Localization of secreted proteins during penetration and invasive growth of the rice blast fungus Magnaporthe oryzae

Plant pathogenic fungi secrete many proteins with diverse roles in plant-microbe interactions. However, information on the temporal and spatial patterns of protein secretion during fungal infection of plants is extremely limited, so in most cases it is not clear which proteins are expressed/secreted at different infection stages. Likewise, it is not known if secretion occurs at specific locations within invasive hyphae, and if secreted proteins are targeted to specific regions of the pathogen-host interface. The goal of this project is to gain a comprehensive insight into protein secretion by Magnaporthe oryzae during infection of rice. We will accomplish this by creating GFP fusions to each of the 939 secreted proteins predicted in the genome sequence. Transformants carrying gene fusions will be inoculated onto leaf sheaths and protein localization during penetration and invasive growth will be determined by live-cell imaging of infection sites using epifluorescence and confocal microscopy. The locations of selected proteins will be validated by immunodetection with specific antibodies. This project will be facilitated by the development of novel vectors for high throughput production of auto fluorescent protein fusions in fungi, and by managing the workflow and data acquisition with a Laboratory Information Management System designed for a similar project. The data generated will be deposited in the MGOS database for access by the broader research community. Knowledge gained in this project will further understanding in many areas of plant microbe interactioris, which in turn will lead to the development of new ways to protect the Nation's food supply.