A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly
Identifieur interne : 001085 ( Istex/Corpus ); précédent : 001084; suivant : 001086A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly
Auteurs : Sigrid Schaper ; Micheline Fromont-Racine ; Patrick Linder ; Jesús De La Cruz ; Abdelkader Namane ; Moshe YanivSource :
- Current Biology [ 0960-9822 ] ; 2001.
English descriptors
- Teeft :
- Brief communication, Cell biol, Chromatin assembly factor, Cleavage site, Control reactions, Copurifying, Current biology, Degradation products, Different regions, Docteur roux, Early precursor rrna copurifies, Final eluate, Free couples, Hybridization, Institut pasteur, Mass spectrometry, Mass spectrometry analysis, Nylon membrane, Oligonucleotides number, Parallel preparation, Paris cedex, Polysome analysis, Polysome profile, Polysomes, Precursor, Precursor rrna, Preribosomal particle, Primer, Primer hybridization assay, Primer number, Primers number, Protein synthesis, Proteins copurify, Ribonuclease, Ribonuclease inhibitor, Ribosomal, Ribosomal protein, Ribosomal proteins, Ribosomal subunit, Ribosomal subunit assembly, Ribosomal subunits, Ribosome, Ribosome assembly, Ribosome biogenesis, Rrb1p, Rrb1p function results, Rrna, Saccharomyces cerevisiae, Same membrane, Spectrometry, Substoichiometrically copurifying, Subunit, Sucrose gradients, Supplementary material, Unpublished data, Yeast, Yeast homolog.
Abstract
Abstract: Cells have a recurrent need for the correct assembly of protein-nucleic acid complexes. We have studied a yeast homolog of the smallest subunit of chromatin assembly factor 1 (CAF1), encoded by YMR131c and termed “RRB1” [1]. Unlike other yeast homologs, Msi1p, and Hat2p, Rrb1p is essential for cell viability. Impairment of Rrb1p function results in decreased levels of free 60S ribosomal subunits and the appearance of half-mer polysomes, suggesting its involvement in ribosome biogenesis. Using tandem affinity purification (TAP [2]) combined with mass spectrometry, we show that Rrb1p is associated with ribosomal protein L3. A fraction of Rrb1p is also found in a protein-precursor rRNA complex containing at least ten other early-assembling ribosomal proteins. We propose that Rrb1p is required for proper assembly of preribosomal particles during early ribosome biogenesis, presumably by targeting L3 onto the 35S precursor rRNA. This action may resemble the mechanism by which CAF1 assembles histones H3/H4 onto newly replicated DNA.
Url:
DOI: 10.1016/S0960-9822(01)00584-X
Links to Exploration step
ISTEX:947E6E140DBABF8D8A3368FF8013F30F08DB0153Le document en format XML
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homolog of chromatin assembly factor 1 is involved in early ribosome assembly</title><author><name sortKey="Schaper, Sigrid" sort="Schaper, Sigrid" uniqKey="Schaper S" first="Sigrid" last="Schaper">Sigrid Schaper</name><affiliation><mods:affiliation>E-mail: schaper@molgen.mpg.de</mods:affiliation></affiliation><affiliation><mods:affiliation>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</mods:affiliation></affiliation></author><author><name sortKey="Fromont Racine, Micheline" sort="Fromont Racine, Micheline" uniqKey="Fromont Racine M" first="Micheline" last="Fromont-Racine">Micheline Fromont-Racine</name><affiliation><mods:affiliation>Unité Génétique des Interactions Macromoléculaires, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</mods:affiliation></affiliation></author><author><name sortKey="Linder, Patrick" sort="Linder, Patrick" uniqKey="Linder P" first="Patrick" last="Linder">Patrick Linder</name><affiliation><mods:affiliation>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</mods:affiliation></affiliation></author><author><name sortKey="De La Cruz, Jesus" sort="De La Cruz, Jesus" uniqKey="De La Cruz J" first="Jesús" last="De La Cruz">Jesús De La Cruz</name><affiliation><mods:affiliation>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</mods:affiliation></affiliation></author><author><name sortKey="Namane, Abdelkader" sort="Namane, Abdelkader" uniqKey="Namane A" first="Abdelkader" last="Namane">Abdelkader Namane</name><affiliation><mods:affiliation>Laboratoire de Chimie Structurale des Macromolécules, URA 2185 du CNRS, Département Biochimie et Génétique Moléculaire, Institut Pasteur, 28, Rue du Docteur Roux, 75724 Paris cedex 15, France</mods:affiliation></affiliation></author><author><name sortKey="Yaniv, Moshe" sort="Yaniv, Moshe" uniqKey="Yaniv M" first="Moshe" last="Yaniv">Moshe Yaniv</name><affiliation><mods:affiliation>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</mods:affiliation></affiliation></author></analytic><monogr></monogr><series><title level="j">Current Biology</title><title level="j" type="abbrev">CURBIO</title><idno type="ISSN">0960-9822</idno><imprint><publisher>ELSEVIER</publisher><date type="published" when="2001">2001</date><biblScope unit="volume">11</biblScope><biblScope unit="issue">23</biblScope><biblScope unit="page" from="1885">1885</biblScope><biblScope unit="page" to="1890">1890</biblScope></imprint><idno type="ISSN">0960-9822</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0960-9822</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="Teeft" xml:lang="en"><term>Brief communication</term><term>Cell biol</term><term>Chromatin assembly factor</term><term>Cleavage site</term><term>Control reactions</term><term>Copurifying</term><term>Current biology</term><term>Degradation products</term><term>Different regions</term><term>Docteur roux</term><term>Early precursor rrna copurifies</term><term>Final eluate</term><term>Free couples</term><term>Hybridization</term><term>Institut pasteur</term><term>Mass spectrometry</term><term>Mass spectrometry analysis</term><term>Nylon membrane</term><term>Oligonucleotides number</term><term>Parallel preparation</term><term>Paris cedex</term><term>Polysome analysis</term><term>Polysome profile</term><term>Polysomes</term><term>Precursor</term><term>Precursor rrna</term><term>Preribosomal particle</term><term>Primer</term><term>Primer hybridization assay</term><term>Primer number</term><term>Primers number</term><term>Protein synthesis</term><term>Proteins copurify</term><term>Ribonuclease</term><term>Ribonuclease inhibitor</term><term>Ribosomal</term><term>Ribosomal protein</term><term>Ribosomal proteins</term><term>Ribosomal subunit</term><term>Ribosomal subunit assembly</term><term>Ribosomal subunits</term><term>Ribosome</term><term>Ribosome assembly</term><term>Ribosome biogenesis</term><term>Rrb1p</term><term>Rrb1p function results</term><term>Rrna</term><term>Saccharomyces cerevisiae</term><term>Same membrane</term><term>Spectrometry</term><term>Substoichiometrically copurifying</term><term>Subunit</term><term>Sucrose gradients</term><term>Supplementary material</term><term>Unpublished data</term><term>Yeast</term><term>Yeast homolog</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Abstract: Cells have a recurrent need for the correct assembly of protein-nucleic acid complexes. We have studied a yeast homolog of the smallest subunit of chromatin assembly factor 1 (CAF1), encoded by YMR131c and termed “RRB1” [1]. Unlike other yeast homologs, Msi1p, and Hat2p, Rrb1p is essential for cell viability. Impairment of Rrb1p function results in decreased levels of free 60S ribosomal subunits and the appearance of half-mer polysomes, suggesting its involvement in ribosome biogenesis. Using tandem affinity purification (TAP [2]) combined with mass spectrometry, we show that Rrb1p is associated with ribosomal protein L3. A fraction of Rrb1p is also found in a protein-precursor rRNA complex containing at least ten other early-assembling ribosomal proteins. We propose that Rrb1p is required for proper assembly of preribosomal particles during early ribosome biogenesis, presumably by targeting L3 onto the 35S precursor rRNA. 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We have studied a yeast homolog of the smallest subunit of chromatin assembly factor 1 (CAF1), encoded by YMR131c and termed “RRB1” [1]. Unlike other yeast homologs, Msi1p, and Hat2p, Rrb1p is essential for cell viability. Impairment of Rrb1p function results in decreased levels of free 60S ribosomal subunits and the appearance of half-mer polysomes, suggesting its involvement in ribosome biogenesis. Using tandem affinity purification (TAP [2]) combined with mass spectrometry, we show that Rrb1p is associated with ribosomal protein L3. A fraction of Rrb1p is also found in a protein-precursor rRNA complex containing at least ten other early-assembling ribosomal proteins. We propose that Rrb1p is required for proper assembly of preribosomal particles during early ribosome biogenesis, presumably by targeting L3 onto the 35S precursor rRNA. This action may resemble the mechanism by which CAF1 assembles histones H3/H4 onto newly replicated DNA.</abstract><qualityIndicators><score>7.351</score><pdfWordCount>3479</pdfWordCount><pdfCharCount>21974</pdfCharCount><pdfVersion>1.3</pdfVersion><pdfPageCount>6</pdfPageCount><pdfPageSize>603 x 783 pts</pdfPageSize><refBibsNative>true</refBibsNative><abstractWordCount>156</abstractWordCount><abstractCharCount>1051</abstractCharCount><keywordCount>0</keywordCount></qualityIndicators><title>A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly</title><pmid><json:string>11728313</json:string></pmid><pii><json:string>S0960-9822(01)00584-X</json:string></pii><genre><json:string>brief-communication</json:string></genre><host><title>Current Biology</title><language><json:string>unknown</json:string></language><publicationDate>2001</publicationDate><issn><json:string>0960-9822</json:string></issn><pii><json:string>S0960-9822(00)X0047-4</json:string></pii><volume>11</volume><issue>23</issue><pages><first>1885</first><last>1890</last></pages><genre><json:string>journal</json:string></genre></host><namedEntities><unitex><date><json:string>40S</json:string><json:string>66S</json:string><json:string>80S</json:string><json:string>18S</json:string><json:string>25S</json:string><json:string>32S</json:string><json:string>2001</json:string><json:string>43S</json:string><json:string>90S</json:string><json:string>35S</json:string><json:string>60S</json:string><json:string>33S</json:string></date><geogName></geogName><orgName><json:string>Elsevier Science Ltd.</json:string><json:string>National Science Foundation and the Ministere</json:string><json:string>Swiss</json:string><json:string>Deutsche Forschungsgemeinschaft</json:string></orgName><orgName_funder><json:string>Swiss</json:string><json:string>Deutsche Forschungsgemeinschaft</json:string></orgName_funder><orgName_provider></orgName_provider><persName><json:string>Moshe Yaniv</json:string><json:string>J.R. Warner</json:string><json:string>B. Seraphin</json:string><json:string>J. Weitzman</json:string><json:string>C. Rousselle</json:string><json:string>Patrick Linder</json:string><json:string>B. Dujon</json:string><json:string>Micheline Fromont-Racine</json:string><json:string>Abdelkader Namane</json:string><json:string>Roux</json:string><json:string>B. Bourachot</json:string><json:string>A. Klochendler-Yeivin</json:string><json:string>C. Saveanu</json:string><json:string>W. Zachariae</json:string><json:string>D. Kressler</json:string></persName><placeName><json:string>Berlin</json:string><json:string>Switzerland</json:string><json:string>Germany</json:string><json:string>France</json:string></placeName><ref_url><json:string>http://images.cellpress.com/ supmat/supmatin.htm</json:string><json:string>http://www.embl-heidelberg</json:string></ref_url><ref_bibl><json:string>[4]</json:string><json:string>[12]</json:string><json:string>[18]</json:string><json:string>[11]</json:string><json:string>[17, 18]</json:string><json:string>[1]</json:string><json:string>[10]</json:string><json:string>[15, 16]</json:string><json:string>[3]</json:string><json:string>[6, 7]</json:string><json:string>[5]</json:string><json:string>[14]</json:string><json:string>[9]</json:string><json:string>[2]</json:string><json:string>[5, 13]</json:string></ref_bibl><bibl></bibl></unitex></namedEntities><ark><json:string>ark:/67375/6H6-WPW40GCX-L</json:string></ark><categories><wos><json:string>1 - science</json:string><json:string>2 - cell biology</json:string><json:string>2 - biochemistry & molecular biology</json:string></wos><scienceMetrix><json:string>1 - health sciences</json:string><json:string>2 - biomedical research</json:string><json:string>3 - developmental biology</json:string></scienceMetrix><scopus><json:string>1 - Life Sciences</json:string><json:string>2 - Agricultural and Biological Sciences</json:string><json:string>3 - General Agricultural and Biological Sciences</json:string><json:string>1 - Life Sciences</json:string><json:string>2 - Biochemistry, Genetics and Molecular Biology</json:string><json:string>3 - General Biochemistry, Genetics and Molecular Biology</json:string></scopus><inist><json:string>1 - sciences appliquees, technologies et medecines</json:string><json:string>2 - sciences biologiques et medicales</json:string><json:string>3 - sciences biologiques fondamentales et appliquees. psychologie</json:string><json:string>4 - immunologie fondamentale</json:string></inist></categories><publicationDate>2001</publicationDate><copyrightDate>2001</copyrightDate><doi><json:string>10.1016/S0960-9822(01)00584-X</json:string></doi><id>947E6E140DBABF8D8A3368FF8013F30F08DB0153</id><score>1</score><fulltext><json:item><extension>pdf</extension><original>true</original><mimetype>application/pdf</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-WPW40GCX-L/fulltext.pdf</uri></json:item><json:item><extension>zip</extension><original>false</original><mimetype>application/zip</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-WPW40GCX-L/bundle.zip</uri></json:item><istex:fulltextTEI uri="https://api.istex.fr/ark:/67375/6H6-WPW40GCX-L/fulltext.tei"><teiHeader><fileDesc><titleStmt><title level="a">A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly</title></titleStmt><publicationStmt><authority>ISTEX</authority><publisher scheme="https://scientific-publisher.data.istex.fr">ELSEVIER</publisher><availability><licence><p>©2001 Elsevier Science Ltd</p></licence><p scheme="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-HKKZVM7B-M">elsevier</p></availability><date>2001</date></publicationStmt><notesStmt><note type="brief-communication" scheme="https://content-type.data.istex.fr/ark:/67375/XTP-S9SX2MFS-0">brief-communication</note><note type="journal" scheme="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</note><note type="content">Section title: Brief communication</note><note type="content">Figure 1: L3 and other early-associating ribosomal proteins copurify with Rrb1-TAP. (a) The purified Rrb1-TAP complex was separated on a 16% SDS-polyacrylamide gel and subjected to a mass spectrometry analysis after Coomassie Blue staining. Identified proteins are indicated on the left. Unlabeled bands were nonidentifiable by mass spectrometry. Note that the designations A or B corresponding to the two gene copies for the r proteins L4, L7, L8, and S1 were omitted here because mass spectrometry analysis did not distinguish between the two gene products. Bands corresponding to degradation products of Rrb1p (*) or ribosomal protein L3 (**) are labeled with asterisks. (b) Two protein extracts containing the Rrb1-TAP complex were treated with ribonuclease inhibitor or ribonuclease A, respectively, and Rrb1-TAP complexes were purified as before. Complexes were separated on 16% SDS-polyacrylamide gels and silver stained. Arrowheads indicate the bands of the substoichiometrically copurifying ribosomal proteins which “disappear” upon the ribonuclease A treatment. A single asterisk indicates degradation products of Rrb1p. The band labeled with two asterisks is a degradation product of L3, possibly due to a protease contamination of ribonuclease A.</note><note type="content">Figure 2: An early precursor rRNA copurifies with the Rrb1-TAP complex. (a) Structure of the 35S pre-rRNA and processing sites (adapted from [5]). This precursor contains the sequences for the mature 18S, 5.8S, and 25S rRNAs that are separated by the two internal transcribed spacers ITS1 and ITS2 and flanked by two external transcribed spacers 5′ETS and 3′ETS. Bars represent mature rRNA species; lines represent the transcribed spacers. The primers used are labeled with their corresponding numbers. Numbered primers with an asterisk indicate that the primers have the reverse orientation. (b) The first two processing steps of the 35S pre-rRNA at the A0 and the A1 cleavage sites (see [5] for a complete scheme). (c) RT reaction and PCR. Purified Rrb1-TAP complex (2 μl) was used to amplify different regions within the precursor rRNA. Purified Sas5-TAP complex (2 μl) or 2.5 μl (2.5 μg) total extracted yeast RNA (Rt) was used in the control reactions. The multiple slower migrating bands in the reactions on total RNA are probably due to nonspecific annealing of the primers. PCR products were resolved on a 1% agarose gel and visualized by ethidium bromide staining. The bands corresponding to products from PCR on genomic DNA (D), using the same sets of primers, are labeled at either side of the gel. The diverse sets of primers used are indicated on top of the panel and are described in the text.</note><note type="content">Figure 3: Primer hybridization assay. RNA was extracted from the final Rrb1-TAP eluate or from a parallel preparation that was subjected to RNase A treatment during the course of TAP-purification or from a Sas5-TAP purification, heated briefly, and spotted (2 μl) onto a nylon membrane. For comparison, 2 μl total extracted RNA corresponding to the amounts indicated were spotted on the same membrane. Hybridizations were performed for 1 hr at 42°C, using successively 32P-labeled oligonucleotides number 15, number 14, or number 9 (see scheme, Figure 2a).</note><note type="content">Figure 4: The temperature-sensitive rrb1-TAP strain exhibits a deficit in free 60S ribosomal subunits and accumulation of half-mer polysomes. (a) The wild-type strain MGD353-13D and (b) the rrb1-TAP strain were grown at 30°C. Cells were harvested at an OD600 of 0.8, and cell extracts were resolved in 7%–50% (w/v) sucrose gradients. Gradients were analyzed by continuous monitoring at A254. Sedimentation is from left to right. The peaks of free 40S and 60S ribosomal subunits, 80S free couples/monosomes, and polysomes are indicated. Half-mers are labeled with asterisks. (c) Analysis of the sedimentation of Rrb1-TAP in the sucrose gradient shown in (b). Collected fractions were precipitated as described [21], resolved on a 15% SDS polyacrylamide gel, and immunoblotted using PAP-soluble antibody complex. The Rrb1-TAP signal and the positions of the 40S, 60S, and 80S ribosomal particles are indicated.</note></notesStmt><sourceDesc><biblStruct type="inbook"><analytic><title level="a">A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly</title><author xml:id="author-0000"><persName><forename type="first">Sigrid</forename><surname>Schaper</surname></persName><email>schaper@molgen.mpg.de</email><note type="biography">Present address: Otto-Warburg-Laboratories, Max-Planck-Institute for Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany.</note><note type="correspondence"><p>Correspondence: Sigrid Schaper</p></note><affiliation>Present address: Otto-Warburg-Laboratories, Max-Planck-Institute for Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany.</affiliation><affiliation>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation></author><author xml:id="author-0001"><persName><forename type="first">Micheline</forename><surname>Fromont-Racine</surname></persName><affiliation>Unité Génétique des Interactions Macromoléculaires, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation></author><author xml:id="author-0002"><persName><forename type="first">Patrick</forename><surname>Linder</surname></persName><affiliation>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</affiliation></author><author xml:id="author-0003"><persName><forename type="first">Jesús</forename><surname>de la Cruz</surname></persName><affiliation>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</affiliation></author><author xml:id="author-0004"><persName><forename type="first">Abdelkader</forename><surname>Namane</surname></persName><affiliation>Laboratoire de Chimie Structurale des Macromolécules, URA 2185 du CNRS, Département Biochimie et Génétique Moléculaire, Institut Pasteur, 28, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation></author><author xml:id="author-0005"><persName><forename type="first">Moshe</forename><surname>Yaniv</surname></persName><affiliation>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation></author><idno type="istex">947E6E140DBABF8D8A3368FF8013F30F08DB0153</idno><idno type="ark">ark:/67375/6H6-WPW40GCX-L</idno><idno type="DOI">10.1016/S0960-9822(01)00584-X</idno><idno type="PII">S0960-9822(01)00584-X</idno></analytic><monogr><title level="j">Current Biology</title><title level="j" type="abbrev">CURBIO</title><idno type="pISSN">0960-9822</idno><idno type="PII">S0960-9822(00)X0047-4</idno><imprint><publisher>ELSEVIER</publisher><date type="published" when="2001"></date><biblScope unit="volume">11</biblScope><biblScope unit="issue">23</biblScope><biblScope unit="page" from="1885">1885</biblScope><biblScope unit="page" to="1890">1890</biblScope></imprint></monogr></biblStruct></sourceDesc></fileDesc><profileDesc><creation><date>2001</date></creation><langUsage><language ident="en">en</language></langUsage><abstract xml:lang="en"><p>Abstract: Cells have a recurrent need for the correct assembly of protein-nucleic acid complexes. We have studied a yeast homolog of the smallest subunit of chromatin assembly factor 1 (CAF1), encoded by YMR131c and termed “RRB1” [1]. Unlike other yeast homologs, Msi1p, and Hat2p, Rrb1p is essential for cell viability. Impairment of Rrb1p function results in decreased levels of free 60S ribosomal subunits and the appearance of half-mer polysomes, suggesting its involvement in ribosome biogenesis. Using tandem affinity purification (TAP [2]) combined with mass spectrometry, we show that Rrb1p is associated with ribosomal protein L3. A fraction of Rrb1p is also found in a protein-precursor rRNA complex containing at least ten other early-assembling ribosomal proteins. We propose that Rrb1p is required for proper assembly of preribosomal particles during early ribosome biogenesis, presumably by targeting L3 onto the 35S precursor rRNA. This action may resemble the mechanism by which CAF1 assembles histones H3/H4 onto newly replicated DNA.</p></abstract></profileDesc><revisionDesc><change when="2001-10-12">Modified</change><change when="2001">Published</change></revisionDesc></teiHeader></istex:fulltextTEI><json:item><extension>txt</extension><original>false</original><mimetype>text/plain</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-WPW40GCX-L/fulltext.txt</uri></json:item></fulltext><metadata><istex:metadataXml wicri:clean="Elsevier, elements deleted: ce:floats; body; tail"><istex:xmlDeclaration>version="1.0" encoding="utf-8"</istex:xmlDeclaration><istex:docType PUBLIC="-//ES//DTD journal article DTD version 4.5.2//EN//XML" URI="art452.dtd" name="istex:docType"><istex:entity SYSTEM="gr1" NDATA="IMAGE" name="gr1"></istex:entity><istex:entity SYSTEM="gr2" NDATA="IMAGE" name="gr2"></istex:entity><istex:entity SYSTEM="gr3" NDATA="IMAGE" name="gr3"></istex:entity><istex:entity SYSTEM="gr4" NDATA="IMAGE" name="gr4"></istex:entity><istex:entity SYSTEM="mmc1" NDATA="APPLICATION" name="mmc1"></istex:entity></istex:docType><istex:document><converted-article version="4.5.2" docsubtype="sco"><item-info><jid>CURBIO</jid><aid>1474</aid><ce:pii>S0960-9822(01)00584-X</ce:pii><ce:doi>10.1016/S0960-9822(01)00584-X</ce:doi><ce:copyright type="full-transfer" year="2001">Elsevier Science Ltd</ce:copyright></item-info><head><ce:dochead><ce:textfn>Brief communication</ce:textfn></ce:dochead><ce:title>A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly</ce:title><ce:author-group><ce:author><ce:given-name>Sigrid</ce:given-name><ce:surname>Schaper</ce:surname><ce:cross-ref refid="FN1">¶</ce:cross-ref><ce:cross-ref refid="AFF1">*</ce:cross-ref><ce:cross-ref refid="CORR1">*</ce:cross-ref><ce:e-address>schaper@molgen.mpg.de</ce:e-address></ce:author><ce:author><ce:given-name>Micheline</ce:given-name><ce:surname>Fromont-Racine</ce:surname><ce:cross-ref refid="AFF2">†</ce:cross-ref></ce:author><ce:author><ce:given-name>Patrick</ce:given-name><ce:surname>Linder</ce:surname><ce:cross-ref refid="AFF3">‡</ce:cross-ref></ce:author><ce:author><ce:given-name>Jesús</ce:given-name><ce:surname>de la Cruz</ce:surname><ce:cross-ref refid="AFF3">‡</ce:cross-ref></ce:author><ce:author><ce:given-name>Abdelkader</ce:given-name><ce:surname>Namane</ce:surname><ce:cross-ref refid="AFF4">§</ce:cross-ref></ce:author><ce:author><ce:given-name>Moshe</ce:given-name><ce:surname>Yaniv</ce:surname><ce:cross-ref refid="AFF1">*</ce:cross-ref></ce:author><ce:affiliation id="AFF1"><ce:label>*</ce:label><ce:textfn>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</ce:textfn></ce:affiliation><ce:affiliation id="AFF2"><ce:label>†</ce:label><ce:textfn>Unité Génétique des Interactions Macromoléculaires, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</ce:textfn></ce:affiliation><ce:affiliation id="AFF3"><ce:label>‡</ce:label><ce:textfn>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</ce:textfn></ce:affiliation><ce:affiliation id="AFF4"><ce:label>§</ce:label><ce:textfn>Laboratoire de Chimie Structurale des Macromolécules, URA 2185 du CNRS, Département Biochimie et Génétique Moléculaire, Institut Pasteur, 28, Rue du Docteur Roux, 75724 Paris cedex 15, France</ce:textfn></ce:affiliation><ce:correspondence id="CORR1"><ce:label>*</ce:label><ce:text>Correspondence: Sigrid Schaper</ce:text></ce:correspondence><ce:footnote id="FN1"><ce:label>¶</ce:label><ce:note-para>Present address: Otto-Warburg-Laboratories, Max-Planck-Institute for Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany.</ce:note-para></ce:footnote></ce:author-group><ce:date-received day="30" month="8" year="2001"></ce:date-received><ce:date-revised day="12" month="10" year="2001"></ce:date-revised><ce:date-accepted day="15" month="10" year="2001"></ce:date-accepted><ce:abstract><ce:section-title>Abstract</ce:section-title><ce:abstract-sec><ce:simple-para id="simple-para0025">Cells have a recurrent need for the correct assembly of protein-nucleic acid complexes. We have studied a yeast homolog of the smallest subunit of chromatin assembly factor 1 (CAF1), encoded by <ce:italic>YMR131c</ce:italic> and termed “<ce:italic>RRB1</ce:italic>” <ce:cross-ref refid="BIB1">[1]</ce:cross-ref>. Unlike other yeast homologs, Msi1p, and Hat2p, Rrb1p is essential for cell viability. Impairment of Rrb1p function results in decreased levels of free 60S ribosomal subunits and the appearance of half-mer polysomes, suggesting its involvement in ribosome biogenesis. Using tandem affinity purification (TAP <ce:cross-ref refid="BIB2">[2]</ce:cross-ref>) combined with mass spectrometry, we show that Rrb1p is associated with ribosomal protein L3. A fraction of Rrb1p is also found in a protein-precursor rRNA complex containing at least ten other early-assembling ribosomal proteins. We propose that Rrb1p is required for proper assembly of preribosomal particles during early ribosome biogenesis, presumably by targeting L3 onto the 35S precursor rRNA. This action may resemble the mechanism by which CAF1 assembles histones H3/H4 onto newly replicated DNA.</ce:simple-para></ce:abstract-sec></ce:abstract></head></converted-article></istex:document></istex:metadataXml><mods version="3.6"><titleInfo><title>A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly</title></titleInfo><titleInfo type="alternative" contentType="CDATA"><title>A yeast homolog of chromatin assembly factor 1 is involved in early ribosome assembly</title></titleInfo><name type="personal"><namePart type="given">Sigrid</namePart><namePart type="family">Schaper</namePart><affiliation>E-mail: schaper@molgen.mpg.de</affiliation><affiliation>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation><description>Correspondence: Sigrid Schaper</description><description>Present address: Otto-Warburg-Laboratories, Max-Planck-Institute for Molecular Genetics, Ihnestrasse 73, D-14195 Berlin, Germany.</description><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Micheline</namePart><namePart type="family">Fromont-Racine</namePart><affiliation>Unité Génétique des Interactions Macromoléculaires, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Patrick</namePart><namePart type="family">Linder</namePart><affiliation>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Jesús</namePart><namePart type="family">de la Cruz</namePart><affiliation>Département de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, 1211 Genève 4, Switzerland</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Abdelkader</namePart><namePart type="family">Namane</namePart><affiliation>Laboratoire de Chimie Structurale des Macromolécules, URA 2185 du CNRS, Département Biochimie et Génétique Moléculaire, Institut Pasteur, 28, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Moshe</namePart><namePart type="family">Yaniv</namePart><affiliation>Unité Virus Oncogènes, URA 1644 du CNRS, Département des Biotechnologies, Institut Pasteur, 25, Rue du Docteur Roux, 75724 Paris cedex 15, France</affiliation><role><roleTerm type="text">author</roleTerm></role></name><typeOfResource>text</typeOfResource><genre type="brief-communication" displayLabel="Short communication" authority="ISTEX" authorityURI="https://content-type.data.istex.fr" valueURI="https://content-type.data.istex.fr/ark:/67375/XTP-S9SX2MFS-0">brief-communication</genre><originInfo><publisher>ELSEVIER</publisher><dateIssued encoding="w3cdtf">2001</dateIssued><dateModified encoding="w3cdtf">2001-10-12</dateModified><copyrightDate encoding="w3cdtf">2001</copyrightDate></originInfo><language><languageTerm type="code" authority="iso639-2b">eng</languageTerm><languageTerm type="code" authority="rfc3066">en</languageTerm></language><abstract lang="en">Abstract: Cells have a recurrent need for the correct assembly of protein-nucleic acid complexes. We have studied a yeast homolog of the smallest subunit of chromatin assembly factor 1 (CAF1), encoded by YMR131c and termed “RRB1” [1]. Unlike other yeast homologs, Msi1p, and Hat2p, Rrb1p is essential for cell viability. Impairment of Rrb1p function results in decreased levels of free 60S ribosomal subunits and the appearance of half-mer polysomes, suggesting its involvement in ribosome biogenesis. Using tandem affinity purification (TAP [2]) combined with mass spectrometry, we show that Rrb1p is associated with ribosomal protein L3. A fraction of Rrb1p is also found in a protein-precursor rRNA complex containing at least ten other early-assembling ribosomal proteins. We propose that Rrb1p is required for proper assembly of preribosomal particles during early ribosome biogenesis, presumably by targeting L3 onto the 35S precursor rRNA. This action may resemble the mechanism by which CAF1 assembles histones H3/H4 onto newly replicated DNA.</abstract><note type="content">Section title: Brief communication</note><note type="content">Figure 1: L3 and other early-associating ribosomal proteins copurify with Rrb1-TAP. (a) The purified Rrb1-TAP complex was separated on a 16% SDS-polyacrylamide gel and subjected to a mass spectrometry analysis after Coomassie Blue staining. Identified proteins are indicated on the left. Unlabeled bands were nonidentifiable by mass spectrometry. Note that the designations A or B corresponding to the two gene copies for the r proteins L4, L7, L8, and S1 were omitted here because mass spectrometry analysis did not distinguish between the two gene products. Bands corresponding to degradation products of Rrb1p (*) or ribosomal protein L3 (**) are labeled with asterisks. (b) Two protein extracts containing the Rrb1-TAP complex were treated with ribonuclease inhibitor or ribonuclease A, respectively, and Rrb1-TAP complexes were purified as before. Complexes were separated on 16% SDS-polyacrylamide gels and silver stained. Arrowheads indicate the bands of the substoichiometrically copurifying ribosomal proteins which “disappear” upon the ribonuclease A treatment. A single asterisk indicates degradation products of Rrb1p. The band labeled with two asterisks is a degradation product of L3, possibly due to a protease contamination of ribonuclease A.</note><note type="content">Figure 2: An early precursor rRNA copurifies with the Rrb1-TAP complex. (a) Structure of the 35S pre-rRNA and processing sites (adapted from [5]). This precursor contains the sequences for the mature 18S, 5.8S, and 25S rRNAs that are separated by the two internal transcribed spacers ITS1 and ITS2 and flanked by two external transcribed spacers 5′ETS and 3′ETS. Bars represent mature rRNA species; lines represent the transcribed spacers. The primers used are labeled with their corresponding numbers. Numbered primers with an asterisk indicate that the primers have the reverse orientation. (b) The first two processing steps of the 35S pre-rRNA at the A0 and the A1 cleavage sites (see [5] for a complete scheme). (c) RT reaction and PCR. Purified Rrb1-TAP complex (2 μl) was used to amplify different regions within the precursor rRNA. Purified Sas5-TAP complex (2 μl) or 2.5 μl (2.5 μg) total extracted yeast RNA (Rt) was used in the control reactions. The multiple slower migrating bands in the reactions on total RNA are probably due to nonspecific annealing of the primers. PCR products were resolved on a 1% agarose gel and visualized by ethidium bromide staining. The bands corresponding to products from PCR on genomic DNA (D), using the same sets of primers, are labeled at either side of the gel. The diverse sets of primers used are indicated on top of the panel and are described in the text.</note><note type="content">Figure 3: Primer hybridization assay. RNA was extracted from the final Rrb1-TAP eluate or from a parallel preparation that was subjected to RNase A treatment during the course of TAP-purification or from a Sas5-TAP purification, heated briefly, and spotted (2 μl) onto a nylon membrane. For comparison, 2 μl total extracted RNA corresponding to the amounts indicated were spotted on the same membrane. Hybridizations were performed for 1 hr at 42°C, using successively 32P-labeled oligonucleotides number 15, number 14, or number 9 (see scheme, Figure 2a).</note><note type="content">Figure 4: The temperature-sensitive rrb1-TAP strain exhibits a deficit in free 60S ribosomal subunits and accumulation of half-mer polysomes. (a) The wild-type strain MGD353-13D and (b) the rrb1-TAP strain were grown at 30°C. Cells were harvested at an OD600 of 0.8, and cell extracts were resolved in 7%–50% (w/v) sucrose gradients. Gradients were analyzed by continuous monitoring at A254. Sedimentation is from left to right. The peaks of free 40S and 60S ribosomal subunits, 80S free couples/monosomes, and polysomes are indicated. Half-mers are labeled with asterisks. (c) Analysis of the sedimentation of Rrb1-TAP in the sucrose gradient shown in (b). Collected fractions were precipitated as described [21], resolved on a 15% SDS polyacrylamide gel, and immunoblotted using PAP-soluble antibody complex. The Rrb1-TAP signal and the positions of the 40S, 60S, and 80S ribosomal particles are indicated.</note><relatedItem type="host"><titleInfo><title>Current Biology</title></titleInfo><titleInfo type="abbreviated"><title>CURBIO</title></titleInfo><genre type="journal" authority="ISTEX" authorityURI="https://publication-type.data.istex.fr" valueURI="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</genre><originInfo><publisher>ELSEVIER</publisher><dateIssued encoding="w3cdtf">2001</dateIssued></originInfo><identifier type="ISSN">0960-9822</identifier><identifier type="PII">S0960-9822(00)X0047-4</identifier><part><date>2001</date><detail type="volume"><number>11</number><caption>vol.</caption></detail><detail type="issue"><number>23</number><caption>no.</caption></detail><extent unit="issue-pages"><start>R947</start><end>R996</end></extent><extent unit="issue-pages"><start>1815</start><end>1902</end></extent><extent unit="pages"><start>1885</start><end>1890</end></extent></part></relatedItem><identifier type="istex">947E6E140DBABF8D8A3368FF8013F30F08DB0153</identifier><identifier type="ark">ark:/67375/6H6-WPW40GCX-L</identifier><identifier type="DOI">10.1016/S0960-9822(01)00584-X</identifier><identifier type="PII">S0960-9822(01)00584-X</identifier><accessCondition type="use and reproduction" contentType="copyright">©2001 Elsevier Science Ltd</accessCondition><recordInfo><recordContentSource authority="ISTEX" authorityURI="https://loaded-corpus.data.istex.fr" valueURI="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-HKKZVM7B-M">elsevier</recordContentSource><recordOrigin>Elsevier Science Ltd, ©2001</recordOrigin></recordInfo></mods><json:item><extension>json</extension><original>false</original><mimetype>application/json</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-WPW40GCX-L/record.json</uri></json:item></metadata><serie></serie></istex></record>Pour manipuler ce document sous Unix (Dilib)
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