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:
The central objective of this project is to elucidate the genetic mechanisms that control resistance to the key pathogens of wheat with particular focus on Fusarium graminearum, the causal agent of Fusarium head blight (FHB). Identification of these mechanisms will give us the tools to develop strategies to engineer improved resistance to some of the most significant biotic threats to wheat production. The contemporary model for plant disease defense has identified two major response networks, basal defense and effector-triggered immunity (ETI). Evolutionarily, basal defense is thought to have arisen earlier and is a broad-spectrum defense that is activated by the perception of molecular features conserved broadly among plant pathogens. This pathway activates a wide range of defense responses but does not culminate in the death of the infected cell. ETI is believed to have evolved later in response to pathogens developing methods of suppressing basal defense. ETI is activated by very specific receptors, which detect pathogen-encoded effector molecules that act to suppress basal defense. Although ETI activates many of the same responses as basal defense, here the response is much more acute and causes the death of the responding cell. When this project began, little was known about the molecular mechanisms underlying resistance to FHB. This project developed a unique functional genomics approach to test if chosen genes have significant roles in FHB resistance. This assay is based on a virus-induced gene silencing (VIGS) system that this project has developed for wheat. Through the VIGS analysis it has been shown that a range of genes in the basal defense pathway make essential contributions to FHB resistance. Additionally, we have shown that chemical stimulation of a portion of this pathway, ethylene-induced signaling, can significantly increase resistance to FHB. This strongly supports the hypothesis that transgenic plants engineered to have elevated expression of an ethylene responsive transcription factor (ERF), which was shown to be essential for FHB resistance by VIGS, will have increased FHB resistance. Transgenic wheat plants have been generated that overexpress this ERF and several other components of the basal defense pathway and these are beginning to be tested for improved resistance to FHB, as well as other pathogens of wheat since the basal defense pathway has a broad spectrum of action. Additionally, progress has been achieved in employing next-generation sequencing to provide a very complete and sensitive survey of gene expression changes in wheat plants that are resisting or succumbing to FHB. This work will provide new candidate genes to be tested for their roles in FHB resistance.