rec-YnH enables simultaneous many-by-many detection of direct protein-protein and protein-RNA interactions

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  • dc.contributor.author Yang, Jae-Seong
  • dc.contributor.author Garriga-Canut, Mireia
  • dc.contributor.author Link, Nele
  • dc.contributor.author Carolis, Carlo
  • dc.contributor.author Broadbent, Katrina
  • dc.contributor.author Beltran-Sastre, Violeta
  • dc.contributor.author Serrano Pubull, Luis, 1982-
  • dc.contributor.author Maurer, Sebastian
  • dc.date.accessioned 2019-11-25T08:32:22Z
  • dc.date.available 2019-11-25T08:32:22Z
  • dc.date.issued 2018
  • dc.description.abstract Knowing which proteins and RNAs directly interact is essential for understanding cellular mechanisms. Unfortunately, discovering such interactions is costly and often unreliable. To overcome these limitations, we developed rec-YnH, a new yeast two and three-hybrid-based screening pipeline capable of detecting interactions within protein libraries or between protein libraries and RNA fragment pools. rec-YnH combines batch cloning and transformation with intracellular homologous recombination to generate bait–prey fusion libraries. By developing interaction selection in liquid–gels and using an ORF sequence-based readout of interactions via next-generation sequencing, we eliminate laborious plating and barcoding steps required by existing methods. We use rec-Y2H to simultaneously map interactions of protein domains and reveal novel putative interactors of PAR proteins. We further employ rec-Y2H to predict the architecture of published coprecipitated complexes. Finally, we use rec-Y3H to map interactions between multiple RNA-binding proteins and RNAs—the first time interactions between protein and RNA pools are simultaneously detected.
  • dc.description.sponsorship We also acknowledge the support from the Spanish Ministry of Economy and Competitiveness (MINECO) for Juan de la Cierva-Incorporación Programme (IJCI‐2014‐22070) to J.-S.Y., L.S. (BFU2015-63571-P), and S.M. (BFU2014-54278-P and BFU2015-62550-ERC). We further acknowledge support of the Spanish Ministry of Economy and Competitiveness, “Centro de Excelencia Severo Ochoa 2013-2017”, SEV-2012-0208 and the CERCA Programme/Generalitat de Catalunya. This work was funded by the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) reference MINECO PE 2013-2016 PN FEDER and the European Regional Development Fund (ERDF). All sequencing was done in the CRG Genomics Core Facility.
  • dc.format.mimetype application/pdf
  • dc.identifier.citation Yang JS, Garriga-Canut M, Link N, Carolis C, Broadbent K, Beltran-Sastre V et al. rec-YnH enables simultaneous many-by-many detection of direct protein-protein and protein-RNA interactions. Nat Commun. 2018;9(1):3747. DOI: 10.1038/s41467-018-06128-x
  • dc.identifier.doi http://dx.doi.org/10.1038/s41467-018-06128-x
  • dc.identifier.issn 2041-1723
  • dc.identifier.uri http://hdl.handle.net/10230/42955
  • dc.language.iso eng
  • dc.publisher Nature Research
  • dc.relation.ispartof Nature Communications. 2018;9(1):3747
  • dc.relation.projectID info:eu-repo/grantAgreement/ES/1PE/BFU2015-63571-P
  • dc.relation.projectID info:eu-repo/grantAgreement/ES/1PE/BFU2014-54278-P
  • dc.relation.projectID info:eu-repo/grantAgreement/ES/1PE/BFU2015-62550-ERC
  • dc.rights © The Author(s) 2018. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
  • dc.rights.accessRights info:eu-repo/semantics/openAccess
  • dc.rights.uri http://creativecommons.org/licenses/by/4.0/
  • dc.subject.keyword Assay systems
  • dc.subject.keyword High-throughput screening
  • dc.title rec-YnH enables simultaneous many-by-many detection of direct protein-protein and protein-RNA interactions
  • dc.type info:eu-repo/semantics/article
  • dc.type.version info:eu-repo/semantics/publishedVersion

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