Role of Extracellular Vesicles in Plant-Pathogen Interactions

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Plant diseases caused by viruses, bacteria, and fungi cause major economic harm to U.S. farmers and greatly limit agricultural production worldwide. In developing regions such as sub-Saharan Africa and India, the combination of rapid population growth and declining agricultural production is a major threat to food security, and hence to political stability. There is thus an urgent need to improve the immune system of plants so that they can defend themselves against disease. This project is focused on understanding an underappreciated component of the plant immune system that appears to be shared with humans, extracellular vesicles (EVs). EVs are microscopic spherical packages that can carry information from one cell to another within an organism. Recent work in animal systems, including humans, has revealed that this information can reprogram recipient cells so that the receiving cell changes the proteins it produces, and hence the properties of the cell, and in some cases, the ability of a cell to prevent infection by a virus. Recent work in the Innes laboratory indicates that EVs may serve a similar role in plants. This research project aims to address basic questions about how plant EVs are produced, what they carry, and how they may contribute to plant immunity. The answers to these questions will facilitate development of disease resistant crops. In addition, this project will provide hands on plant science activities to elementary, high school and undergraduate students, with a special focus on underrepresented minorities. This project focuses on the genesis and roles of extracellular vesicles (EVs) in the context of plant immune systems. Although EV release by plant cells is known to be stimulated by fungal and bacterial infection, nothing is known about their function. Over the last two years, the Innes laboratory has developed a method for purifying and quantifying EVs from Arabidopsis leaves. Analysis of purified EVs revealed that they carry microRNAs and are highly enriched in proteins associated with response to biotic and abiotic stress, suggesting that EVs may participate in intercellular communication. The experiments described in this proposal will address the following questions: 1) Does the protein and RNA content of plant EVs change in response to specific biotic stresses? 2) Are known endomembrane trafficking proteins required for production of plant EVs? 3) Are EVs required for host-induced gene silencing (HIGS) of genes in a fungal pathogen? Question 1 will be addressed using proteomic and RNA-seq analysis of EVs purified from Arabidopsis leaves following treatment with various biotic stresses. Question 2 will be addressed by quantifying EV production in mutant Arabidopsis lines deficient in specific endomembrane trafficking proteins. Question 3 will be addressed using a forward genetics screen to identify Arabidopsis mutants that are unable to perform HIGS of genes in the fungus Fusarium graminearum. This screen will uncover Arabidopsis genes required for HIGS, and will establish whether EVs transport small RNAs from the host to the pathogen. This award is co-funded by the Plant Biotic Interactions program in the Division of Integrative Organismal Systems division and the Cellular Dynamics and Function program in the Molecular and Cellular Biosciences division.