Mechanisms of siRNA mediated broad-spectrum resistance to eukaryotic pathogens

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The sustainability of agriculture is threatened by pathogens, which cause substantial damage to crop yield and food safety. Although plants have evolved a myriad of immunity mechanisms to defend themselves, successful pathogens can overcome this surveillance system and cause disease. The dynamic interactions between virulence factors of pathogens and the innate immunity of a host determine whether disease will occur. To protect crops from pathogens, it is essential to identify new defense mechanisms and understand the molecular basis of their functions in order to design innovative approaches to elevate disease resistance.Recent research discovered a small RNA (sRNA)-based defense mechanism in plants. Small RNAs are short RNA molecules (usually 20-24 nucleotide in length) that guide the inhibition of target gene expression based on sequence complementarity. Gene silencing or RNA interference (RNAi) triggered by sRNAs is a fundamental and universal regulatory mechanism in eukaryotes that impacts a wide range of biological processes. During host-pathogen interactions, sRNAs produced from one organism have been observed to affect gene expression in the opposing organism. Although this "trans-species" RNAi is an exciting concept that represents a new perspective in host-pathogen arms race, many challenges remain. A fundamental challenge is to identify which sRNAs in plant execute target genes silencing in invading pathogens. Furthermore, direct experimental evidence demonstrating gene silencing guided by plant sRNAs in invading pathogens is lacking. These major gaps of knowledge need to be filled before sRNAs can be effectively deployed in disease control. In this project, we aim to investigate the mechanism underlying pathogen gene silencing by plant sRNAs during natural infection. This project builds on our recent discoveries suggesting that a specific family of plant sRNAs confer resistance to a filamentous eukaryotic pathogen. These sRNAs are produced from a unique biogenesis pathway that leads to the generation of a diverse sRNA pool, which has the potential to silence target genes in a broad range of eukaryotic pathogens. As such, we identified a designated family of "antimicrobial" sRNAs that may confer broad-spectrum resistance.We will use a combination of genetics, molecular biology, biochemistry, synthetic biology, and plant pathology approaches and the model Arabidopsis thaliana-Phytophthora capsici pathosystem to investigate the function of this specific family of sRNAs in plant immunity. Trans-species gene silencing will be monitored by detection of plant sRNAs in the protein complex that guides gene silencing in the invading pathogen. We will test the hypothesis that this novel defense mechanism confers broad-spectrum resistance by examining plant mutants defective in sRNA production for susceptibility to additional eukaryotic pathogens. We will further explore how sRNA-spawning sequences could be edited to produce bespoke sRNAs that can increase pathogen gene silencing and elevate resistance. The outcome of this project will be to provide novel insight into fundamental principles of plant immunity and offer new opportunities for the development of sustainable disease control strategies with far-reaching implications to a broad range of pathosystems.