1a. Objectives (from AD-416):
It is estimated that production of small grain cereals will need to increase by 50% to meet the growing worldwide demand. Fusarium head blight (FHB) has been ranked as one of the greatest threats to the production of wheat and barley. This project is focused on identifying the wheat genes with essential functions in FHB resistance. This will be accomplished using a virus-induced gene silencing (VIGS) assay which can down-regulate, or “knockdown” the expression of chosen genes so that their function can be inferred from the phenotypic change. This functional assay for genes involved in FHB resistance has been validated and a key role for genes in the ethylene-signaling pathway has already been established. In the next 5 years of work, we will conduct an in depth survey for genes functioning in FHB resistance, and determine if these genes can be used to engineer improved levels of FHB resistance. Objective 1: Determine if over-expression of genes involved in the ethylene signaling pathway can confer improved resistance to Fusarium head blight in wheat. Objective 2: Determine whether pathogen recognition receptors have essential roles in Fusarium head blight resistance in wheat. Objective 3: Test if elevated expression of pathogen recognition receptors can be used to improve resistance to Fusarium head blight and other fungal pathogens of wheat. Objective 4: Determine expression of genes in floral tissue from FHB resistant and susceptible wheat genotypes as they respond to challenge by Fusarium graminearum. Subobjectives 4A and 4B: A. Employ VIGS to test if the candidate genes identified in the RNA-seq survey encode proteins with functions that are critical for FHB resistance in wheat. B. If new genes encoding functions essential for FHB resistance are identified initiate transgenic studies to test if they can be used to improve FHB resistance.
1b. Approach (from AD-416):
This project is designed to identify the key genes involved in resistance to Fusarium head blight (FHB), and where possible, test if these genes can be utilized to improve FHB resistance. The project will utilize RNA-seq technology to identify candidate genes with possible roles in FHB resistance and susceptibility. Candidate genes will be tested by virus-induced gene silencing to assess whether or not they have functional roles in FHB resistance. Two genetic pathways, ethylene-signaling and pathogen-associated molecular pattern-induced signaling, have already been identified as having significant roles in FHB resistance, and will therefore be an important focus for this research. Genes in these pathways and others will be utilized in transgenic experiments attempting to improve FHB resistance.
3. Progress Report:
In the first five months since the certification of this project efforts have been focused on two goals. The first is the characterization of transgenic wheat plants that overexpress an ethylene-responsive transcription factor. Our previous work has shown that silencing this gene causes FHB resistant wheat to become susceptible. It is hoped that overexpression of this gene will result in an improved FHB resistance. Before these tests are possible, plants expressing the transgene at elevated levels must be identified and adequate seed must be generated. The second goal has been performing RNA-Seq experiments of resistant and susceptible wheat plants undergoing interactions with Fusarium graminearum, the fungus that causes FHB. “RNA-Seq” is the name given to newest method of detecting and measuring RNA expression changes. It is based on the use of “next generation” massively parallel DNA sequencing methods. The RNA-Seq studies will provide a state-of the-art catalog of all the changes of wheat gene expression as resistant and susceptible plants interact with F. graminearum. This will provide a roadmap for identifying genes with key functions in FHB resistance.