolus to the nucleoplasmic foci. Simultaneous depletion of either Rna14p or Rna15p and the nuclear exosome component Rrp6p induces accumulation of poly(A)؉ RNA at the snoRNP-containing foci. We propose that the foci detected after CFIA inactivation correspond to quality control centers in the nucleoplasm. INTRODUCTIONSmall nucleolar RNAs (snoRNAs) are an abundant group of nonprotein coding RNAs in eukaryotic cells (Kiss, 2002). The snoRNAs associate with specific proteins forming small nucleolar ribonucleoprotein (snoRNP) particles, the majority of which participates in the biogenesis of ribosomes in the nucleolus (for recent reviews, see Bachellerie et al., 2002;Filipowicz and Pogacic, 2002;Kiss, 2002;Tran et al., 2004). The known snoRNAs fall into two major classes named box C/D and box H/ACA, which are characterized by distinctive sequence elements and associated proteins.The snoRNAs are transcribed as precursors that undergo posttranscriptional processing to generate the mature functional forms. Although most vertebrate snoRNAs are processed from the intronic regions of mRNA precursors Fournier, 1995, Bachellerie et al., 2002), only a few yeast snoRNAs are intron encoded. The majority of yeast snoRNAs are encoded in independent monocistronic or polycistronic transcripts that are transcribed by RNA polymerase II. Intronic snoRNAs can be produced either by splicing-dependent or by splicing-independent processing, whereas the maturation of independently transcribed units involves both endonucleolytic cleavage and exonucleolytic trimming (Filipowicz and Pogacic, 2002).In yeast, the U14 and U3 box C/D snoRNAs constitute paradigms for the study of snoRNA biosynthesis. U14 is cotranscribed with snR190, another box C/D snoRNA (Li et al., 1990). Cleavage of the dicistronic precursor by the endonuclease Rnt1 separates snR190 from U14 (Chanfreau et al., 1998). The pre-U3 transcript, by contrast, is produced from a single gene and contains the 7-methyl guanosine (m 7 G) cap structure, which is characteristic of mRNAs, and a short 3Ј extension. During maturation, the m 7 G cap on the pre-U3 is hypermethylated to 2,2,7-trimethylguanosine (TMG) and the 3Ј end is cleaved by Rnt1p, followed by removal of the 3Ј-terminal extension (Kufel et al., 2000). The yeast pre-U3 also contains an intron that is removed by the pre-mRNA splicing machinery, but t...

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“…The Nup84p complex consists of the nups Nup84p, Nup85p, Nup120p, Nup133p, Nup145-Cp, and Seh1p (Siniossoglou et al, 1996;Allen et al, 2002;Lutzmann et al, 2002). Mutations in members of the Nup84p complex display a variety of phenotypes including NPC clustering and NE organization defects (Aitchison et al, 1995a;Siniossoglou et al, 1996;Belgareh and Doye, 1997), suggesting a role for this complex in NPC assembly. Similarly, the vertebrate counterpart, the Nup107-160 complex, comprised of Nup107, Nup160, Nup133, Nup96, Nup85, Sec13, Nup43, Nup37, and Seh1 (Belgareh et al, 2001;Vasu et al, 2001;Harel et al, 2003;Loïodice et al, 2004) is also required for correct NPC assembly (Boehmer et al, 2003;Harel et al, 2003;Walther et al, 2003).…”

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confidence: 99%

Vertebrate Nup53 Interacts with the Nuclear Lamina and Is Required for the Assembly of a Nup93-containing Complex

Hawryluk-Gara

Shibuya

Wozniak

2005

MBoC

125

149

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The nuclear pore complex (NPC) is an evolutionarily conserved structure that mediates exchange of macromolecules across the nuclear envelope (NE). It is comprised of ϳ30 proteins termed nucleoporins that are each present in multiple copies. We have investigated the function of the human nucleoporin Nup53, the ortholog of Saccharomyces cerevisiae Nup53p. Both cell fractionation and in vitro binding data suggest that Nup53 is tightly associated with the NE membrane and the lamina where it interacts with lamin B. We have also shown that Nup53 is capable of physically interacting with a group of nucleoporins including Nup93, Nup155, and Nup205. Consistent with this observation, depletion of Nup53 using small interfering RNAs causes a decrease in the cellular levels of these nucleoporins as well as the spindle checkpoint protein Mad1, likely due to destabilization of Nup53-containing complexes. The cellular depletion of this group of nucleoporins, induced by depleting either Nup53 or Nup93, severely alters nuclear morphology producing phenotypes similar to that previously observed in cells depleted of lamin A and Mad1. On basis of these data, we propose a model in which Nup53 is positioned near the pore membrane and the lamina where it anchors an NPC subcomplex containing Nup93, Nup155, and Nup205. INTRODUCTIONThe nuclear envelope (NE) is a specialized membrane system that functions to separate the eukaryotic genome from the cytoplasm. The NE consists of an inner and an outer nuclear membrane. The former contains a distinct set of associated proteins, whereas the latter is structurally equivalent to the endoplasmic reticulum (reviewed in Mattaj, 2004). The nucleoplasmic face of the metazoan inner nuclear membrane is connected to a fibrous protein meshwork termed the nuclear lamina that forms a shell around the underlying chromatin mass. The lamina is composed of Aand B-type lamins, which are closely related to one another and to the intermediate filament-like family of proteins (reviewed in Burke, 2001). The nuclear lamina has been suggested to be involved in maintaining the structural integrity of the NE and influencing chromatin structure and function. The association of the lamina with transcriptionally inactive heterochromatin has led to the suggestion that it may play a role in regulating gene expression and some recent data support this idea (reviewed in Mattout-Drubezki and Gruenbaum, 2003).At numerous points along the NE the inner and outer nuclear membranes fuse to form pores ϳ100 nm in diameter. These pores are occupied by complex macromolecular structures termed nuclear pore complexes (NPCs). The NPCs are associated with euchromatin channels that extend into the interior of the nucleus, interrupting the continuity of the lamina and the heterochromatin. These complex structures have an estimated mass of ϳ60 million Daltons, but are composed of a relatively small number of proteins (ϳ30; Rout et al., 2000;Cronshaw et al., 2002) termed nucleoporins or nups. This is explained by the fact that all of these prot...

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Dynamics of Nuclear Pore Distribution in Nucleoporin Mutant Yeast Cells

Cited by 133 publications

References 63 publications

Yeast Nuclear Envelope Subdomains with Distinct Abilities to Resist Membrane Expansion

Yeast Nuclear Envelope Subdomains with Distinct Abilities to Resist Membrane Expansion

Inactivation of Cleavage Factor I Components Rna14p and Rna15p Induces Sequestration of Small Nucleolar Ribonucleoproteins at Discrete Sites in the Nucleus

Vertebrate Nup53 Interacts with the Nuclear Lamina and Is Required for the Assembly of a Nup93-containing Complex

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