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The general objective of this proposal is to use systems biology approaches that combine metabolomics and transcriptomics to study the F. graminearum-corn interaction in planta and further our understanding of mycotoxin accumulation in the disease process. In addition, we will use this information to begin to understand mechanisms of resistance in corn. <P>Specifically we aim to: <br/>1. Survey a panel of maize germplasm with different levels of susceptibility to F. graminearum for metabolite production during ear and stalk rot disease. We will use a F. graminearum wild type and mycotoxin mutant strains to obtain metabolite data from infected corn ears and stalks. This experiment will expand our knowledge of host factors that affect mycotoxin production, and provide information about host resistance mechanisms. <br/>2. Characterize gene expression and metabolite production during the F. graminearum-corn interaction as ear and stalk rot disease progresses. We will use F. graminearum wild type and mycotoxin mutant strains to obtain metabolite and transcriptome data from infected corn samples at successive time points with the goal to identify host and pathogen genes important for disease, mycotoxin production and host response. <br/>3. Build gene-metabolite association networks to identify candidate genes host and pathogen involved in modulation of mycotoxin production throughout disease. Gene association networks can reveal relationships between genes in biochemical pathways as well as relationships between regulatory genes and their targets. Use of a systems biology approach in combination with the mycotoxin mutants will enable the identification of genes and metabolites relevant to infection, mycotoxin production and host resistance. These experiments will also reveal commonalities and differences in metabolite production and transcriptional changes in stalk rot versus ear rot.
Non-Technical Summary:<br/>
From an agricultural standpoint, fungal diseases such as ear and stalk rot of corn are primary factors limiting corn production in many parts of the U.S. Thus, understanding the genetic basis for the F. graminearum-corn interaction and the mechanisms affecting mycotoxin production in the host is key to the long-term improvement of this industry. In the U. S., the corn industry occupies approximately 92.3 million acres, with an estimates market value of ~$77 billion. In recent years however, the high content of mycotoxins in food, and ethanol byproducts used for feed has raised significant concerns on the corn industry. The F. graminearum-corn pathosystem provides a unique and timely opportunity for investigating transcriptional and metabolite changes in pathogen and host during the progression of ear and stalk rot. First, the release and publication of the corn and F. graminearum genomes provides a wealth of underexploited genomic potential for understanding the pathogen-host interaction and regulation and production of mycotoxins. Second, working with a crop and pathogen of significant agricultural importance such as corn and F. graminearum allows bypassing the use of traditional lab models such as Arabidopsis, and learn directly from the crop to translate this knowledge into solutions for disease and mycotoxin management, not only in corn, but cereal crops in general. This project will characterize differential responses of maize germplasm to F. graminearum infection and determine throughout the disease cycle, not only the types and levels of mycotoxin production in both infected stalks and ears but also gene expression and metabolite profiles in both corn and the pathogen thereby providing the first data on how host processes affect mycotoxin production by the pathogen while causing disease. These data will form a systems biology approach to identify key fungal and host genes and mechanisms involved in mycotoxin accumulation, a first step in developing control measures to reduce the impact of the pathogen and its toxins on corn-derived food and feed.
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Approach:<br/>
We will perform a survey of metabolites produced during ear and stalk rot of corn caused by F. graminearum in a panel of 20-24 corn varieties. We will grow corn plants in the greenhouse and inoculate their stalks with a wild type and two mycotoxin (DON and ZEN) mutant strains of F. graminearum. For ear rot experiments, plants will be grown in the field and ears will be collected to perform detached ear inoculations in humid chambers in the lab with F. graminearum. We will perform disease ratings to determine the susceptibility of corn varieties to F. graminearum and we will collect infected tissue at four time points for metabolite analysis. We will generate a rich dataset of metabolite profiles across a panel of corn varieties with different levels of tolerance to F. graminearum. We expect to detect corn varieties with resistance to disease or mycotoxin accumulation, as well as host and pathogen metabolites induced in resistant vs. susceptible varieties. We will perform metabolite and transcriptomics analysis on a resistant to moderately resistant and a susceptible corn variety identified in the first set of experiments. We will sample at four time points during ear and stalk rot disease development caused by the wild type F. graminearum and two mycotoxin mutants. We will inoculate stalks (greenhouse) and ears (detached and field-grown) of these varieties with F. graminearum wild type, a DON, and a ZEN mutant. We will perform disease ratings and collect infected tissue at four time points for transcriptome and metabolome profiling. These data will allow us to characterize host and pathogen gene and metabolite expression as disease progresses, differential expression of genes and metabolites in stalk vs. ear rot as well as in wild type F. graminearum vs. DON or ZEN mutants. Using systems biology approaches, we will integrate metabolome and transcriptome data to construct gene-metabolite association networks. We expect to identify host and pathogen networks affecting mycotoxin production during stalk vs. ear rot, when wild type vs. mutant strains cause disease, and at specific time points during disease. This detailed characterization of the F. graminearum-corn interaction and of genes and metabolites affecting mycotoxin production will provide new insight about how mycotoxins accumulate in corn tissues, which is required information to develop control measures.