Project description:TRI6 is a positive regulator of the trichothecene gene cluster and the production of trichothecene mycotoxins (deoxynivalenol [DON] and acetylated forms such as 15-ADON) in the cereal pathogen F. graminearum. As a global transcriptional regulator, TRI6 expression is modulated by nitrogen-limiting conditions, sources of nitrogen and carbon, pH, and light. However, the mechanism by which these diverse environmental factors affect TRI6 expression remains under-explored. In our effort to understand how nutrients affect TRI6 regulation, comparative digital expression profiling was performed with a wildtype F. graminearum and a Δtri6 mutant strain, grown in nutrient-rich conditions. Analysis showed that TRI6 negatively regulates genes of the branched-chain amino acid (BCAA) metabolic pathway. Feeding studies with deletion mutants of MCC, encoding methylcrotonyl-CoA-carboxylase, one of the key enzymes of leucine metabolism, showed that addition of leucine specifically down regulated TRI6 expression and reduced 15-ADON accumulation. Constitutive expression of TRI6 in the Δmcc mutant strain restored 15-ADON production. A combination of cellophane breach assays and pathogenicity experiments on wheat demonstrated that disrupting the leucine metabolic pathway significantly reduced disease. These findings suggest a complex interaction between one of the primary metabolic pathways with a global regulator of mycotoxin biosynthesis and virulence in F. graminearum. Triplicate samples of digital gene expression profiling data for the Tri6 mutant and wildtype strains were compared. **Please note that the raw data files for the current study are no longer available, and therefore are not included in the submission.
Project description:TRI6 is a positive regulator of the trichothecene gene cluster and the production of trichothecene mycotoxins (deoxynivalenol [DON] and acetylated forms such as 15-ADON) in the cereal pathogen F. graminearum. As a global transcriptional regulator, TRI6 expression is modulated by nitrogen-limiting conditions, sources of nitrogen and carbon, pH, and light. However, the mechanism by which these diverse environmental factors affect TRI6 expression remains under-explored. In our effort to understand how nutrients affect TRI6 regulation, comparative digital expression profiling was performed with a wildtype F. graminearum and a Δtri6 mutant strain, grown in nutrient-rich conditions. Analysis showed that TRI6 negatively regulates genes of the branched-chain amino acid (BCAA) metabolic pathway. Feeding studies with deletion mutants of MCC, encoding methylcrotonyl-CoA-carboxylase, one of the key enzymes of leucine metabolism, showed that addition of leucine specifically down regulated TRI6 expression and reduced 15-ADON accumulation. Constitutive expression of TRI6 in the Δmcc mutant strain restored 15-ADON production. A combination of cellophane breach assays and pathogenicity experiments on wheat demonstrated that disrupting the leucine metabolic pathway significantly reduced disease. These findings suggest a complex interaction between one of the primary metabolic pathways with a global regulator of mycotoxin biosynthesis and virulence in F. graminearum.
Project description:In F. graminearum, the transcriptional regulator TRI6 is encoded within the trichothecene gene cluster and regulates genes involved in the biosynthesis of the secondary metabolite deoxynivalenol (DON). Targeted disruption of TRI6 confirmed its role as a positive regulator of trichothecene genes and previous studies designated Tri6 as a pathway-specific transcriptional regulator. The Tri6 protein with its Cys2His2 zinc-finger may also conform to the class of broad-domain transcription regulators. This class of global transcriptional regulators mediate various environmental cues and generally responds to the demands of cellular metabolism. Expression profiling of F. graminearum grown under nitrogen-limiting conditions revealed that 49 out of 198 target genes are differentially regulated by TRI6. The identification of potential new targets together with deciphering novel binding site for Tri6, casts new light into the role of this transcriptional regulator in the overall growth and development of F. graminearum. Three biological replicates of Fusarium graminearum wildtype strain GZ3639 (NRRL 38155) (reference) and a tri6∆ mutant derived from GZ3639 were grown under nitrogen-limiting conditions in liquid culture for 5 hrs at 28oC
Project description:In F. graminearum, the transcriptional regulator TRI6 is encoded within the trichothecene gene cluster and regulates genes involved in the biosynthesis of the secondary metabolite deoxynivalenol (DON). Targeted disruption of TRI6 confirmed its role as a positive regulator of trichothecene genes and previous studies designated Tri6 as a pathway-specific transcriptional regulator. The Tri6 protein with its Cys2His2 zinc-finger may also conform to the class of broad-domain transcription regulators. This class of global transcriptional regulators mediate various environmental cues and generally responds to the demands of cellular metabolism. Expression profiling of F. graminearum grown under nitrogen-limiting conditions revealed that 49 out of 198 target genes are differentially regulated by TRI6. The identification of potential new targets together with deciphering novel binding site for Tri6, casts new light into the role of this transcriptional regulator in the overall growth and development of F. graminearum.
Project description:Fusarium head blight (FHB) is a major disease of cereal crops caused by the fungus Fusarium graminearum (Fg). FHB affects the flowering heads (or spikes) and developing seeds. This study compares the gene expression profile of heads from the susceptible cultivar Roblin after inoculation with either water (W) or the disarmed Fg strain tri6Δ (T6). Tri6 is a global transcription regulator affecting the trichothecene biosynthesis pathway.
Project description:Identifying the role of TRI6 in Fusarium graminearum isolate NRRL29169 by high-throughput RNA sequencing under two environemental conditions.
Project description:affy_brachy_2011_11 - affy_brachy_2011_11 - Fusarium graminearum is the causal agent of Fusarium head blight (FHB) of small-grain cereals, including wheat. Besides direct grain losses, this disease is of major concern because of the production by the pathogen of mycotoxins which are hazardous to animals, thus making the grain unfit for food or feed. Major mycotoxins produced by the fungus are trichothecens, including deoxynivalenol (DON). In our laboratory, we use Brachypodium distachyon as a model plant for cereals because of its amenability (short life cycle, numerous genomic and genetic resources, ...). We have recently shown that F. graminearum does induce head blight symptoms on this species and that DON is produced on infected spikes. We have also evidenced that a F. graminearum strain unable to produce DON exhibits reduced virulence on B. distachyon spikes, as previously shown on wheat. The aim of this project is to analyse and compare the plant response to DON producing and non-producing strains of F. graminearum. This analysis will allow to decipher the mechanisms of detoxification set up by the plant and also more specific responses due to the impact of the mycotoxin on plant metabolism and physiology. -Three conditions on B. distachyon spikes: 1-Mock inoculation (Tween 20 0,01%) 2-Inoculation by a F. graminearum wild-type strain 3-Inoculation by a F. graminearum mutant strain, unable to produce DON Spikes were point inoculated with 3ul of either Tween 20 0.01%, wild-type strain or mutant strain (300 spores) and incubated for 96 hours. Six inoculated spikes were collected and pooled for each condition and biological replicate. Three independent biological replicates were conducted. 9 arrays - Brachypodium; normal vs disease comparison,time course
Project description:Fusarium head blight (FHB) is a major disease of cereal crops caused by the fungus Fusarium graminearum (Fg). FHB affects the flowering heads (or spikes). This study compare the gene expression profile in wheat spikelets from the very susceptible spring wheat cultivar Roblin inoculated with either water (H2O), a Fg strain (GZ3639) producing the mycotoxin deoxynivalenol (+DON), or a GZ3639-derived Fg strain which has been inactivated at the Tri5 locus (-DON).
Project description:affy_brachy_2011_11 - affy_brachy_2011_11 - Fusarium graminearum is the causal agent of Fusarium head blight (FHB) of small-grain cereals, including wheat. Besides direct grain losses, this disease is of major concern because of the production by the pathogen of mycotoxins which are hazardous to animals, thus making the grain unfit for food or feed. Major mycotoxins produced by the fungus are trichothecens, including deoxynivalenol (DON). In our laboratory, we use Brachypodium distachyon as a model plant for cereals because of its amenability (short life cycle, numerous genomic and genetic resources, ...). We have recently shown that F. graminearum does induce head blight symptoms on this species and that DON is produced on infected spikes. We have also evidenced that a F. graminearum strain unable to produce DON exhibits reduced virulence on B. distachyon spikes, as previously shown on wheat. The aim of this project is to analyse and compare the plant response to DON producing and non-producing strains of F. graminearum. This analysis will allow to decipher the mechanisms of detoxification set up by the plant and also more specific responses due to the impact of the mycotoxin on plant metabolism and physiology. -Three conditions on B. distachyon spikes: 1-Mock inoculation (Tween 20 0,01%) 2-Inoculation by a F. graminearum wild-type strain 3-Inoculation by a F. graminearum mutant strain, unable to produce DON Spikes were point inoculated with 3ul of either Tween 20 0.01%, wild-type strain or mutant strain (300 spores) and incubated for 96 hours. Six inoculated spikes were collected and pooled for each condition and biological replicate. Three independent biological replicates were conducted.
Project description:Fusarium graminearum is the major causal agent of Fusarium head blight (FHB) in North America and other regions of the world. The pathogen causes direct yield losses and produces various types of trichothecenes mycotoxins [Deoxynivalenol (DON) and its acetylated forms (3-acetyl-4-deoxynivalenol=3ADON and 15-acetyl-4-deoxvevalenol=15ADON), nivalenol (NIV). Recent studies indicated that 3ADON-type isolates were significantly increased in North America and China in recent years and appears to be more aggressive based on growth rate, disease severity on different cultivars, and DON production in vitro. Thus the overall objective of this this study was to understand the molecular mechanisms that make 3ADON- and 15ADON-type populations different during infection using a susceptible cultivar, and to compare the transcriptomes of the 3ADON- and 15ADON-type populations in vitro and in planta using the RNA-seq approach as well as to identify the expression differences of candidate genes related to aggressiveness and DON production.