Proteomic Analysis of SRm160-containing Complexes Reveals a Conserved Association with Cohesin
In this study, we describe a rapid immunoaffinity purification procedure for gel-free tandem mass spectrometry-based analysis of endogenous protein complexes and apply it to the characterization of complexes containing the SRm160 (serine/arginine repeat-related nuclear matrix protein of 160 kDa) spl...
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| Veröffentlicht in: | The Journal of biological chemistry 2005-12, Vol.280 (51), p.42227-42236 |
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| container_title | The Journal of biological chemistry |
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| creator | McCracken, Susan Longman, Dasa Marcon, Edyta Moens, Peter Downey, Michael Nickerson, Jeffrey A. Jessberger, Rolf Wilde, Andrew Caceres, Javier F. Emili, Andrew Blencowe, Benjamin J. |
| description | In this study, we describe a rapid immunoaffinity purification procedure for gel-free tandem mass spectrometry-based analysis of endogenous protein complexes and apply it to the characterization of complexes containing the SRm160 (serine/arginine repeat-related nuclear matrix protein of 160 kDa) splicing coactivator. In addition to promoting splicing, SRm160 stimulates 3′-end processing via its N-terminal PWI nucleic acid-binding domain and is found in a post-splicing exon junction complex that has been implicated in coupling splicing with mRNA turnover, export, and translation. Consistent with these known functional associations, we found that the majority of proteins identified in SRm160-containing complexes are associated with pre-mRNA processing. Interestingly, SRm160 is also associated with factors involved in chromatin regulation and sister chromatid cohesion, specifically the cohesin subunits SMC1α, SMC3, RAD21, and SA2. Gradient fractionation suggested that there are two predominant SRm160-containing complexes, one enriched in splicing components and the other enriched in cohesin subunits. Co-immunoprecipitation and co-localization experiments, as well as combinatorial RNA interference in Caenorhabditis elegans, support the existence of conserved and functional interactions between SRm160 and cohesin. |
| doi_str_mv | 10.1074/jbc.M507410200 |
| format | Article |
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In addition to promoting splicing, SRm160 stimulates 3′-end processing via its N-terminal PWI nucleic acid-binding domain and is found in a post-splicing exon junction complex that has been implicated in coupling splicing with mRNA turnover, export, and translation. Consistent with these known functional associations, we found that the majority of proteins identified in SRm160-containing complexes are associated with pre-mRNA processing. Interestingly, SRm160 is also associated with factors involved in chromatin regulation and sister chromatid cohesion, specifically the cohesin subunits SMC1α, SMC3, RAD21, and SA2. Gradient fractionation suggested that there are two predominant SRm160-containing complexes, one enriched in splicing components and the other enriched in cohesin subunits. Co-immunoprecipitation and co-localization experiments, as well as combinatorial RNA interference in Caenorhabditis elegans, support the existence of conserved and functional interactions between SRm160 and cohesin.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1074/jbc.M507410200</identifier><identifier>PMID: 16159877</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Animals ; Antigens, Nuclear - chemistry ; Antigens, Nuclear - genetics ; Antigens, Nuclear - metabolism ; Caenorhabditis elegans ; Caenorhabditis elegans - genetics ; Cell Cycle Proteins - metabolism ; Chromosomal Proteins, Non-Histone ; Cohesins ; Fungal Proteins - metabolism ; HeLa Cells ; Humans ; Immunoprecipitation ; Mass Spectrometry ; Nuclear Matrix-Associated Proteins - chemistry ; Nuclear Matrix-Associated Proteins - genetics ; Nuclear Matrix-Associated Proteins - metabolism ; Nuclear Proteins - metabolism ; Protein Binding ; Proteome ; RNA Splicing ; RNA, Messenger - genetics ; RNA-Binding Proteins - chemistry ; RNA-Binding Proteins - genetics ; RNA-Binding Proteins - metabolism</subject><ispartof>The Journal of biological chemistry, 2005-12, Vol.280 (51), p.42227-42236</ispartof><rights>2005 © 2005 ASBMB. 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Co-immunoprecipitation and co-localization experiments, as well as combinatorial RNA interference in Caenorhabditis elegans, support the existence of conserved and functional interactions between SRm160 and cohesin.</description><subject>Animals</subject><subject>Antigens, Nuclear - chemistry</subject><subject>Antigens, Nuclear - genetics</subject><subject>Antigens, Nuclear - metabolism</subject><subject>Caenorhabditis elegans</subject><subject>Caenorhabditis elegans - genetics</subject><subject>Cell Cycle Proteins - metabolism</subject><subject>Chromosomal Proteins, Non-Histone</subject><subject>Cohesins</subject><subject>Fungal Proteins - metabolism</subject><subject>HeLa Cells</subject><subject>Humans</subject><subject>Immunoprecipitation</subject><subject>Mass Spectrometry</subject><subject>Nuclear Matrix-Associated Proteins - chemistry</subject><subject>Nuclear Matrix-Associated Proteins - genetics</subject><subject>Nuclear Matrix-Associated Proteins - metabolism</subject><subject>Nuclear Proteins - metabolism</subject><subject>Protein Binding</subject><subject>Proteome</subject><subject>RNA Splicing</subject><subject>RNA, Messenger - genetics</subject><subject>RNA-Binding Proteins - chemistry</subject><subject>RNA-Binding Proteins - genetics</subject><subject>RNA-Binding Proteins - metabolism</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2005</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkE1r3DAQhkVISbZprzkGH0Jv3o5kyZaOy9IvSGlJW-ipQpbHsYItbSTvpvn31bILOYXqMkLzzMvoIeSSwpJCw9_ft3b5VeQbBQZwQhYUZFVWgv4-JQsARkvFhDwnr1O6h3y4omfknNZUKNk0C_LnewwzhsnZYuXN-JRcKkJf_LidaA2lDX42zjt_V6zDtBnxL6biFndoxlSY_OYTxh12xSqlYJ2ZXfDFo5uH3BowOf-GvOozi2-P9YL8-vjh5_pzefPt05f16qa0nLO57Dgowaji2FaKWdmqRlowwvSNFVJyK0HUjAOtDLO2gqaq-7bru0oY1mP-8QV5d8jdxPCwxTTrySWL42g8hm3StVSwV_RfkKoG6qwzg8sDaGNIKWKvN9FNJj5pCnqvXmf1-ll9Hrg6Jm_bCbtn_Og6A9cHYHB3w6OLqFsX7ICTZhK0oJozxvaYPGCYfe0cRp2sQ2-xyyN21l1wL63wD1jinRk</recordid><startdate>20051223</startdate><enddate>20051223</enddate><creator>McCracken, Susan</creator><creator>Longman, Dasa</creator><creator>Marcon, Edyta</creator><creator>Moens, Peter</creator><creator>Downey, Michael</creator><creator>Nickerson, Jeffrey A.</creator><creator>Jessberger, Rolf</creator><creator>Wilde, Andrew</creator><creator>Caceres, Javier F.</creator><creator>Emili, Andrew</creator><creator>Blencowe, Benjamin J.</creator><general>Elsevier Inc</general><general>American Society for Biochemistry and Molecular Biology</general><scope>6I.</scope><scope>AAFTH</scope><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>7TM</scope><scope>7X8</scope></search><sort><creationdate>20051223</creationdate><title>Proteomic Analysis of SRm160-containing Complexes Reveals a Conserved Association with Cohesin</title><author>McCracken, Susan ; 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| source | MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; PubMed Central; Alma/SFX Local Collection |
| subjects | Animals Antigens, Nuclear - chemistry Antigens, Nuclear - genetics Antigens, Nuclear - metabolism Caenorhabditis elegans Caenorhabditis elegans - genetics Cell Cycle Proteins - metabolism Chromosomal Proteins, Non-Histone Cohesins Fungal Proteins - metabolism HeLa Cells Humans Immunoprecipitation Mass Spectrometry Nuclear Matrix-Associated Proteins - chemistry Nuclear Matrix-Associated Proteins - genetics Nuclear Matrix-Associated Proteins - metabolism Nuclear Proteins - metabolism Protein Binding Proteome RNA Splicing RNA, Messenger - genetics RNA-Binding Proteins - chemistry RNA-Binding Proteins - genetics RNA-Binding Proteins - metabolism |
| title | Proteomic Analysis of SRm160-containing Complexes Reveals a Conserved Association with Cohesin |
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