hNOP2/NSUN1 Regulates Ribosome Biogenesis through Stabilization of snoRNP Complexes and Cytosine-5 Methylation of 28S rRNA.

5-Methylcytosine (m5C) is a base modification broadly found on a variety of RNAs in the human transcriptome. In eukaryotes m5C is catalyzed by enzymes of the NSUN family, which is composed of seven members in humans (NSUN1-7). NOP2/NSUN1 has been mostly characterized in budding yeast as an essential ribosome biogenesis factor required for the deposition of m5C on the 25S rRNA. Although human NOP2/NSUN1 has been known to be an oncogene overexpressed in several types of cancer, its functions remain poorly characterized. To define the roles of human NOP2/NSUN1, we used an miCLIP-seq approach to identify its RNA substrates. Our analysis reveals that vault RNA 1.2 and rRNA are NOP2/NSUN1-specific methylated targets and we further confirm by bisulfite sequencing that NOP2/NSUN1 is responsible for the deposition of m5C at residue 4447 on the 28S rRNA. Depletion of NOP2/NSUN1 impairs cell proliferation, rRNA processing and 60S ribosome biogenesis. Additionally, we find that NOP2/NSUN1 binds to the 5′ETS region of the pre-rRNA transcript and regulates pre-rRNA processing in part through non-catalytic complex formation with box C/D snoRNAs. Our study identifies for the first time the RNA substrates of human NOP2/NSUN1 and reveals additional functions in rRNA processing beyond catalyzing m5C base modification.

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Las1L Is a Nucleolar Protein Required for Cell Proliferation and Ribosome Biogenesis

Molecular and Cellular Biology ◽

10.1128/mcb.00358-10 ◽

2010 ◽

Vol 30 (18) ◽

pp. 4404-4414 ◽

Cited By ~ 35

Author(s):

Christopher D. Castle ◽

Erica K. Cassimere ◽

Jinho Lee ◽

Catherine Denicourt

Keyword(s):

Cell Proliferation ◽

Cell Growth ◽

Ribosome Biogenesis ◽

Ribosomal Subunit ◽

Nucleolar Protein ◽

28S Rrna ◽

Rrna Processing ◽

Tumor Suppressor P53 ◽

Growth Increase ◽

Multistep Process

ABSTRACT Ribosome biogenesis is a highly regulated process ensuring that cell growth (increase in biomass) is coordinated with cell proliferation. The formation of eukaryotic ribosomes is a multistep process initiated by the transcription and processing of rRNA in the nucleolus. Concomitant with this, several preribosomal particles, which transiently associate with numerous nonribosomal factors before mature 60S and 40S subunits are formed and exported in the cytoplasm, are generated. Here we identify Las1L as a previously uncharacterized nucleolar protein required for ribosome biogenesis. Depletion of Las1L causes inhibition of cell proliferation characterized by a G1 arrest dependent on the tumor suppressor p53. Moreover, we demonstrate that Las1L is crucial for ribosome biogenesis and that depletion of Las1L leads to inhibition of rRNA processing and failure to synthesize the mature 28S rRNA. Taken together, our data demonstrate that Las1L is essential for cell proliferation and biogenesis of the 60S ribosomal subunit.

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Small nucleolar RNA

Biochemistry and Cell Biology ◽

10.1139/o95-092 ◽

1995 ◽

Vol 73 (11-12) ◽

pp. 845-858 ◽

Cited By ~ 27

Author(s):

Susan A. Gerbi

Keyword(s):

Rna Polymerase Ii ◽

Binding Sites ◽

Ribosome Biogenesis ◽

Internal Transcribed Spacers ◽

28S Rrna ◽

Small Nucleolar Rnas ◽

Rrna Processing ◽

Conserved Sequence ◽

Nucleolar Rna ◽

Nucleolar Rnas

A growing list of small nucleolar RNAs (snoRNAs) has been characterized in eukaryotes. They are transcribed by RNA polymerase II or III; some snoRNAs are encoded in the introns of other genes. The nonintronic polymerase II transcribed snoRNAs receive a trimethylguanosine cap, probably in the nucleus, and move to the nucleolus. snoRNAs are complexed with proteins, sometimes including fibrillarin. Localization and maintenance in the nucleolus of some snoRNAs requires the presence of initial precursor rRNA (pre-rRNA). Many snoRNAs have conserved sequence boxes C and D and a 3′ terminal stem; the roles of these features are discussed. Functional assays done for a few snoRNAs indicate their roles in rRNA processing for cleavage of the external and internal transcribed spacers (ETS and ITS). U3 is the most abundant snoRNA and is needed for cleavage of ETS1 and ITS1; experimental results on U3 binding sites in pre-rRNA are reviewed. 18S rRNA production also needs U14, U22, and snR30 snoRNAs, whereas U8 snoRNA is needed for 5.8S and 28S rRNA production. Other snoRNAs that are complementary to 18S or 28S rRNA might act as chaperones to mediate RNA folding. Whether snoRNAs join together in a large rRNA processing complex (the "processome") is not yet clear. It has been hypothesized that such complexes could anchor the ends of loops in pre-rRNA containing 18S or 28S rRNA, thereby replacing base-paired stems found in pre-rRNA of prokaryotes.Key words: RNA processing, small nucleolar RNAs, nucleolus, ribosome biogenesis, rRNA processing complex.

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The sequence of the 5' end of the U8 small nucleolar RNA is critical for 5.8S and 28S rRNA maturation.

Molecular and Cellular Biology ◽

10.1128/mcb.17.7.3702 ◽

1997 ◽

Vol 17 (7) ◽

pp. 3702-3713 ◽

Cited By ~ 48

Author(s):

B A Peculis

Keyword(s):

Ribosome Biogenesis ◽

Critical Role ◽

Functional Site ◽

28S Rrna ◽

Rrna Processing ◽

Hybrid Molecules ◽

Evolutionarily Conserved ◽

Rrna Maturation ◽

Base Pairing Interaction

Ribosome biogenesis in eucaryotes involves many small nucleolar ribonucleoprotein particles (snoRNP), a few of which are essential for processing pre-rRNA. Previously, U8 snoRNA was shown to play a critical role in pre-rRNA processing, being essential for accumulation of mature 28S and 5.8S rRNAs. Here, evidence which identifies a functional site of interaction on the U8 RNA is presented. RNAs with mutations, insertions, or deletions within the 5'-most 15 nucleotides of U8 do not function in pre-rRNA processing. In vivo competitions in Xenopus oocytes with 2'O-methyl oligoribonucleotides have confirmed this region as a functional site of a base-pairing interaction. Cross-species hybrid molecules of U8 RNA show that this region of the U8 snoRNP is necessary for processing of pre-rRNA but not sufficient to direct efficient cleavage of the pre-rRNA substrate; the structure or proteins comprising, or recruited by, the U8 snoRNP modulate the efficiency of cleavage. Intriguingly, these 15 nucleotides have the potential to base pair with the 5' end of 28S rRNA in a region where, in the mature ribosome, the 5' end of 28S interacts with the 3' end of 5.8S. The 28S-5.8S interaction is evolutionarily conserved and critical for pre-rRNA processing in Xenopus laevis. Taken together these data strongly suggest that the 5' end of U8 RNA has the potential to bind pre-rRNA and in so doing, may regulate or alter the pre-rRNA folding pathway. The rest of the U8 particle may then facilitate cleavage or recruitment of other factors which are essential for pre-rRNA processing.

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Role of Pre-rRNA Base Pairing and 80S Complex Formation in Subnucleolar Localization of the U3 snoRNP

Molecular and Cellular Biology ◽

10.1128/mcb.24.19.8600-8610.2004 ◽

2004 ◽

Vol 24 (19) ◽

pp. 8600-8610 ◽

Cited By ~ 33

Author(s):

Sander Granneman ◽

Judith Vogelzangs ◽

Reinhard Lührmann ◽

Walther J. van Venrooij ◽

Ger J. M. Pruijn ◽

...

Keyword(s):

Complex Formation ◽

Ribosome Biogenesis ◽

Specific Protein ◽

Direct Result ◽

Rrna Processing ◽

Base Pairing ◽

Active Involvement ◽

U3 Snorna ◽

Fibrillar Component

ABSTRACT In the nucleolus the U3 snoRNA is recruited to the 80S pre-rRNA processing complex in the dense fibrillar component (DFC). The U3 snoRNA is found throughout the nucleolus and has been proposed to move with the preribosomes to the granular component (GC). In contrast, the localization of other RNAs, such as the U8 snoRNA, is restricted to the DFC. Here we show that the incorporation of the U3 snoRNA into the 80S processing complex is not dependent on pre-rRNA base pairing sequences but requires the B/C motif, a U3-specific protein-binding element. We also show that the binding of Mpp10 to the 80S U3 complex is dependent on sequences within the U3 snoRNA that base pair with the pre-rRNA adjacent to the initial cleavage site. Furthermore, mutations that inhibit 80S complex formation and/or the association of Mpp10 result in retention of the U3 snoRNA in the DFC. From this we propose that the GC localization of the U3 snoRNA is a direct result of its active involvement in the initial steps of ribosome biogenesis.

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Formation and Trapping of the Thermodynamically Unfavoured Inverted-Hemicucurbit[6]uril

10.26434/chemrxiv.7977566 ◽

2019 ◽

Author(s):

Elena Prigorchenko ◽

Sandra Kaabel ◽

Triin Narva ◽

Anastassia Baškir ◽

Maria Fomitšenko ◽

...

Keyword(s):

Complex Formation ◽

Combinatorial Chemistry ◽

Trifluoroacetic Acid ◽

Chloride Anion ◽

Binding Affinities ◽

Dynamic Covalent Chemistry ◽

Reaction Selectivity ◽

Low Concentration ◽

Dynamic Combinatorial Chemistry ◽

First Time

Amplification of a thermodynamically unfavoured macrocyclic product through the directed shift of the equilibrium between dynamic covalent chemistry library members is difficult to achieve. We show for the first time that during condensation of formaldehyde and cis-N,N'-cyclohexa-1,2-diylurea formation of inverted-cis-cyclohexanohemicucurbit[6]uril (i-cis-cycHC[6]) can be induced at the expense of thermodynamically favoured cis-cyclohexanohemicucurbit[6]uril (cis-cycHC[6]). The formation of i-cis-cycHC[6] is enhanced in low concentration of the templating chloride anion and suppressed in excess of this template. We found that reaction selectivity is governed by the solution-based template-aided dynamic combinatorial chemistry and continuous removal of the formed cycHC[6] macrocycles from the equilibrating solution by precipitation. Notably, the i-cis-cycHC[6] was isolated with 33% yield. Different binding affinities of three diastereomeric i-cis-, cis-cycHC[6] and their chiral isomer (R,R)-cycHC[6] for trifluoroacetic acid demonstrate the influence of macrocycle geometry on complex formation.

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Interactome Analysis of KIN (Kin17) Shows New Functions of This Protein

Current Issues in Molecular Biology ◽

10.3390/cimb43020056 ◽

2021 ◽

Vol 43 (2) ◽

pp. 767-781

Author(s):

Vanessa Pinatto Gaspar ◽

Anelise Cardoso Ramos ◽

Philippe Cloutier ◽

José Renato Pattaro Junior ◽

Francisco Ferreira Duarte Junior ◽

...

Keyword(s):

Dna Replication ◽

Protein Interactions ◽

Ribosome Biogenesis ◽

Cell Metabolism ◽

Cell Types ◽

Protein Protein Interactions ◽

Cellular Mechanisms ◽

Cancerous Cell ◽

First Time

KIN (Kin17) protein is overexpressed in a number of cancerous cell lines, and is therefore considered a possible cancer biomarker. It is a well-conserved protein across eukaryotes and is ubiquitously expressed in all cell types studied, suggesting an important role in the maintenance of basic cellular function which is yet to be well determined. Early studies on KIN suggested that this nuclear protein plays a role in cellular mechanisms such as DNA replication and/or repair; however, its association with chromatin depends on its methylation state. In order to provide a better understanding of the cellular role of this protein, we investigated its interactome by proximity-dependent biotin identification coupled to mass spectrometry (BioID-MS), used for identification of protein–protein interactions. Our analyses detected interaction with a novel set of proteins and reinforced previous observations linking KIN to factors involved in RNA processing, notably pre-mRNA splicing and ribosome biogenesis. However, little evidence supports that this protein is directly coupled to DNA replication and/or repair processes, as previously suggested. Furthermore, a novel interaction was observed with PRMT7 (protein arginine methyltransferase 7) and we demonstrated that KIN is modified by this enzyme. This interactome analysis indicates that KIN is associated with several cell metabolism functions, and shows for the first time an association with ribosome biogenesis, suggesting that KIN is likely a moonlight protein.

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The Complexity of Human Ribosome Biogenesis Revealed by Systematic Nucleolar Screening of Pre-rRNA Processing Factors

Molecular Cell ◽

10.1016/j.molcel.2013.08.011 ◽

2013 ◽

Vol 51 (4) ◽

pp. 539-551 ◽

Cited By ~ 219

Author(s):

Lionel Tafforeau ◽

Christiane Zorbas ◽

Jean-Louis Langhendries ◽

Sahra-Taylor Mullineux ◽

Vassiliki Stamatopoulou ◽

...

Keyword(s):

Ribosome Biogenesis ◽

Rrna Processing ◽

Processing Factors

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Bop1 Is a Mouse WD40 Repeat Nucleolar Protein Involved in 28S and 5.8S rRNA Processing and 60S Ribosome Biogenesis

Molecular and Cellular Biology ◽

10.1128/mcb.20.15.5516-5528.2000 ◽

2000 ◽

Vol 20 (15) ◽

pp. 5516-5528 ◽

Cited By ~ 117

Author(s):

Žaklina Strezoska ◽

Dimitri G. Pestov ◽

Lester F. Lau

Keyword(s):

Ribosome Biogenesis ◽

Ribosomal Subunit ◽

Weight Fraction ◽

Rnase A ◽

Mutant Form ◽

Rrna Processing ◽

Ribonucleoprotein Complexes ◽

Nuclear Extracts ◽

Mouse Tissues ◽

Mouse Cells

ABSTRACT We have identified and characterized a novel mouse protein, Bop1, which contains WD40 repeats and is highly conserved through evolution. bop1 is ubiquitously expressed in all mouse tissues examined and is upregulated during mid-G1 in serum-stimulated fibroblasts. Immunofluorescence analysis shows that Bop1 is localized predominantly to the nucleolus. In sucrose density gradients, Bop1 from nuclear extracts cosediments with the 50S-80S ribonucleoprotein particles that contain the 32S rRNA precursor. RNase A treatment disrupts these particles and releases Bop1 into a low-molecular-weight fraction. A mutant form of Bop1, Bop1Δ, which lacks 231 amino acids in the N- terminus, is colocalized with wild-type Bop1 in the nucleolus and in ribonucleoprotein complexes. Expression of Bop1Δ leads to cell growth arrest in the G1phase and results in a specific inhibition of the synthesis of the 28S and 5.8S rRNAs without affecting 18S rRNA formation. Pulse-chase analyses show that Bop1Δ expression results in a partial inhibition in the conversion of the 36S to the 32S pre-rRNA and a complete inhibition of the processing of the 32S pre-rRNA to form the mature 28S and 5.8S rRNAs. Concomitant with these defects in rRNA processing, expression of Bop1Δ in mouse cells leads to a deficit in the cytosolic 60S ribosomal subunits. These studies thus identify Bop1 as a novel, nonribosomal mammalian protein that plays a key role in the formation of the mature 28S and 5.8S rRNAs and in the biogenesis of the 60S ribosomal subunit.

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SRN1, a yeast gene involved in RNA processing, is identical to HEX2/REG1, a negative regulator in glucose repression

Molecular and Cellular Biology ◽

10.1128/mcb.12.6.2673-2680.1992 ◽

1992 ◽

Vol 12 (6) ◽

pp. 2673-2680

Author(s):

K S Tung ◽

L L Norbeck ◽

S L Nolan ◽

N S Atkinson ◽

A K Hopper

Keyword(s):

Rna Processing ◽

Carbon Catabolite Repression ◽

Glucose Repression ◽

Complementation Group ◽

Negative Regulator ◽

Rrna Processing ◽

Yeast Gene ◽

Trna Splicing ◽

Intervening Sequences ◽

First Time

The yeast RNA1 gene encodes a cytosolic protein that affects pre-tRNA splicing, pre-rRNA processing, the production of mRNA, and the export of RNA from the nucleus to the cytosol. In an attempt to understand how the RNA1 protein affects such a diverse set of processes, we sought second-site suppressors of a mutation, rna1-1, of the RNA1 locus. Mutations in a single complementation group were obtained. These lesions proved to be in the same gene, SRN1, identified previously in a search for second-site suppressors of mutations that affect the removal of intervening sequences from pre-mRNAs. The SRN1 gene was mapped, cloned, and sequenced. DNA sequence analysis and the phenotype of disruption mutations showed that, surprisingly, SRN1 is identical to HEX2/REG1, a gene that negatively regulates glucose-repressible genes. Interestingly, SRN1 is not a negative regulator of RNA1 at the transcriptional, translational, or protein stability level. However, SRN1 does regulate the level of two newly discovered antigens, p43 and p70, one of which is not glucose repressible. These studies for the first time link RNA processing and carbon catabolite repression.

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Ribonucleoproteins in Archaeal Pre-rRNA Processing and Modification

Archaea ◽

10.1155/2013/614735 ◽

2013 ◽

Vol 2013 ◽

pp. 1-14 ◽

Cited By ~ 13

Author(s):

W. S. Vincent Yip ◽

Nicholas G. Vincent ◽

Susan J. Baserga

Keyword(s):

Ribosome Biogenesis ◽

Gene Organization ◽

Rrna Gene ◽

Rrna Processing ◽

Ribosomal Rnas ◽

Intensive Research ◽

Complex Process ◽

Nucleotide Modification

Given that ribosomes are one of the most important cellular macromolecular machines, it is not surprising that there is intensive research in ribosome biogenesis. Ribosome biogenesis is a complex process. The maturation of ribosomal RNAs (rRNAs) requires not only the precise cleaving and folding of the pre-rRNA but also extensive nucleotide modifications. At the heart of the processing and modifications of pre-rRNAs in Archaea and Eukarya are ribonucleoprotein (RNP) machines. They are called small RNPs (sRNPs), in Archaea, and small nucleolar RNPs (snoRNPs), in Eukarya. Studies on ribosome biogenesis originally focused on eukaryotic systems. However, recent studies on archaeal sRNPs have provided important insights into the functions of these RNPs. This paper will introduce archaeal rRNA gene organization and pre-rRNA processing, with a particular focus on the discovery of the archaeal sRNP components, their functions in nucleotide modification, and their structures.

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