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. 2015 Dec 17:6:346.
doi: 10.3389/fgene.2015.00346. eCollection 2015.

Next-Generation Sequencing for Binary Protein-Protein Interactions

Bernhard Suter  1 , Xinmin Zhang  2 , C Gustavo Pesce  1 , Andrew R Mendelsohn  3 , Savithramma P Dinesh-Kumar  4 , Jian-Hua Mao  5
Affiliations

Next-Generation Sequencing for Binary Protein-Protein Interactions

Bernhard Suter et al. Front Genet. .

Abstract

The yeast two-hybrid (Y2H) system exploits host cell genetics in order to display binary protein-protein interactions (PPIs) via defined and selectable phenotypes. Numerous improvements have been made to this method, adapting the screening principle for diverse applications, including drug discovery and the scale-up for proteome wide interaction screens in human and other organisms. Here we discuss a systematic workflow and analysis scheme for screening data generated by Y2H and related assays that includes high-throughput selection procedures, readout of comprehensive results via next-generation sequencing (NGS), and the interpretation of interaction data via quantitative statistics. The novel assays and tools will serve the broader scientific community to harness the power of NGS technology to address PPI networks in health and disease. We discuss examples of how this next-generation platform can be applied to address specific questions in diverse fields of biology and medicine.

Keywords: interactome mapping; next-generation sequencing; protein–protein interactions; quantitative interaction profiles; yeast two-hybrid.

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Figures

FIGURE 1
FIGURE 1
Pipeline for Next-generation interaction sequencing (NGIS). Specific target binding in Y2H (or related assays) results in distinct populations of cDNAs that are identified and quantified via NGS. Interactions are scored and interpreted in a bioinformatics pipeline with quantitative statistics.
FIGURE 2
FIGURE 2
Screening principle and applications for NGIS. Experimental scheme for NGS based interaction profiling. Bait-specific screening results/profiles are compared to original cDNA pools and control screens to uncover background and non-specific interactions. Prey cDNAs that interact with the bait are enriched and quantitated using next-generation sequencing (NGS) and bioinformatics analysis. Bait-specific enrichments (blue) can be quantitatively distinguished from non-specific enrichments (red) and non-selected preys (orange, green).
See this image and copyright information in PMC

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