Characterization of Staufen 1 ribonucleoprotein complexes
In Drosophila oocytes and neuroblasts, the double-stranded RNA binding protein Staufen assembles into ribonucleoprotein particles, which mediate cytoplasmic mRNA trafficking and translation. Two different mammalian orthologues also appear to reside in distinct RNA-containing particles. To date, rela...
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description | In Drosophila oocytes and neuroblasts, the double-stranded RNA binding protein Staufen assembles into ribonucleoprotein particles, which mediate cytoplasmic mRNA trafficking and translation. Two different mammalian orthologues also appear to reside in distinct RNA-containing particles. To date, relatively little is known about the molecular composition of Staufen-containing ribonucleoprotein complexes. Here, we have used a novel one-step affinity purification protocol to identify components of Staufen 1-containing particles. Whereas the nucleocytoplasmic RNA-binding protein nucleolin is linked to Staufen in an RNA-dependent manner, the association of protein phosphatase 1, the microtubule-dependent motor protein kinesin and several components of the large and small ribosomal subunits with Staufen ribonucleoprotein complexes is RNA-independent. Notably, all these components do not co-purify with a second RNA-binding protein, hnRNPK (heterogeneous ribonucleoprotein K), demonstrating the high specificity of the purification protocol. Furthermore, pull-down and immunoprecipitation experiments suggest a direct interaction between Staufen 1 and the ribosomal protein P0 in vitro as well as in cells. In cell fractionation and sucrose gradient assays, Staufen co-fractionates with intact ribosomes and polysomes, but not with the isolated 40 S ribosomal subunit. Taken together, these findings imply that, in the cytoplasm of mammalian cells, an association with the ribosomal P-stalk protein P0 recruits Staufen 1 into ribosome-containing ribonucleoprotein particles, which also contain kinesin, protein phosphatase 1 and nucleolin. |
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Two different mammalian orthologues also appear to reside in distinct RNA-containing particles. To date, relatively little is known about the molecular composition of Staufen-containing ribonucleoprotein complexes. Here, we have used a novel one-step affinity purification protocol to identify components of Staufen 1-containing particles. Whereas the nucleocytoplasmic RNA-binding protein nucleolin is linked to Staufen in an RNA-dependent manner, the association of protein phosphatase 1, the microtubule-dependent motor protein kinesin and several components of the large and small ribosomal subunits with Staufen ribonucleoprotein complexes is RNA-independent. Notably, all these components do not co-purify with a second RNA-binding protein, hnRNPK (heterogeneous ribonucleoprotein K), demonstrating the high specificity of the purification protocol. Furthermore, pull-down and immunoprecipitation experiments suggest a direct interaction between Staufen 1 and the ribosomal protein P0 in vitro as well as in cells. In cell fractionation and sucrose gradient assays, Staufen co-fractionates with intact ribosomes and polysomes, but not with the isolated 40 S ribosomal subunit. Taken together, these findings imply that, in the cytoplasm of mammalian cells, an association with the ribosomal P-stalk protein P0 recruits Staufen 1 into ribosome-containing ribonucleoprotein particles, which also contain kinesin, protein phosphatase 1 and nucleolin.</description><identifier>ISSN: 0264-6021</identifier><identifier>EISSN: 1470-8728</identifier><identifier>DOI: 10.1042/BJ20040812</identifier><identifier>PMID: 15303970</identifier><language>eng</language><publisher>England: Portland Press Ltd</publisher><subject>Animals ; Brain - cytology ; Brain - metabolism ; Cell Fractionation - methods ; Cells, Cultured ; Chromatography, Affinity - methods ; Drosophila ; Humans ; Kidney - chemistry ; Kidney - cytology ; Kidney - embryology ; Kidney - metabolism ; Multiprotein Complexes - chemistry ; Neurons - chemistry ; Neurons - metabolism ; Nucleolin ; Phosphoproteins - metabolism ; Polyribosomes - chemistry ; Rats ; Ribonucleoproteins - chemistry ; Ribosomal Proteins - metabolism ; Ribosomes - chemistry ; RNA - metabolism ; RNA-Binding Proteins - chemistry ; RNA-Binding Proteins - metabolism ; Transfection - methods</subject><ispartof>Biochemical journal, 2004-12, Vol.384 (Pt 2), p.239-246</ispartof><rights>The Biochemical Society, London 2004</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c438t-7b9ba1e9ed94595a99500c24201d6ab20a33d2aefd4d1941d7ccbc3cb5eb3ef53</citedby><cites>FETCH-LOGICAL-c438t-7b9ba1e9ed94595a99500c24201d6ab20a33d2aefd4d1941d7ccbc3cb5eb3ef53</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC1134106/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC1134106/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,723,776,780,881,27901,27902,53766,53768</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15303970$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Brendel, Cornelia</creatorcontrib><creatorcontrib>Rehbein, Monika</creatorcontrib><creatorcontrib>Kreienkamp, Hans-Jürgen</creatorcontrib><creatorcontrib>Buck, Friedrich</creatorcontrib><creatorcontrib>Richter, Dietmar</creatorcontrib><creatorcontrib>Kindler, Stefan</creatorcontrib><title>Characterization of Staufen 1 ribonucleoprotein complexes</title><title>Biochemical journal</title><addtitle>Biochem J</addtitle><description>In Drosophila oocytes and neuroblasts, the double-stranded RNA binding protein Staufen assembles into ribonucleoprotein particles, which mediate cytoplasmic mRNA trafficking and translation. Two different mammalian orthologues also appear to reside in distinct RNA-containing particles. To date, relatively little is known about the molecular composition of Staufen-containing ribonucleoprotein complexes. Here, we have used a novel one-step affinity purification protocol to identify components of Staufen 1-containing particles. Whereas the nucleocytoplasmic RNA-binding protein nucleolin is linked to Staufen in an RNA-dependent manner, the association of protein phosphatase 1, the microtubule-dependent motor protein kinesin and several components of the large and small ribosomal subunits with Staufen ribonucleoprotein complexes is RNA-independent. Notably, all these components do not co-purify with a second RNA-binding protein, hnRNPK (heterogeneous ribonucleoprotein K), demonstrating the high specificity of the purification protocol. Furthermore, pull-down and immunoprecipitation experiments suggest a direct interaction between Staufen 1 and the ribosomal protein P0 in vitro as well as in cells. In cell fractionation and sucrose gradient assays, Staufen co-fractionates with intact ribosomes and polysomes, but not with the isolated 40 S ribosomal subunit. Taken together, these findings imply that, in the cytoplasm of mammalian cells, an association with the ribosomal P-stalk protein P0 recruits Staufen 1 into ribosome-containing ribonucleoprotein particles, which also contain kinesin, protein phosphatase 1 and nucleolin.</description><subject>Animals</subject><subject>Brain - cytology</subject><subject>Brain - metabolism</subject><subject>Cell Fractionation - methods</subject><subject>Cells, Cultured</subject><subject>Chromatography, Affinity - methods</subject><subject>Drosophila</subject><subject>Humans</subject><subject>Kidney - chemistry</subject><subject>Kidney - cytology</subject><subject>Kidney - embryology</subject><subject>Kidney - metabolism</subject><subject>Multiprotein Complexes - chemistry</subject><subject>Neurons - chemistry</subject><subject>Neurons - metabolism</subject><subject>Nucleolin</subject><subject>Phosphoproteins - metabolism</subject><subject>Polyribosomes - chemistry</subject><subject>Rats</subject><subject>Ribonucleoproteins - chemistry</subject><subject>Ribosomal Proteins - metabolism</subject><subject>Ribosomes - chemistry</subject><subject>RNA - metabolism</subject><subject>RNA-Binding Proteins - chemistry</subject><subject>RNA-Binding Proteins - metabolism</subject><subject>Transfection - methods</subject><issn>0264-6021</issn><issn>1470-8728</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqN0T1PwzAQBmALgWj5WPgBKBMDUuDOdj68IEHFpyoxALPlOBcalMbFThDw62nVisIE0w336NWdXsYOEE4QJD-9uOMAEnLkG2yIMoM4z3i-yYbAUxmnwHHAdkJ4AUA5d9tsgIkAoTIYMjWaGG9sR77-NF3t2shV0UNn-oraCCNfF67tbUNu5l1HdRtZN5019E5hj21Vpgm0v5q77Onq8nF0E4_vr29H5-PYSpF3cVaowiApKpVMVGKUSgAslxywTE3BwQhRckNVKUtUEsvM2sIKWyRUCKoSscvOlrmzvphSaantvGn0zNdT4z-0M7X-vWnriX52bxpRSIR0HnC0CvDutafQ6WkdLDWNacn1QacZ5PPL8j8hKiWBwz9glshMqQU8XkLrXQiequ-zEfSiO73ubo4Pfz66pquyxBcewZTv</recordid><startdate>20041201</startdate><enddate>20041201</enddate><creator>Brendel, Cornelia</creator><creator>Rehbein, Monika</creator><creator>Kreienkamp, Hans-Jürgen</creator><creator>Buck, Friedrich</creator><creator>Richter, Dietmar</creator><creator>Kindler, Stefan</creator><general>Portland Press Ltd</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7SS</scope><scope>7TM</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20041201</creationdate><title>Characterization of Staufen 1 ribonucleoprotein complexes</title><author>Brendel, Cornelia ; Rehbein, Monika ; Kreienkamp, Hans-Jürgen ; Buck, Friedrich ; Richter, Dietmar ; Kindler, Stefan</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c438t-7b9ba1e9ed94595a99500c24201d6ab20a33d2aefd4d1941d7ccbc3cb5eb3ef53</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2004</creationdate><topic>Animals</topic><topic>Brain - cytology</topic><topic>Brain - metabolism</topic><topic>Cell Fractionation - methods</topic><topic>Cells, Cultured</topic><topic>Chromatography, Affinity - methods</topic><topic>Drosophila</topic><topic>Humans</topic><topic>Kidney - chemistry</topic><topic>Kidney - cytology</topic><topic>Kidney - embryology</topic><topic>Kidney - metabolism</topic><topic>Multiprotein Complexes - chemistry</topic><topic>Neurons - chemistry</topic><topic>Neurons - metabolism</topic><topic>Nucleolin</topic><topic>Phosphoproteins - metabolism</topic><topic>Polyribosomes - chemistry</topic><topic>Rats</topic><topic>Ribonucleoproteins - chemistry</topic><topic>Ribosomal Proteins - metabolism</topic><topic>Ribosomes - chemistry</topic><topic>RNA - metabolism</topic><topic>RNA-Binding Proteins - chemistry</topic><topic>RNA-Binding Proteins - metabolism</topic><topic>Transfection - methods</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Brendel, Cornelia</creatorcontrib><creatorcontrib>Rehbein, Monika</creatorcontrib><creatorcontrib>Kreienkamp, Hans-Jürgen</creatorcontrib><creatorcontrib>Buck, Friedrich</creatorcontrib><creatorcontrib>Richter, Dietmar</creatorcontrib><creatorcontrib>Kindler, Stefan</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Nucleic Acids Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Biochemical journal</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Brendel, Cornelia</au><au>Rehbein, Monika</au><au>Kreienkamp, Hans-Jürgen</au><au>Buck, Friedrich</au><au>Richter, Dietmar</au><au>Kindler, Stefan</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Characterization of Staufen 1 ribonucleoprotein complexes</atitle><jtitle>Biochemical journal</jtitle><addtitle>Biochem J</addtitle><date>2004-12-01</date><risdate>2004</risdate><volume>384</volume><issue>Pt 2</issue><spage>239</spage><epage>246</epage><pages>239-246</pages><issn>0264-6021</issn><eissn>1470-8728</eissn><abstract>In Drosophila oocytes and neuroblasts, the double-stranded RNA binding protein Staufen assembles into ribonucleoprotein particles, which mediate cytoplasmic mRNA trafficking and translation. Two different mammalian orthologues also appear to reside in distinct RNA-containing particles. To date, relatively little is known about the molecular composition of Staufen-containing ribonucleoprotein complexes. Here, we have used a novel one-step affinity purification protocol to identify components of Staufen 1-containing particles. Whereas the nucleocytoplasmic RNA-binding protein nucleolin is linked to Staufen in an RNA-dependent manner, the association of protein phosphatase 1, the microtubule-dependent motor protein kinesin and several components of the large and small ribosomal subunits with Staufen ribonucleoprotein complexes is RNA-independent. Notably, all these components do not co-purify with a second RNA-binding protein, hnRNPK (heterogeneous ribonucleoprotein K), demonstrating the high specificity of the purification protocol. Furthermore, pull-down and immunoprecipitation experiments suggest a direct interaction between Staufen 1 and the ribosomal protein P0 in vitro as well as in cells. In cell fractionation and sucrose gradient assays, Staufen co-fractionates with intact ribosomes and polysomes, but not with the isolated 40 S ribosomal subunit. Taken together, these findings imply that, in the cytoplasm of mammalian cells, an association with the ribosomal P-stalk protein P0 recruits Staufen 1 into ribosome-containing ribonucleoprotein particles, which also contain kinesin, protein phosphatase 1 and nucleolin.</abstract><cop>England</cop><pub>Portland Press Ltd</pub><pmid>15303970</pmid><doi>10.1042/BJ20040812</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record> |
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subjects | Animals Brain - cytology Brain - metabolism Cell Fractionation - methods Cells, Cultured Chromatography, Affinity - methods Drosophila Humans Kidney - chemistry Kidney - cytology Kidney - embryology Kidney - metabolism Multiprotein Complexes - chemistry Neurons - chemistry Neurons - metabolism Nucleolin Phosphoproteins - metabolism Polyribosomes - chemistry Rats Ribonucleoproteins - chemistry Ribosomal Proteins - metabolism Ribosomes - chemistry RNA - metabolism RNA-Binding Proteins - chemistry RNA-Binding Proteins - metabolism Transfection - methods |
title | Characterization of Staufen 1 ribonucleoprotein complexes |
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