DISCOVERY, CHARACTERIZATION, AND EVOLUTION OF INSECT-DERIVED EFFECTORS USING WILD AND CULTIVATED GRAPE AS A MODEL SYSTEM.

The long-term goal of this research is to identify how insect secretions target conserved plant processes to enable adaptation to new genotypes. Based upon my preliminary data, I propose to: 1)Correlate insect effector gene expression to the altered plant phenotype across novel (including cultivated) and wild hosts using clonal and diverse genotypes.The preliminary data suggest variation in effector genes among phylloxera populations underlies both the physiological and morphological plant phenotype. To better understand which genes can be linked to a specific phenotype, I will profile insect gene expression across relatedVitisspecies that differ in observed traits induced by a single insect clone. My hypothesis is that differentially expressed effector genes will correlate with colonization success, and that co-expressed genes will reveal insect gene networks underlying variation in host phenotypes. I will profile defensive, nutritional, and morphological host traits alongside insect performance. Together these data will provide a phylogenetically informed understanding of what genes underlie complex phenotypes and provide context for validation studies. 2)Resolve effector diversity among naturally occurring populations using population whole genome resequencing.Naturally occurring populations of phylloxera represent novel genetic resources that may underlie the next invasion into cultivated grape or provide insights into how native grape and phylloxera co-exist with minimal impact on plant health. To better understand the genetic structure of natural populations, I will collect both plantand insect genotypes from locations across the southwestern US and Mexico. My hypothesis is that effector genes will show signatures of stabilizing selection for genes underlying coexistence and purifying selection relative to effectors that drive virulence. 3)Establish a tractable system to functionally validate effectors through CRISPR-Cas9 gene editing and immune target interaction profiling using select genes identified as unique in objectives 1 and 2.Preliminary data indicate select effector genes are amenable to protein-interaction assays that provide insight into functional interactions during colonization events. To better understand what plant proteins interact with phylloxera gene products, the phylloxera genes that are overexpressed and co-expressed across host species (Objective 1), and maintained and/or under selection within the southwestern population (Objective 2) will be chosen for study. These genes will be characterized for their plant targets using a combination of protein-interaction and gene-editing techniques. My hypothesis is that genes identified as conserved among hosts will target transcription factors and regulatory genes linked to plant immune and cell cycle pathways.