UC Riverside

Since microorganisms first evolved to attack plants, there has been a constant evolutionary arms race. Plants evolve defensive mechanisms to protect themselves from pathogens that evolve countermeasures. One instance is host-induced gene silencing (HIGS), where small RNAs (sRNAs) endogenous to the plant confer resistance presumably by silencing their target gene(s) in the invading pathogen. The pathogen counter may be effectors such as Phytophthora suppressor of RNA silencing 2 (PSR2), which suppresses the sRNA processes in the plant host. This thesis aims to understand the mechanisms underlying HIGS and suppressing of HIGS by PSR2.In chapter 1, I investigated the virulence activities of PSR2, which was previously found to be associated with the Double-stranded RNA binding protein 4 (DRB4) in Arabidopsis thaliana. Unpublished data in the lab also indicate that PSR2 associates with the plant Serine/Threonine protein phosphatase 2A (PP2A). I tested the hypothesis that PSR2 may regulate the phosphorylation levels of DRB4. By examining the phosphorylation of DRB4 in Arabidopsis expressing PSR2, it appears that PSR2 may not affect the overall level of DRB4 phosphorylation. This suggests another mechanism underlies potential manipulation of DRB4 by PSR2. In chapter 2, I tested whether small interfering RNAs (siRNAs) spawned from the pentatricopeptide repeat protein (PPR) transcripts can directly silence target genes in Phytophthora pathogen(s). I attempted to generate Phytophthora capsici mutants that contain various mutations in Phyca_554980, which was previously identified to be potentially targeted by PPR-siRNAs. I was unable to manipulate Phyca_554980 through point mutation, replacement or overexpression. This could indicate a key role of this gene in the general fitness of P. capsici. In chapter 3, I explored to enhance HIGS in soybean by introducing miRNA-PPR circuits and tested whether this approach could lead to the accumulation of PPR-siRNAs that confer resistance to Phytophthora sojae. The constructs were introduced into soybean using the Agrobacterium rhizogenes-mediated hairy root gene expression system. However, I was unable to detect PPR-siRNAs despite the production of mature miRNAs in the roots. Further investigations are required to further optimize the system. My thesis research provides insight into plant-pathogen arms race centered on sRNA-mediated defense and counter-defense.