Functional and dynamic aspects of the mammalian nucleolus

Scheer, Ulrich; Benavente, Ricardo

doi:10.1002/bies.950120104

Cited by 260 publications

(163 citation statements)

References 33 publications

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“…U3 and U14 are transiently associated with some 5Ј-ETS and 18S sequences of pre-rRNA (Beltrame and Tollervey, 1995;Liang and Fournier, 1995), and they have been suggested to assemble into a hypothetical multiple-snoRNP complex (Maxwell and Fournier, 1995;Ghisolfi-Nieto et al, 1996). In addition, electron-dense "terminal balls" at the leading ends of nascent pre-rRNA transcripts have been identified as the 5Ј-ETS primary processing complexes (Kass et al, 1990;Mougey et al, 1993a), which appear to contain U3 snoRNA (Kass et al, 1990), fibrillarin (Scheer and Benavente, 1990;Mougey et al, 1993b), and nucleolin (Ghisolfi-Nieto et al, 1996;Ginisty et al, 1998). Thus, the NDF may be a means of preserving the integrity of these processing complexes until the reformed postmitotic nucleoli become fully functional.…”

Section: Discussionmentioning

confidence: 99%

Partially Processed pre-rRNA Is Preserved in Association with Processing Components in Nucleolus-derived Foci during Mitosis

Dundr

Olson

1998

MBoC

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Previous studies showed that components implicated in pre-rRNA processing, including U3 small nucleolar (sno)RNA, fibrillarin, nucleolin, and proteins B23 and p52, accumulate in perichromosomal regions and in numerous mitotic cytoplasmic particles, termed nucleolus-derived foci (NDF) between early anaphase and late telophase. The latter structures were analyzed for the presence of pre-rRNA by fluorescence in situ hybridization using probes for segments of pre-rRNA with known half-lives. The NDF did not contain the short-lived 5Ј-external transcribed spacer (ETS) leader segment upstream from the primary processing site in 47S pre-rRNA. However, the NDF contained sequences from the 5Ј-ETS core, 18S, internal transcribed spacer 1 (ITS1), and 28S segments and also had detectable, but significantly reduced, levels of the 3Ј-ETS sequence. Northern analyses showed that in mitotic cells, the latter sequences were present predominantly in 45S-46S pre-rRNAs, indicating that high-molecular weight processing intermediates are preserved during mitosis. Two additional essential processing components were also found in the NDF: U8 snoRNA and hPop1 (a protein component of RNase MRP and RNase P). Thus, the NDF appear to be large complexes containing partially processed pre-rRNA associated with processing components in which processing has been significantly suppressed. The NDF may facilitate coordinated assembly of postmitotic nucleoli. INTRODUCTIONThe nucleolus is a prominent membraneless subnuclear compartment assembled around clusters of tandemly repeated rDNA genes from which prerRNA is transcribed and subsequently folded, processed, modified, and assembled into small and large ribosomal subunits (Scheer et al., 1993;Shaw and Jordan, 1995;Maden and Hughes, 1997). A remarkable aspect of the cell cycle is the disintegration of the nucleolus during mitosis and its reassembly as the daughter cells proceed toward G 1 phase. A pivotal event in this process is the repression of RNA polymerase (pol) I-driven pre-rRNA synthesis between prophase and telophase (Prescott and Bender, 1962). At the same time, nucleolar components disperse to various subcellular locations as the maternal nucleoli diassemble (Goessens, 1984).A comprehensive understanding of the locations and fates of nucleolar components during mitosis is slowly emerging (Scheer et al., 1993;Hernandez-Verdun and Gautier, 1994). For example, much of the RNA pol I transcription machinery and DNA topoisomerase I remain associated with the nucleolus organizer regions (NORs) 1 of chromosomes between nucleolar disassembly in late prophase and reassembly in telophase (Scheer et al., 1993; Weisenberger and * Corresponding author. 1 Abbrevations used: DFC, dense fibrillar component; ETS, external transcribed spacer; GC, granular component; ITS, internal transcribed spacer; NDF, nucleolus-derived foci; NOR, nucleolus organizer region; PNB, prenucleolar body; pol, polymerase; snoRNA, small nucleolar RNA; sno-RNP, small nucleolar ribonucleoprotein.© 1998 by The American Society for Ce...

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Section: Discussionmentioning

confidence: 99%

Partially Processed pre-rRNA Is Preserved in Association with Processing Components in Nucleolus-derived Foci during Mitosis

Dundr

Olson

1998

MBoC

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“…In this respect, as recently discussed by Scheer and Benavente (1990), both fibrillar nucleolar components might be considered as a functional unit whose structural integrity is essential for the transition of the primary transcripts from the template-bound to the released state. Subsequent steps of assembly and maturation then occur in the granular component before the ribosomal subunits are finally transported through the nuclear pores complexes into the cytoplasm.…”

Section: B Functional Organization Of the Nucleolusmentioning

confidence: 97%

“…a time span required for single RNA polymerase I molecule to transcribe half or even entirely through a rRNA gene unit. Thus it is conceivable that the distribution of silver grains primarily reflects the sites where the template-released pre-rRNA molecules accumulate to high local concentrations, thereby facilitating their detection by autoradiography (Goessens, 1976;Scheer and Benavente, 1990). Of course, the rRNA transcription units must also be labelled but the resulting silver grains in the fibrillar centers probably escaped detection so far because of a low signal to noise ratio.…”

Section: B Functional Organization Of the Nucleolusmentioning

confidence: 99%

“…Selective inhibition of rRNA transcription by microinjection of antibodies to RNA polymerase I into mitotic or interphasic mammalian cells have clearly shown that ongoing transcription is required for the integration of the dense fibrillar component into the nucleolus (Benavente et al, 1987(Benavente et al, , 1988. Furthermore, these experiments have demonstrated that the dense fibrillar component represents a structure sui generis which exists independent of the transcriptional apparatus (for review see Scheer and Benavente, 1990). Consequently, this nucleolar component cannot be considered as a transient structure formed by the superposition of active rRNA genes and their transcripts.…”

Section: B Functional Organization Of the Nucleolusmentioning

confidence: 99%

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Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

Thiry

Scheer

Goessens

1991

Electron Microscopy Reviews

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Abstract-Nucleoli are the morphological expression of the activity of a defined set of chromosomal segments bearing rRNA genes. The topological distribution and composition of the intranucleolar chromatin as well as the definition of nucleolar structures in which enzymes of the rDNA transcription machinery reside have been investigated in mammalian cells by various immunogold labelling approaches at the ultrastructural level. The precise intranucleolar location of rRNA genes has been further specified by electron microscopic in situ hybridization with a non-autoradiographic procedure.Our results indicate that the fibrillar centers are the sole nucleolar structures where rDNA, core histones, RNA polymerase I and DNA to po isomerase I are located together.Taking into account the potential value and limitations of immunoelectron microscopic techniques, we propose that transcription of the rRNA genes takes place within the confines of the fibrillar centers, probably close to the boundary regions to the surrounding dense fibrillar component.

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“…The appeal of this model was that it explained the rapid reinitiation rate on ribosomal gene promoters evidenced by the high density of polymerase molecules observed on active genes (Miller and Bakken 1972;Puvion-Dutilleul 1983;Scheer and Benavente 1990). Recently, however, this view of transcription has been challenged.…”

Section: Org Downloaded Frommentioning

confidence: 99%

UBF-binding site arrays form pseudo-NORs and sequester the RNA polymerase I transcription machinery

Mais¹,

Wright²,

Prieto³

et al. 2004

Genes Dev.

164

191

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Human ribosomal genes (rDNA) are located in nucleolar organizer regions (NORs) on the short arms of acrocentric chromosomes. Metaphase NORs that were transcriptionally active in the previous cell cycle appear as prominent chromosomal features termed secondary constrictions that are achromatic in chromosome banding and positive in silver staining. The architectural RNA polymerase I (pol I) transcription factor UBF binds extensively across rDNA throughout the cell cycle. To determine if UBF binding underpins NOR structure, we integrated large arrays of heterologous UBF-binding sequences at ectopic sites on human chromosomes. These arrays efficiently recruit UBF even to sites outside the nucleolus and, during metaphase, form novel silver stainable secondary constrictions, termed pseudo-NORs, morphologically similar to NORs. We demonstrate for the first time that in addition to UBF the other components of the pol I machinery are found associated with sequences across the entire human rDNA repeat. Remarkably, a significant fraction of these same pol I factors are sequestered by pseudo-NORs independent of both transcription and nucleoli. Because of the heterologous nature of the sequence employed, we infer that sequestration is mediated primarily by protein-protein interactions with UBF. These results suggest that extensive binding of UBF is responsible for formation and maintenance of the secondary constriction at active NORs. Furthermore, we propose that UBF mediates recruitment of the pol I machinery to nucleoli independently of promoter elements.[Keywords: RNA polymerase I; ribosomal DNA; nucleolar organiser region; nucleolus; upstream-binding factor] Supplemental material is available at http://www.genesdev.org.

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Functional and dynamic aspects of the mammalian nucleolus

Cited by 260 publications

References 33 publications

Partially Processed pre-rRNA Is Preserved in Association with Processing Components in Nucleolus-derived Foci during Mitosis

Partially Processed pre-rRNA Is Preserved in Association with Processing Components in Nucleolus-derived Foci during Mitosis

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

UBF-binding site arrays form pseudo-NORs and sequester the RNA polymerase I transcription machinery

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