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Title: Quantitative Interactor Screening with next-generation Sequencing (QIS-Seq) identifies Arabidopsis thaliana MLO2 as a target of the Psuedomonas syringe type III effector HopZ2

Author
item LEWIS, J - University Of Toronto
item WAN, J - University Of Toronto
item FORD, R - University Of Toronto
item GONG, Y - University Of Toronto
item FUNG, P - University Of Toronto
item WANG, P - University Of Toronto
item DESVEAUX, D - University Of Toronto
item GUTTMAN, D - University Of Toronto

Submitted to: BMC Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/9/2012
Publication Date: 1/20/2012
Citation: Lewis, J.D., Wan, J., Ford, R., Gong, Y., Fung, P., Wang, P.W., Desveaux, D., Guttman, D.S. 2012. Quantitative Interactor Screening with next-generation Sequencing (QIS-Seq) identifies Arabidopsis thaliana MLO2 as a target of the Psuedomonas syringe type III effector HopZ2. Biomed Central (BMC) Genomics. 13:8.

Interpretive Summary: Bacterial diseases cause substantial crop losses. Bacteria use the type III secretion system to inject type III secreted effectors into plants. Type III effector proteins are a primary mechanism for causing disease, as they can suppress defense responses in the plant. The YopJ superfamily of type III effector proteins is evolutionarily conserved and found in both animal and plant pathogens. We briefly review the literature on YopJ, and synthesize the literature on members of the YopJ superfamily in plant-associated bacteria. We discuss the functions of each homolog, their similarities and their differences. Analysis of the YopJ superfamily provides a unique opportunity to understand type III effector function within its evolutionary context.

Technical Abstract: Background: Identification of protein-protein interactions is a fundamental aspect of understanding protein function. A commonly used method for identifying protein interactions is the yeast two-hybrid system. Results: Here we describe the application of next-generation sequencing to yeast two-hybrid interaction screens and develop Quantitative Interactor Screen Sequencing (QIS-Seq). QIS-Seq provides a quantitative measurement of enrichment for each interactor relative to its frequency in the library as well as its general stickiness (non-specific binding). The QIS-Seq approach is scalable and can be used with any yeast two-hybrid screen and with any next-generation sequencing platform. The quantitative nature of QIS-Seq data makes it amenable to statistical evaluation, and importantly, facilitates the standardization of experimental design, data collection, and data analysis. We applied QIS-Seq to identify the Arabidopsis thaliana MLO2 protein as a target of the Pseudomonas syringae type III secreted effector protein HopZ2. We validate the interaction between HopZ2 and MLO2 in planta and show that the interaction is required for HopZ2-associated virulence. Conclusions: We demonstrate that QIS-Seq is a high-throughput quantitative interactor screen and validate MLO2 as an interactor and novel virulence target of the P. syringae type III secreted effector HopZ2.