Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS
Receptor for Activated protein C kinase 1 (RACK1) is a scaffold protein that has been found in association with several signaling complexes, and with the 40S subunit of the ribosome. Using the model organism Drosophila melanogaster, we recently showed that RACK1 is required at the ribosome for inter...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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Oxford University Press
2017-07-01
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| Series: | G3: Genes, Genomes, Genetics |
| Subjects: | |
| Online Access: | http://g3journal.org/lookup/doi/10.1534/g3.117.042564 |
| _version_ | 1818571634356781056 |
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| author | Lauriane Kuhn Karim Majzoub Evelyne Einhorn Johana Chicher Julien Pompon Jean-Luc Imler Philippe Hammann Carine Meignin |
| author_facet | Lauriane Kuhn Karim Majzoub Evelyne Einhorn Johana Chicher Julien Pompon Jean-Luc Imler Philippe Hammann Carine Meignin |
| author_sort | Lauriane Kuhn |
| collection | DOAJ |
| description | Receptor for Activated protein C kinase 1 (RACK1) is a scaffold protein that has been found in association with several signaling complexes, and with the 40S subunit of the ribosome. Using the model organism Drosophila melanogaster, we recently showed that RACK1 is required at the ribosome for internal ribosome entry site (IRES)-mediated translation of viruses. Here, we report a proteomic characterization of the interactome of RACK1 in Drosophila S2 cells. We carried out Label-Free quantitation using both Data-Dependent and Data-Independent Acquisition (DDA and DIA, respectively) and observed a significant advantage for the Sequential Window Acquisition of all THeoretical fragment-ion spectra (SWATH) method, both in terms of identification of interactants and quantification of low abundance proteins. These data represent the first SWATH spectral library available for Drosophila and will be a useful resource for the community. A total of 52 interacting proteins were identified, including several molecules involved in translation such as structural components of the ribosome, factors regulating translation initiation or elongation, and RNA binding proteins. Among these 52 proteins, 15 were identified as partners by the SWATH strategy only. Interestingly, these 15 proteins are significantly enriched for the functions translation and nucleic acid binding. This enrichment reflects the engagement of RACK1 at the ribosome and highlights the added value of SWATH analysis. A functional screen did not reveal any protein sharing the interesting properties of RACK1, which is required for IRES-dependent translation and not essential for cell viability. Intriguingly however, 10 of the RACK1 partners identified restrict replication of Cricket paralysis virus (CrPV), an IRES-containing virus. |
| first_indexed | 2024-12-14T13:58:38Z |
| format | Article |
| id | doaj.art-853b068f78b7454ca409975f3530513e |
| institution | Directory Open Access Journal |
| issn | 2160-1836 |
| language | English |
| last_indexed | 2024-12-14T13:58:38Z |
| publishDate | 2017-07-01 |
| publisher | Oxford University Press |
| record_format | Article |
| series | G3: Genes, Genomes, Genetics |
| spelling | doaj.art-853b068f78b7454ca409975f3530513e2022-12-21T22:58:47ZengOxford University PressG3: Genes, Genomes, Genetics2160-18362017-07-01772249225810.1534/g3.117.04256422Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MSLauriane KuhnKarim MajzoubEvelyne EinhornJohana ChicherJulien PomponJean-Luc ImlerPhilippe HammannCarine MeigninReceptor for Activated protein C kinase 1 (RACK1) is a scaffold protein that has been found in association with several signaling complexes, and with the 40S subunit of the ribosome. Using the model organism Drosophila melanogaster, we recently showed that RACK1 is required at the ribosome for internal ribosome entry site (IRES)-mediated translation of viruses. Here, we report a proteomic characterization of the interactome of RACK1 in Drosophila S2 cells. We carried out Label-Free quantitation using both Data-Dependent and Data-Independent Acquisition (DDA and DIA, respectively) and observed a significant advantage for the Sequential Window Acquisition of all THeoretical fragment-ion spectra (SWATH) method, both in terms of identification of interactants and quantification of low abundance proteins. These data represent the first SWATH spectral library available for Drosophila and will be a useful resource for the community. A total of 52 interacting proteins were identified, including several molecules involved in translation such as structural components of the ribosome, factors regulating translation initiation or elongation, and RNA binding proteins. Among these 52 proteins, 15 were identified as partners by the SWATH strategy only. Interestingly, these 15 proteins are significantly enriched for the functions translation and nucleic acid binding. This enrichment reflects the engagement of RACK1 at the ribosome and highlights the added value of SWATH analysis. A functional screen did not reveal any protein sharing the interesting properties of RACK1, which is required for IRES-dependent translation and not essential for cell viability. Intriguingly however, 10 of the RACK1 partners identified restrict replication of Cricket paralysis virus (CrPV), an IRES-containing virus.http://g3journal.org/lookup/doi/10.1534/g3.117.042564Drosophila melanogastermass spectrometrytranslationRACK1ribosomevirusIRESLarkAGO2 |
| spellingShingle | Lauriane Kuhn Karim Majzoub Evelyne Einhorn Johana Chicher Julien Pompon Jean-Luc Imler Philippe Hammann Carine Meignin Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS G3: Genes, Genomes, Genetics Drosophila melanogaster mass spectrometry translation RACK1 ribosome virus IRES Lark AGO2 |
| title | Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS |
| title_full | Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS |
| title_fullStr | Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS |
| title_full_unstemmed | Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS |
| title_short | Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS |
| title_sort | definition of a rack1 interaction network in drosophila melanogaster using swath ms |
| topic | Drosophila melanogaster mass spectrometry translation RACK1 ribosome virus IRES Lark AGO2 |
| url | http://g3journal.org/lookup/doi/10.1534/g3.117.042564 |
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