Differential 14-3-3 Affinity Capture Reveals New Downstream Targets of Phosphatidylinositol 3-Kinase Signaling
We devised a strategy of 14-3-3 affinity capture and release, isotope differential (d0/d4) dimethyl labeling of tryptic digests, and phosphopeptide characterization to identify novel targets of insulin/IGF1/phosphatidylinositol 3-kinase signaling. Notably four known insulin-regulated proteins (PFK-2...
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| Veröffentlicht in: | Molecular & cellular proteomics 2009-11, Vol.8 (11), p.2487-2499 |
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| container_title | Molecular & cellular proteomics |
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| creator | Dubois, Fanny Vandermoere, Franck Gernez, Aurélie Murphy, Jane Toth, Rachel Chen, Shuai Geraghty, Kathryn M. Morrice, Nick A. MacKintosh, Carol |
| description | We devised a strategy of 14-3-3 affinity capture and release, isotope differential (d0/d4) dimethyl labeling of tryptic digests, and phosphopeptide characterization to identify novel targets of insulin/IGF1/phosphatidylinositol 3-kinase signaling. Notably four known insulin-regulated proteins (PFK-2, PRAS40, AS160, and MYO1C) had high d0/d4 values meaning that they were more highly represented among 14-3-3-binding proteins from insulin-stimulated than unstimulated cells. Among novel candidates, insulin receptor substrate 2, the proapoptotic CCDC6, E3 ubiquitin ligase ZNRF2, and signaling adapter SASH1 were confirmed to bind to 14-3-3s in response to IGF1/phosphatidylinositol 3-kinase signaling. Insulin receptor substrate 2, ZNRF2, and SASH1 were also regulated by phorbol ester via p90RSK, whereas CCDC6 and PRAS40 were not. In contrast, the actin-associated protein vasodilator-stimulated phosphoprotein and lipolysis-stimulated lipoprotein receptor, which had low d0/d4 scores, bound 14-3-3s irrespective of IGF1 and phorbol ester. Phosphorylated Ser19 of ZNRF2 (RTRAYpS19GS), phospho-Ser90 of SASH1 (RKRRVpS90QD), and phospho- Ser493 of lipolysis-stimulated lipoprotein receptor (RPRARpS493LD) provide one of the 14-3-3-binding sites on each of these proteins. Differential 14-3-3 capture provides a powerful approach to defining downstream regulatory mechanisms for specific signaling pathways. |
| doi_str_mv | 10.1074/mcp.M800544-MCP200 |
| format | Article |
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Notably four known insulin-regulated proteins (PFK-2, PRAS40, AS160, and MYO1C) had high d0/d4 values meaning that they were more highly represented among 14-3-3-binding proteins from insulin-stimulated than unstimulated cells. Among novel candidates, insulin receptor substrate 2, the proapoptotic CCDC6, E3 ubiquitin ligase ZNRF2, and signaling adapter SASH1 were confirmed to bind to 14-3-3s in response to IGF1/phosphatidylinositol 3-kinase signaling. Insulin receptor substrate 2, ZNRF2, and SASH1 were also regulated by phorbol ester via p90RSK, whereas CCDC6 and PRAS40 were not. In contrast, the actin-associated protein vasodilator-stimulated phosphoprotein and lipolysis-stimulated lipoprotein receptor, which had low d0/d4 scores, bound 14-3-3s irrespective of IGF1 and phorbol ester. Phosphorylated Ser19 of ZNRF2 (RTRAYpS19GS), phospho-Ser90 of SASH1 (RKRRVpS90QD), and phospho- Ser493 of lipolysis-stimulated lipoprotein receptor (RPRARpS493LD) provide one of the 14-3-3-binding sites on each of these proteins. Differential 14-3-3 capture provides a powerful approach to defining downstream regulatory mechanisms for specific signaling pathways.</description><identifier>ISSN: 1535-9476</identifier><identifier>EISSN: 1535-9484</identifier><identifier>DOI: 10.1074/mcp.M800544-MCP200</identifier><identifier>PMID: 19648646</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>14-3-3 Proteins - metabolism ; Actins - chemistry ; Apoptosis ; Binding Sites ; Biochemistry, Molecular Biology ; Chromatography, Liquid - methods ; Genomics ; HeLa Cells ; Humans ; Life Sciences ; Mass Spectrometry - methods ; Models, Biological ; Peptides - chemistry ; Phosphatidylinositol 3-Kinases - metabolism ; Proteome ; Proteomics - methods ; Signal Transduction ; Trypsin - chemistry</subject><ispartof>Molecular & cellular proteomics, 2009-11, Vol.8 (11), p.2487-2499</ispartof><rights>2009 © 2009 ASBMB. Currently published by Elsevier Inc; originally published by American Society for Biochemistry and Molecular Biology.</rights><rights>Distributed under a Creative Commons Attribution 4.0 International License</rights><rights>2009 by The American Society for Biochemistry and Molecular Biology, Inc.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c580t-6781c8d3f9983338057a563dabb605dbc4b1b25518595dca83bf7bb50c1fdc53</citedby><cites>FETCH-LOGICAL-c580t-6781c8d3f9983338057a563dabb605dbc4b1b25518595dca83bf7bb50c1fdc53</cites><orcidid>0000-0001-5724-7037</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC2773716/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC2773716/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,881,27901,27902,53766,53768</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/19648646$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://hal.science/hal-03030169$$DView record in HAL$$Hfree_for_read</backlink></links><search><creatorcontrib>Dubois, Fanny</creatorcontrib><creatorcontrib>Vandermoere, Franck</creatorcontrib><creatorcontrib>Gernez, Aurélie</creatorcontrib><creatorcontrib>Murphy, Jane</creatorcontrib><creatorcontrib>Toth, Rachel</creatorcontrib><creatorcontrib>Chen, Shuai</creatorcontrib><creatorcontrib>Geraghty, Kathryn M.</creatorcontrib><creatorcontrib>Morrice, Nick A.</creatorcontrib><creatorcontrib>MacKintosh, Carol</creatorcontrib><title>Differential 14-3-3 Affinity Capture Reveals New Downstream Targets of Phosphatidylinositol 3-Kinase Signaling</title><title>Molecular & cellular proteomics</title><addtitle>Mol Cell Proteomics</addtitle><description>We devised a strategy of 14-3-3 affinity capture and release, isotope differential (d0/d4) dimethyl labeling of tryptic digests, and phosphopeptide characterization to identify novel targets of insulin/IGF1/phosphatidylinositol 3-kinase signaling. Notably four known insulin-regulated proteins (PFK-2, PRAS40, AS160, and MYO1C) had high d0/d4 values meaning that they were more highly represented among 14-3-3-binding proteins from insulin-stimulated than unstimulated cells. Among novel candidates, insulin receptor substrate 2, the proapoptotic CCDC6, E3 ubiquitin ligase ZNRF2, and signaling adapter SASH1 were confirmed to bind to 14-3-3s in response to IGF1/phosphatidylinositol 3-kinase signaling. Insulin receptor substrate 2, ZNRF2, and SASH1 were also regulated by phorbol ester via p90RSK, whereas CCDC6 and PRAS40 were not. In contrast, the actin-associated protein vasodilator-stimulated phosphoprotein and lipolysis-stimulated lipoprotein receptor, which had low d0/d4 scores, bound 14-3-3s irrespective of IGF1 and phorbol ester. Phosphorylated Ser19 of ZNRF2 (RTRAYpS19GS), phospho-Ser90 of SASH1 (RKRRVpS90QD), and phospho- Ser493 of lipolysis-stimulated lipoprotein receptor (RPRARpS493LD) provide one of the 14-3-3-binding sites on each of these proteins. Differential 14-3-3 capture provides a powerful approach to defining downstream regulatory mechanisms for specific signaling pathways.</description><subject>14-3-3 Proteins - metabolism</subject><subject>Actins - chemistry</subject><subject>Apoptosis</subject><subject>Binding Sites</subject><subject>Biochemistry, Molecular Biology</subject><subject>Chromatography, Liquid - methods</subject><subject>Genomics</subject><subject>HeLa Cells</subject><subject>Humans</subject><subject>Life Sciences</subject><subject>Mass Spectrometry - methods</subject><subject>Models, Biological</subject><subject>Peptides - chemistry</subject><subject>Phosphatidylinositol 3-Kinases - metabolism</subject><subject>Proteome</subject><subject>Proteomics - methods</subject><subject>Signal Transduction</subject><subject>Trypsin - chemistry</subject><issn>1535-9476</issn><issn>1535-9484</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kV-P1CAUxRujcdfVL-CDwSfjQ1coUGhiTCazrmuc1Y3OO6H0tsW0pUJnJvPtl0mb9c-D4QFy-Z3DvZwkeUnwJcGCvevNeHkrMeaMpbfruwzjR8k54ZSnBZPs8cNZ5GfJsxB-YpxhIvjT5IwUOZM5y8-T4crWNXgYJqs7RFhKU4pWdW0HOx3RWo_TzgP6DnvQXUBf4YCu3GEIkwfdo632DUwBuRrdtS6MrZ5sdezs4IKdXIdo-sUOOgD6YZtBx3rzPHlSRyN4sewXyfb643Z9k26-ffq8Xm1SwyWe0lxIYmRF66KQlFKJudA8p5UuyxzzqjSsJGXGOZG84JXRkpa1KEuODakrw-lF8mG2HXdlD5WJ43ndqdHbXvujctqqv28G26rG7VUmBBUkjwZvZ4P2H9nNaqNONUzjInmxJ5F9szzm3a8dhEn1NhjoOj2A2wUlKIsg5yfXbCaNdyF4qB-sCVanSFWMVC2RqjnSKHr15yy_JUuGEXi99Gqb9mA9qNI600KvpCJEZUyKyLyfGYifvrfgVTAWBgNV5M2kKmf_18M9QPC9vg</recordid><startdate>20091101</startdate><enddate>20091101</enddate><creator>Dubois, Fanny</creator><creator>Vandermoere, Franck</creator><creator>Gernez, Aurélie</creator><creator>Murphy, Jane</creator><creator>Toth, Rachel</creator><creator>Chen, Shuai</creator><creator>Geraghty, Kathryn M.</creator><creator>Morrice, Nick A.</creator><creator>MacKintosh, Carol</creator><general>Elsevier Inc</general><general>American Society for Biochemistry and Molecular Biology</general><general>The 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>7X8</scope><scope>1XC</scope><scope>VOOES</scope><scope>5PM</scope><orcidid>https://orcid.org/0000-0001-5724-7037</orcidid></search><sort><creationdate>20091101</creationdate><title>Differential 14-3-3 Affinity Capture Reveals New Downstream Targets of Phosphatidylinositol 3-Kinase Signaling</title><author>Dubois, Fanny ; Vandermoere, Franck ; Gernez, Aurélie ; Murphy, Jane ; Toth, Rachel ; Chen, Shuai ; Geraghty, Kathryn M. ; Morrice, Nick A. ; MacKintosh, Carol</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c580t-6781c8d3f9983338057a563dabb605dbc4b1b25518595dca83bf7bb50c1fdc53</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>14-3-3 Proteins - metabolism</topic><topic>Actins - chemistry</topic><topic>Apoptosis</topic><topic>Binding Sites</topic><topic>Biochemistry, Molecular Biology</topic><topic>Chromatography, Liquid - methods</topic><topic>Genomics</topic><topic>HeLa Cells</topic><topic>Humans</topic><topic>Life Sciences</topic><topic>Mass Spectrometry - methods</topic><topic>Models, Biological</topic><topic>Peptides - chemistry</topic><topic>Phosphatidylinositol 3-Kinases - metabolism</topic><topic>Proteome</topic><topic>Proteomics - methods</topic><topic>Signal Transduction</topic><topic>Trypsin - chemistry</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Dubois, Fanny</creatorcontrib><creatorcontrib>Vandermoere, Franck</creatorcontrib><creatorcontrib>Gernez, Aurélie</creatorcontrib><creatorcontrib>Murphy, Jane</creatorcontrib><creatorcontrib>Toth, Rachel</creatorcontrib><creatorcontrib>Chen, Shuai</creatorcontrib><creatorcontrib>Geraghty, Kathryn M.</creatorcontrib><creatorcontrib>Morrice, Nick A.</creatorcontrib><creatorcontrib>MacKintosh, Carol</creatorcontrib><collection>ScienceDirect Open Access Titles</collection><collection>Elsevier:ScienceDirect:Open Access</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>Hyper Article en Ligne (HAL)</collection><collection>Hyper Article en Ligne (HAL) (Open Access)</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Molecular & cellular proteomics</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Dubois, Fanny</au><au>Vandermoere, Franck</au><au>Gernez, Aurélie</au><au>Murphy, Jane</au><au>Toth, Rachel</au><au>Chen, Shuai</au><au>Geraghty, Kathryn M.</au><au>Morrice, Nick A.</au><au>MacKintosh, Carol</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Differential 14-3-3 Affinity Capture Reveals New Downstream Targets of Phosphatidylinositol 3-Kinase Signaling</atitle><jtitle>Molecular & cellular proteomics</jtitle><addtitle>Mol Cell Proteomics</addtitle><date>2009-11-01</date><risdate>2009</risdate><volume>8</volume><issue>11</issue><spage>2487</spage><epage>2499</epage><pages>2487-2499</pages><issn>1535-9476</issn><eissn>1535-9484</eissn><abstract>We devised a strategy of 14-3-3 affinity capture and release, isotope differential (d0/d4) dimethyl labeling of tryptic digests, and phosphopeptide characterization to identify novel targets of insulin/IGF1/phosphatidylinositol 3-kinase signaling. Notably four known insulin-regulated proteins (PFK-2, PRAS40, AS160, and MYO1C) had high d0/d4 values meaning that they were more highly represented among 14-3-3-binding proteins from insulin-stimulated than unstimulated cells. Among novel candidates, insulin receptor substrate 2, the proapoptotic CCDC6, E3 ubiquitin ligase ZNRF2, and signaling adapter SASH1 were confirmed to bind to 14-3-3s in response to IGF1/phosphatidylinositol 3-kinase signaling. Insulin receptor substrate 2, ZNRF2, and SASH1 were also regulated by phorbol ester via p90RSK, whereas CCDC6 and PRAS40 were not. In contrast, the actin-associated protein vasodilator-stimulated phosphoprotein and lipolysis-stimulated lipoprotein receptor, which had low d0/d4 scores, bound 14-3-3s irrespective of IGF1 and phorbol ester. Phosphorylated Ser19 of ZNRF2 (RTRAYpS19GS), phospho-Ser90 of SASH1 (RKRRVpS90QD), and phospho- Ser493 of lipolysis-stimulated lipoprotein receptor (RPRARpS493LD) provide one of the 14-3-3-binding sites on each of these proteins. Differential 14-3-3 capture provides a powerful approach to defining downstream regulatory mechanisms for specific signaling pathways.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>19648646</pmid><doi>10.1074/mcp.M800544-MCP200</doi><tpages>13</tpages><orcidid>https://orcid.org/0000-0001-5724-7037</orcidid><oa>free_for_read</oa></addata></record> |
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| subjects | 14-3-3 Proteins - metabolism Actins - chemistry Apoptosis Binding Sites Biochemistry, Molecular Biology Chromatography, Liquid - methods Genomics HeLa Cells Humans Life Sciences Mass Spectrometry - methods Models, Biological Peptides - chemistry Phosphatidylinositol 3-Kinases - metabolism Proteome Proteomics - methods Signal Transduction Trypsin - chemistry |
| title | Differential 14-3-3 Affinity Capture Reveals New Downstream Targets of Phosphatidylinositol 3-Kinase Signaling |
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