Distribution of proteins among chromatin components of nucleoli

Olson, Matthew S.; Thompson, Barbara A.

doi:10.1021/bi00282a023

Cited by 86 publications

(19 citation statements)

References 38 publications

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“…Therefore, our results argue rather that the pol I promoter contains all of the information required for the repression by nucleolin. It has been shown that nucleolin binds tightly to chromatin (42,43), and to DNA fragments containing the region of the nontranscribed rDNA spacer upstream of the initiation site (19). Furthermore, nucleolin is able to interact with histone H1 and to modulate chromatin structure (20,21) suggesting that nucleolin could mediate repression of transcription through an interaction with the chromosomal intergenic spacer.…”

Section: Discussionmentioning

confidence: 99%

Repression of RNA Polymerase I Transcription by Nucleolin Is Independent of the RNA Sequence That Is Transcribed

Roger¹,

Moisand²,

Amalríc³

et al. 2002

Journal of Biological Chemistry

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Nucleolin is one of the most abundant non-ribosomal proteins of the nucleolus. Several studies in vitro have shown that nucleolin is involved in several steps of ribosome biogenesis, including the regulation of rDNA transcription, rRNA processing, and ribosome assembly. However, the different steps of ribosome biogenesis are highly coordinated, and therefore it is not clear to what extent nucleolin is involved in each of these steps. It has been proposed that the interaction of nucleolin with the rDNA sequence and with nascent pre-rRNA leads to the blocking of RNA polymerase I (RNA pol I) transcription. To test this model and to get molecular insights into the role of nucleolin in RNA pol I transcription, we studied the function of nucleolin in Xenopus oocytes. We show that injection of a 2-4-fold excess of Xenopus or hamster nucleolin in stage VI Xenopus oocytes reduces the accumulation of 40 S pre-rRNA 3-fold, whereas transcription by RNA polymerase II and III is not affected. Direct analysis of rDNA transcription units by electron microscopy reveals that the number of polymerase complexes/ rDNA unit is drastically reduced in the presence of increased amounts of nucleolin and corresponds to the level of reduction of 40 S pre-rRNA. Transcription from DNA templates containing various combinations of RNA polymerase I or II promoters in fusion with rDNA or CAT sequences was analyzed in the presence of elevated amounts of nucleolin. It was shown that nucleolin leads to transcription repression from a minimal polymerase I promoter, independently of the nature of the RNA sequence that is transcribed. Therefore, we propose that nucleolin affects RNA pol I transcription by acting directly on the transcription machinery or on the rDNA promoter sequences and not, as previously thought, through interaction with the nascent pre-rRNA.The synthesis of functional ribosomes is a major task for the cell. Ribosomal gene transcription can account for as much as 40% of all cellular transcription and ribosomal RNA for about 80% of the RNA content of living cells (1). The different steps of ribosome biogenesis take place in a subcompartment of the nucleus called the nucleolus (2-4). The localization of the different steps of ribosome biogenesis in a single nuclear compartment probably allows an efficient coordination and regulation of ribosome assembly. The formation of mature ribosomes is one of the most complex assembly of ribonucleoparticles involving the interaction of four different RNAs and about 80 ribosomal proteins (5). In addition, several nucleolar non-ribosomal proteins are required for this process (6 -8). An ordered interaction of ribosomal and non-ribosomal proteins with pre-rRNA is probably required for the formation of functional ribosomes. The molecular details of this highly integrated process are still largely unknown.The non-ribosomal proteins fibrillarin and nucleolin as well as some ribosomal proteins have been detected on nascent prerRNA (9 -11) suggesting that they interact with the pre-rRNA during transcrip...

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Section: Discussionmentioning

confidence: 99%

Repression of RNA Polymerase I Transcription by Nucleolin Is Independent of the RNA Sequence That Is Transcribed

Roger¹,

Moisand²,

Amalríc³

et al. 2002

Journal of Biological Chemistry

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“…It is also interesting that yeast topoisomerase II, which is implicated in chromosome condensation, contains a Cdc2 phosphorylation consensus sequence (50,51) and is readily phosphorylated in mitosis (26). A subclass of nucleolin is associated with chromosomal NORs in metaphase and with chromatin in interphase (19,20,40). Furthermore, nucleolin reveals structural analogies with histone Hi around Cdc2-phosphorylated threonine in the TPXK motif.…”

Section: Discussionmentioning

confidence: 99%

“…These proteins are candidates for controlling the switch-off of rRNA synthesis and structural reorganization of the nucleolus following mitosis. Among these, nucleolin (20) is a multifunctional protein (37) that is involved in nucleolar chromatin organization (19,40) and in pre-rRNA packaging (10, 27).The idea that nucleolin function is coupled to growth control by phosphorylation is supported by the observation that active rRNA transcription is correlated to highly phosphorylated nucleolin (22,44,48,49). In growing cells, a casein kinase II (CKII) phosphorylates nucleolin on serine during interphase, while in confluent cells, nucleolin is unphosphorylated and rRNA synthesis goes down to 5% of the growing level (4, 12).…”

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confidence: 99%

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Mitosis-specific phosphorylation of nucleolin by p34cdc2 protein kinase.

Bélenguer¹,

Caizergues‐Ferrer²,

Labbé³

et al. 1990

Mol. Cell. Biol.

145

100

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Nucleolin is a ubiquitous multifunctional protein involved in preribosome assembly and associated with both nucleolar chromatin in interphase and nucleolar organizer regions on metaphasic chromosomes in mitosis. Extensive nucleolin phosphorylation by a casein kinase (CKII) occurs on serine in growing cells. Here we report that while CKII phosphorylation is achieved in interphase, threonine phosphorylation occurs during mitosis. We provide evidence that this type of in vivo phosphorylation involves a mammalian homolog of the cell cycle control Cdc2 kinase. In vitro M-phase Hl kinase from starfish oocytes phosphorylated threonines in a TPXK motif present nine times in the amino-terminal part of the protein. The same sites which matched the p34Cdc2 consensus phosphorylation sequence were used in vivo during mitosis. We propose that successive Cdc2 and CKII phosphorylation could modulate nucleolin function in controlling cell cycle-dependent nucleolar function and organization. Our results, along with previous studies, suggest that while serine phosphorylation is related to nucleolin function in the control of rDNA transcription, threonine phosphorylation is linked to mitotic reorganization of nucleolar chromatin.Ribosome biogenesis in eucaryotic cells takes place within the nucleolus (see reference 24 for a review). This fundamental process is directly related to cell growth and modulated during progression through the cell cycle. Fluctuations in ribosomal particle formation are linked to changes in the structural organization of the nucleolus. During interphase, a functional structure in which rDNA transcription occurs is established by the contribution of DNA bearing the ribosomal genes (known as the nucleolar organizer regions [NORs]) from several chromosomes. During prophase, chromosome condensation is accompanied by the disaggregation of the nucleolus. A subset of nucleolar proteins remains associated with the chromosomal NORs (see reference 46 for a review). These proteins are candidates for controlling the switch-off of rRNA synthesis and structural reorganization of the nucleolus following mitosis. Among these, nucleolin (20) is a multifunctional protein (37) that is involved in nucleolar chromatin organization (19,40) and in pre-rRNA packaging (10, 27).The idea that nucleolin function is coupled to growth control by phosphorylation is supported by the observation that active rRNA transcription is correlated to highly phosphorylated nucleolin (22,44,48,49). In growing cells, a casein kinase II (CKII) phosphorylates nucleolin on serine during interphase, while in confluent cells, nucleolin is unphosphorylated and rRNA synthesis goes down to 5% of the growing level (4, 12). CKII phosphorylation sites have been mapped in the amino-terminal domain of the molecule next to long acidic domains. Similar stretches are also found in a class of anionic nuclear proteins (see reference 17 for review). A basic motif (TPXK) repeated between the first two acidic domains is reminiscent of sequences present in the amino-and ...

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“…Interestingly, in addition to its role in preribosome biogenesis, nucleolin is associated with both nucleolar chromatin in interphase [6,71 and nucleolar organiser regions in mitosis [8, 91. Indeed, we have shown that nucleolin can modulate chromatin condensation [7,10,111 and that this function correlates well with the observed homology between the 280-amino-acid N-terminal domain of nucleolin and high-mobility-group proteins.…”

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confidence: 99%

Concerted activities of the RNA recognition and the glycine‐rich C‐terminal domains of nucleolin are required for efficient complex formation with pre‐ribosomal RNA

Ghisolfi

Kharrat

Joseph

et al. 1992

European Journal of Biochemistry

114

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Nucleolin is an abundant nucleolar protein which is involved in the early stages of ribosome assembly. The central 40-kDa domain of nucleolin comprises four RNA recognition motifs (RRM) which are presumed to be involved in specific interactions with pre-rRNA. In order to examine in detail the role of this central domain and the contribution of the N-terminal and C-terminal domains of nucleolin to RNA binding, we have used an Escherichiu coli expression system to synthezise polypeptides corresponding to various combinations of the three domains and their subdomains. By means of an in-vitro binding assay and a synthetic RNA corresponding to a specific recognition site in pre-rRNA we have been able to demonstrate conclusively that the central 40-kDa domain is indeed responsible for the specificity of RNA recognition and that the N-terminal domain can be removed without affecting RNA binding. Most interestingly, it appears that the C-terminal 10-kDa domain, which is rich in glycine and arginine residues, is essential for efficient binding of nucleolin to RNA, but does not itself contribute to the specificity of the interaction. Circular dichroic spectroscopic probing of the RNA component shows that the C-terminal domain significantly modifies the RNAbinding properties of the central RRM core. Finally, infrared spectroscopic studies reveal that the central 40-kDa domain is structured in a helices and p sheets and that the interaction with the specific pre-rRNA site induces subtle changes in the p sheet conformation.Just as newly synthesized hnRNA associates in the nucleus with a discrete set of proteins, pre-rRNA is organised within the nucleolus into specific ribonucleoproteic complexes. One of the most abundant proteic factors to have been identified in the dense fibrillar component of the nucleolus is a 100-kDa phosphoprotein, called nucleolin [l, 21. Based on this finding and the observed association between nucleolin and prerRNA [3, 41, it was supposed that this nucleolar protein was implicated in the early stages of ribosoinal assembly. Recently, direct evidence of a molecular interaction between nucleolin and pre-rRNA has been obtained. Several sites of interaction have been mapped within the 18s and 28s ribosomal RNA sequences as well as in the 5' external transcribed spacer (5'-ETS), and it has been shown that this last site can be specifically recognised by nucleolin in an in-vitro system IS].Interestingly, in addition to its role in preribosome biogenesis, nucleolin is associated with both nucleolar chromatin in interphase [6, 71 and nucleolar organiser regions in mitosis [8, 91. Indeed, we have shown that nucleolin can modulate chromatin condensation [7,10, 111 and that this function correlates well with the observed homology between the 280-amino-acid N-terminal domain of nucleolin and high-mobility-group proteins. The remaining 430 amino acids of the protein sequence comprise two other domains: a central do- main, made up of four closely related 80 -90-amino-acid repeats and a C-terminal domain, 85 re...

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Distribution of proteins among chromatin components of nucleoli

Cited by 86 publications

References 38 publications

Repression of RNA Polymerase I Transcription by Nucleolin Is Independent of the RNA Sequence That Is Transcribed

Repression of RNA Polymerase I Transcription by Nucleolin Is Independent of the RNA Sequence That Is Transcribed

Mitosis-specific phosphorylation of nucleolin by p34cdc2 protein kinase.

Concerted activities of the RNA recognition and the glycine‐rich C‐terminal domains of nucleolin are required for efficient complex formation with pre‐ribosomal RNA

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