Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671
Performing Department
(N/A)
Non Technical Summary
OverviewTo successfully infect plants, symbiotic microbes must locate, enter, and manipulate host plants. Therefore, host range of free-living, symbiotic microbes depends on whether the microbes can locate potential plant hosts. This project will provide mechanistic understanding of how a group of soil-borne pathogens locates roots of preferred host plants. Soil-borne, motile symbionts locate host roots by recognizing and responding to root exudates via chemotaxis. To sense the complex milieu of root exudate chemicals, root-colonizing bacteria have large repertoires of chemosensors (called methyl-accepting chemotaxis proteins, or MCPs) that bind chemicals and initiate an intracellular signaling cascade that drives bacteria to swim up gradients of the chemical attractants. We hypothesize that MCP repertoires (and the respective ligand preferences) contribute to the host range of bacterial strains by allowing them to locate roots of preferred hosts. We will test this hypothesis using the agriculturally important phytopathogens in the Ralstonia solanacearum species complex (RSSC). The experimental approach leverages the natural genetic diversity of these model pathogens to understand how chemotaxis genotypes influence host range phenotypes. The project will use emerging tools (HyChemosensors, random barcoded transposon site sequencing [RB-TnSeq], and EcoFabs) to connect the molecular mechanisms to microbial ecosystems. The project has two aims that will independently yield complementary, mechanistic insights into the chemical ecology of bacteria-root interactions:Aim 1: Define the chemical specificities of core and accessory MCP chemoreceptors.Aim 2: Investigate the role of chemotactic specificity in driving host preferenceIntellectual MeritThis project will provide mechanistic insight into the chemical ecology of the rhizosphere. It is well-appreciated that pathogen host range is influenced by repertoires of effectors that pathogens use to manipulate host physiology. In contrast, there is little mechanistic understanding of how chemosensor repertoires impact host preference, and therefore, host range. Although chemosensors can be readily identified in symbiont genomes, it is not yet possible to predict which phytochemicals are sensed by each chemosensor based on their amino acid sequence alone. The molecular and phenomic data generated in this project will empower future computational biology approaches to decipher new ligand-binding motifs. Moreover, understanding the specificity of chemosensors will enable scientists to understand how host-produced chemicals drive the assembly of microbial communities.Broader ImpactsThe project will build STEM talent by broadening participation in science. UC Davis and University of Arizone are an "emerging Hispanic-Serving Institution" due to institutional initiatives that broadenaccess to STEM Bachelor's degrees for students that start their undergraduate education at local community colleges. In the CURE, cohorts of students will be immersed in scientific discovery by dissecting the specificity of chemoreceptors. Success of the broader impacts will be quantitatively evaluated by the Conversational Networking Tool and qualitatively evaluated based on students' continued engagement with research through opportunities including: presenting their results at a campus research conference, enrolling as Learning Assistants for the CURE, applying for research fellowships, and applying for graduate school.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Goals / Objectives
Host range is the result of molecular interactions between symbiotic microbes and their hosts. To successfully infect a host, pathogenic, commensal, and mutualistic symbionts must accomplish three goals: (1) locate the host, (2) enter the host, and (3) manipulate host physiology and immunity to enable the symbiont's growth1,2. Although it is well-established that motile symbionts locate plant hosts by using chemosensors to perceive phytochemicals3-6, it is unclear how the specificity of bacterial chemosensors impacts host preference7-8. To understand how chemosensor genotypes influence host range phenotypes, we propose to dissect the chemical dialogue between plant roots and agriculturally important bacterial wilt pathogens with a complex host range: the Ralstonia solanacearum species complex (RSSC).Aim 1: Define the chemical specificities of core and accessory MCP chemoreceptors.Aim 1.1: Identify the substrate specificities of core RSSC MCPs using HyChemosensor screens.Aim 1.2: Identify and functionally characterize strain-specific accessory MCPs(Integrated with the project's BROADER IMPACTS via the "Sensory SynBio CURE")Aim 1.3: Screen mutant libraries with RB-TnSeq to identify RSSC mutants with chemotaxis defects.Aim 1.4: Quantify physiological responses of RSSC Δmcp mutants.Aim 2: Investigate the role of chemotactic specificity in driving host preferenceAim 2.1: Quantify disease outcome of model RSSC strains against diverse plant hosts.Aim 2.2: Quantify RSSC chemoattraction to root exudates from phylogenetically diverse plant hosts.Aim 2.3: Profile primary metabolites in tomato, melon, and impatiens root exudates.Aim 2.4: Determine whether distinct MCPs contribute to RSSC colonization of roots from preferred hosts.Aim 2.5: Quantify mutants' responses to root exudates and visualize their root colonization phenotypes.
Project Methods
We will combine several cutting-edge methodsto address the scientific goals. We will use a forward genomic screen, random-barcoded transposon mutant sequencing (RB-TnSeq), to identify genes that contribute to (1) bacterial sensing/movement to specific chemicals from plant roots and (2) bacterial colonization of roots. We will use synthetic biology to develop "hybrid biosensor" proteins that allow us to perform chemical genomics screens to pinpoint the chemical specificity of individual chemosensors. We will optimize reproducible systems for collecting root exudate chemicals from hydroponic culture to identify the chemical milleu that mediates interactions between plants and the root microbiota, including pathogen members of the microbiota.