Open AccessCrY2H-seq: a massively multiplexed assay for deep-coverage interactome mapping
Author(s) -
Shelly A. Trigg,
Renee M. Garza,
Andrew MacWilliams,
Joseph R. Nery,
Anna Bartlett,
Rosa Casta,
Adeline Goubil,
Joseph Feeney,
Ronan O’Malley,
Shaoshan Carol Huang,
Zhuzhu Z. Zhang,
Mary Galli,
Joseph R. Ecker
Publication year - 2017
Publication title -
nature methods
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 19.469
H-Index - 318
eISSN - 1548-7105
pISSN - 1548-7091
DOI - 10.1038/nmeth.4343
Subject(s) - interactome , computational biology , biology , transcription factor , protein–protein interaction , interaction network , computer science , genetics , gene
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.