A scalable double-barcode sequencing platform for characterization of dynamic protein-protein interactions.
A scalable double-barcode sequencing platform for characterization of dynamic protein-protein interactions.
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DOI：
10.1038/ncomms15586
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发表时间：
2017-05-25
影响因子：
16.6
通讯作者：
Levy SF
中科院分区：
文献类型：
--
作者：
Schlecht U;Liu Z;Blundell JR;St Onge RP;Levy SF
Several large-scale efforts have systematically catalogued protein-protein interactions (PPIs) of a cell in a single environment. However, little is known about how the protein interactome changes across environmental perturbations. Current technologies, which assay one PPI at a time, are too low throughput to make it practical to study protein interactome dynamics. Here, we develop a highly parallel protein-protein interaction sequencing (PPiSeq) platform that uses a novel double barcoding system in conjunction with the dihydrofolate reductase protein-fragment complementation assay in Saccharomyces cerevisiae. PPiSeq detects PPIs at a rate that is on par with current assays and, in contrast with current methods, quantitatively scores PPIs with enough accuracy and sensitivity to detect changes across environments. Both PPI scoring and the bulk of strain construction can be performed with cell pools, making the assay scalable and easily reproduced across environments. PPiSeq is therefore a powerful new tool for large-scale investigations of dynamic PPIs. Protein-protein interactions (PPIs) play a major role in defining biological functions. Here, the authors present PPiSeq, a method to quantitatively score dynamic PPIs in yeast combining fragment complementation assays, genomic double-barcoding, and an analytical framework to precisely call fitness from barcode lineage trajectories.
