Assembly and Functional Organization of the Nucleolus: Ultrastructural Analysis of<i>Saccharomyces cerevisiae</i>Mutants

Trumtel, Stéphanie; Léger-Silvestre, Isabelle; Gleizes, Pierre‐Emmanuel; Teulières, Frédéric; Gas, Nicole

doi:10.1091/mbc.11.6.2175

Cited by 62 publications

(41 citation statements)

References 39 publications

(54 reference statements)

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“…In this strain, most of the chromosomal rDNA copies have been deleted and this lethal mutation is rescued by the introduction of a multicopy plasmid carrying a full rDNA unit controlled by its regular RNA polymerase I promoter (Oakes et al 1998). Strains such as NOY758 have been shown to display a few (one to three) mini nucleoli that are believed to be organized around several gathered plasmids from which high amounts of primary ribosomal RNA transcripts emanate (Oakes et al 1998;Trumtel et al 2000). If Rnt1p accumulates at the sites of active rDNA transcription driven from these plasmids, we expected to observe multiple dots of Rnt1p-GFP.…”

Section: Rnt1pgfp Strongly Accumulates Within a Nucleolar Structure Dmentioning

confidence: 99%

A cotranscriptional model for 3′-end processing of the Saccharomyces cerevisiae pre-ribosomal RNA precursor

Henras¹,

Bertrand²,

Chanfreau³

2004

RNA

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Cleavage of the Saccharomyces cerevisiae primary ribosomal RNA (rRNA) transcript in the 3 external transcribed spacer (ETS) by Rnt1p generates the 35S pre-rRNA, the earliest detectable species in the pre-rRNA processing pathway. In this study we show that Rnt1p is concentrated in a subnucleolar dot-shaped territory distinct from the nucleolar body. The 35S pre-rRNA is localized at the periphery of the Rnt1p dot, in a pattern that suggests a diffusion of the 35S pre-rRNA from the site of Rnt1p processing. When plasmid-borne versions of the rDNA are used to express rRNAs, the Rnt1p territory reorganizes around these plasmids, suggesting a close association between Rnt1p and the plasmid-borne rDNA units. Rnt1p was found associated with the endogenous rDNA by chromatin immunoprecipitation. Deletion of functionally important Rnt1p domains result in a loss of the dot-shaped territory, showing that this subnucleolar territory corresponds to a functional site of processing. These results show that a large fraction of Rnt1p is localized at the site of transcription of the rDNA, suggesting that the cleavage of the primary pre-rRNA transcript to generate the 35S pre-rRNA is a cotranscriptional event.

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Section: Rnt1pgfp Strongly Accumulates Within a Nucleolar Structure Dmentioning

confidence: 99%

A cotranscriptional model for 3′-end processing of the Saccharomyces cerevisiae pre-ribosomal RNA precursor

Henras¹,

Bertrand²,

Chanfreau³

2004

RNA

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“…Intriguingly, different yeast strains have different numbers of tandem repeats, extending from 100 to 200 (Pasero and Marilley 1993). Assembly of the nucleolus is thought to be a selfdriven process, initiated by production of rRNA (Trumtel et al 2000;Hernandez-Verdun et al 2002). This argument stems in part from studies in which the rDNA repeat was transcribed by RNA Pol II, rather than by the endogenous RNA Pol I.…”

Section: Dna-based Compartmentsmentioning

confidence: 99%

“…This argument stems in part from studies in which the rDNA repeat was transcribed by RNA Pol II, rather than by the endogenous RNA Pol I. This led to massive alterations in nucleolar structure, arguing that factors associated with RNA Pol I play a key role in proper nucleolar assembly (Oakes et al 1993;Trumtel et al 2000).…”

Section: Dna-based Compartmentsmentioning

confidence: 99%

Structure and Function in the Budding Yeast Nucleus

2012

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Budding yeast, like other eukaryotes, carries its genetic information on chromosomes that are sequestered from other cellular constituents by a double membrane, which forms the nucleus. An elaborate molecular machinery forms large pores that span the double membrane and regulate the traffic of macromolecules into and out of the nucleus. In multicellular eukaryotes, an intermediate filament meshwork formed of lamin proteins bridges from pore to pore and helps the nucleus reform after mitosis. Yeast, however, lacks lamins, and the nuclear envelope is not disrupted during yeast mitosis. The mitotic spindle nucleates from the nucleoplasmic face of the spindle pole body, which is embedded in the nuclear envelope. Surprisingly, the kinetochores remain attached to short microtubules throughout interphase, influencing the position of centromeres in the interphase nucleus, and telomeres are found clustered in foci at the nuclear periphery. In addition to this chromosomal organization, the yeast nucleus is functionally compartmentalized to allow efficient gene expression, repression, RNA processing, genomic replication, and repair. The formation of functional subcompartments is achieved in the nucleus without intranuclear membranes and depends instead on sequence elements, protein-protein interactions, specific anchorage sites at the nuclear envelope or at pores, and long-range contacts between specific chromosomal loci, such as telomeres. Here we review the spatial organization of the budding yeast nucleus, the proteins involved in forming nuclear subcompartments, and evidence suggesting that the spatial organization of the nucleus is important for nuclear function. TABLE OF CONTENTS Abstract 107Introduction 108

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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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Assembly and Functional Organization of the Nucleolus: Ultrastructural Analysis ofSaccharomyces cerevisiaeMutants

Cited by 62 publications

References 39 publications

A cotranscriptional model for 3′-end processing of the Saccharomyces cerevisiae pre-ribosomal RNA precursor

A cotranscriptional model for 3′-end processing of the Saccharomyces cerevisiae pre-ribosomal RNA precursor

Structure and Function in the Budding Yeast Nucleus

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

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