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

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

doi:10.1038/NMETH.4343

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

Journal Article · Mon Jun 26 00:00:00 EDT 2017 · Nature Methods

DOI:https://doi.org/10.1038/NMETH.4343· OSTI ID:1473848

Trigg, Shelly A. ^[1]; Garza, Renee M. ^[2]; MacWilliams, Andrew ^[2]; Nery, Joseph R. ^[2]; Bartlett, Anna ^[2]; Castanon, Rosa ^[2]; Goubil, Adeline ^[3]; Feeney, Joseph ^[3]; O'Malley, Ronan ^[3]; Huang, Shao-shan Carol ^[2]; Zhang, Zhuzhu Z. ^[2]; Galli, Mary ^[3]; Ecker, Joseph R. ^[4]

Salk Inst. for Biological Studies, La Jolla, CA (United States). Genomic Analysis and Plant Biology Lab.; Univ. of California, San Diego, CA (United States). Division of Biological Sciences
Salk Inst. for Biological Studies, La Jolla, CA (United States). Genomic Analysis and Plant Biology Lab.
Salk Inst. for Biological Studies, La Jolla, CA (United States). Genomic Analysis and Plant Biology Lab.; National Inst. for Agricultural Research, Paris (France)
Salk Inst. for Biological Studies, La Jolla, CA (United States). Genomic Analysis and Plant Biology Lab., and Howard Hughes Medical Inst.; Univ. of California, San Diego, CA (United States). Division of Biological Sciences

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.

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Research Organization:: Salk Inst. for Biological Studies, La Jolla, CA (United States)

Sponsoring Organization:: USDOE; National Science Foundation (NSF)

Grant/Contract Number:: SC0007078; IOS-1650227; IOS1456950; IOS1546873; DGE-1650112

OSTI ID:: 1473848

Journal Information:: Nature Methods, Vol. 14, Issue 8; ISSN 1548-7091

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 101 works

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