CrY2H-seq: A massively multiplexed assay for deep-coverage interactome mapping

Shelly A. Trigg, Renee M. Garza, Andrew MacWilliams, Joseph R. Nery, Anna Bartlett, Rosa Castanon, Adeline Goubil, Joseph Feeney, Ronan O'Malley, Shao Shan C. Huang, Zhuzhu Z. Zhang, Mary Galli, Joseph R. Ecker

Research output: Contribution to journalArticle

Abstract

Broad-scale protein-protein interaction mapping is a major challenge given the cost, time, and sensitivity constraints of existing technologies. Here, we present a massively multiplexed yeast two-hybrid method, CrY2H-seq, which uses a Cre recombinase interaction reporter to intracellularly fuse the coding sequences of two interacting proteins and next-generation DNA sequencing to identify these interactions en masse. We applied CrY2H-seq to investigate sparsely annotated Arabidopsis thaliana transcription factors interactions. By performing ten independent screens testing a total of 36 million binary interaction combinations, and uncovering a network of 8,577 interactions among 1,453 transcription factors, we demonstrate CrY2H-seq?s improved screening capacity, efficiency, and sensitivity over those of existing technologies. The deep-coverage network resource we call AtTFIN-1 recapitulates one-third of previously reported interactions derived from diverse methods, expands the number of known plant transcription factor interactions by three-fold, and reveals previously unknown family-specific interaction module associations with plant reproductive development, root architecture, and circadian coordination.

Original languageEnglish (US)
Pages (from-to)819-825
Number of pages7
JournalNature methods
Volume14
Issue number8
DOIs
StatePublished - Jul 28 2017

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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  • Cite this

    Trigg, S. A., Garza, R. M., MacWilliams, A., Nery, J. R., Bartlett, A., Castanon, R., Goubil, A., Feeney, J., O'Malley, R., Huang, S. S. C., Zhang, Z. Z., Galli, M., & Ecker, J. R. (2017). CrY2H-seq: A massively multiplexed assay for deep-coverage interactome mapping. Nature methods, 14(8), 819-825. https://doi.org/10.1038/nmeth.4343