A High Throughput Protoplast System for Rice Functional Genomics and Proteomics: Protein-Protein Interactions at the Host-Pathogen Interface

点击翻译按钮获取中文摘要

A High Throughput Protoplast System for Rice Functional Genomics and Proteomics: Protein-Protein Interactions at the Host-Pathogen InterfaceRice is the food staple for over half the world's population. Rice blast disease, caused by the fungal pathogen Magnaporthe grisea, is highly devastating and persists as a major threat to global food security. The ability of host and pathogen to detect and respond to each other's presence is governed by protein interactions, however knowledge of these interactions remains fragmentary at best. Access to near complete genomes of both rice and M. grisea, along with the extensive biological resources and a rich history of investigation, provides the opportunity to investigate interactions between host and pathogen. This research project will develop and test a novel system based on the reassembly of dissected fragments of light/fluorescence generating proteins fused to associating fungal and/or rice peptides to directly visualize protein interactions in living plant cells. Rice protoplasts (cells devoid of their cell wall) will be using since they are easy to manipulate, are amenable to large-scale analyses and typically mimic whole plants. As a proof-of-concept, the system will be first tested using rice proteins that are known to interact. The applicability of the system to interrogate the interaction of proteins from rice with those of the rice blast fungus will be subsequently tested using a known defense elicitor (avirulence) protein and its cognate rice resistance receptor. The system will be finally scaled up to screen for unknown fungal and plant protein associations that are predicted to control the deployment of host defenses. The high throughput protoplast system will be also applied in a feasibility study to screen for unknown fungal elicitors of plant host defense responses, of which a select few will advance into the novel screen developed to identify interacting plant proteins. An extensive education plan for high school students and activities promoting recruitment and participation by members of underrepresented groups are planned. In addition to furthering our basic understanding of plant-pathogen interactions, the technologies to be developed will have far-reaching utility for the discovery and evaluation of other protein-protein or protein-ligand interactions in plants systems outside rice and rice blast. Data from this project will be made available in refereed publications. Data, protocols, and reagents will also be made avaiable via www.fsgc.net (Fungal Genetics Stock Center) and www.mgosdb.org (Magnaportha grisea Oriza sativa database) as they are generated and qualities checked.