A wheat NAC interacts with an orphan protein and enhances resistance to Fusarium head blight disease.

Taxonomically-restricted orphan genes play an important role in environmental adaptation, as recently demonstrated by the fact that the Pooideae-specific orphan TaFROG (Triticum aestivum Fusarium Resistance Orphan Gene) enhanced wheat resistance to the economically devastating Fusarium head blight (FHB) disease. Like most orphan genes, little is known about the cellular function of the encoded protein TaFROG, other than it interacts with the central stress regulator TaSnRK1α. Here, we functionally characterized a wheat (T. aestivum) NAC-like transcription factor TaNACL-D1 that interacts with TaFROG and investigated its' role in FHB using studies to assess motif analyses, yeast transactivation, protein-protein interaction, gene expression and the disease response of wheat lines overexpressing TaNACL-D1. TaNACL-D1 is a Poaceae-divergent NAC transcription factor that encodes a Triticeae-specific protein C-terminal region with transcriptional activity and a nuclear localisation signal. The TaNACL-D1/TaFROG interaction was detected in yeast and confirmed in planta, within the nucleus. Analysis of multi-protein interactions indicated that TaFROG could form simultaneously distinct protein complexes with TaNACL-D1 and TaSnRK1α in planta. TaNACL-D1 and TaFROG are co-expressed as an early response to both the causal fungal agent of FHB, Fusarium graminearum and its virulence factor deoxynivalenol (DON). Wheat lines overexpressing TaNACL-D1 were more resistant to FHB disease than wild type plants. Thus, we conclude that the orphan protein TaFROG interacts with TaNACL-D1, a NAC transcription factor that forms part of the disease response evolved within the Triticeae.

Mitochondrial phosphate transporter and methyltransferase genes contribute to Fusarium head blight Type II disease resistance and grain development in wheat.

Fusarium head blight (FHB) is an economically important disease of wheat that results in yield loss and grain contaminated with fungal mycotoxins that are harmful to human and animal health. Herein we characterised two wheat genes involved in the FHB response in wheat: a wheat mitochondrial phosphate transporter (TaMPT) and a methyltransferase (TaSAM). Wheat has three sub-genomes (A, B, and D) and gene expression studies demonstrated that TaMPT and TaSAM homoeologs were differentially expressed in response to FHB infection and the mycotoxigenic Fusarium virulence factor deoxynivalenol (DON) in FHB resistant wheat cv. CM82036 and susceptible cv. Remus. Virus-induced gene silencing (VIGS) of either TaMPT or TaSAM enhanced the susceptibility of cv. CM82036 to FHB disease, reducing disease spread (Type II disease resistance). VIGS of TaMPT and TaSAM significantly reduced grain number and grain weight. This indicates TaSAM and TaMPT genes also contribute to grain development in wheat and adds to the increasing body of evidence linking FHB resistance genes to grain development. Hence, Fusarium responsive genes TaSAM and TaMPT warrant further study to determine their potential to enhance both disease resistance and grain development in wheat.

Analysis of the chromosomal clustering of Fusarium-responsive wheat genes uncovers new players in the defence against head blight disease.

There is increasing evidence that some functionally related, co-expressed genes cluster within eukaryotic genomes. We present a novel pipeline that delineates such eukaryotic gene clusters. Using this tool for bread wheat, we uncovered 44 clusters of genes that are responsive to the fungal pathogen Fusarium graminearum. As expected, these Fusarium-responsive gene clusters (FRGCs) included metabolic gene clusters, many of which are associated with disease resistance, but hitherto not described for wheat. However, the majority of the FRGCs are non-metabolic, many of which contain clusters of paralogues, including those implicated in plant disease responses, such as glutathione transferases, MAP kinases, and germin-like proteins. 20 of the FRGCs encode nonhomologous, non-metabolic genes (including defence-related genes). One of these clusters includes the characterised Fusarium resistance orphan gene, TaFROG. Eight of the FRGCs map within 6 FHB resistance loci. One small QTL on chromosome 7D (4.7 Mb) encodes eight Fusarium-responsive genes, five of which are within a FRGC. This study provides a new tool to identify genomic regions enriched in genes responsive to specific traits of interest and applied herein it highlighted gene families, genetic loci and biological pathways of importance in the response of wheat to disease.

The wheat SnRK1α family and its contribution to Fusarium toxin tolerance.

Deoxynivalenol (DON) is a mycotoxin produced by phytopathogenic Fusarium fungi in cereal grain and plays a role as a disease virulence factor. TaFROG (Triticum aestivum Fusarium Resistance Orphan Gene) enhances wheat resistance to DON and it interacts with a sucrose non-fermenting-1 (SNF1)-related protein kinase 1 catalytic subunit α (SnRK1α). This protein kinase family is central integrator of stress and energy signalling, regulating plant metabolism and growth. Little is known regarding the role of SnRK1α in the biotic stress response, especially in wheat. In this study, 15 wheat (Triticum aestivum) SnRK1α genes (TaSnRK1αs) belonging to four homoeologous groups were identified in the wheat genome. TaSnRK1αs are expressed ubiquitously in all organs and developmental stages apart from two members predominantly detected in grain. While DON treatment had either no effect or downregulated the transcription of TaSnRK1αs, it increased both the kinase activity associated with SnRK1α and the level of active (phosphorylated) SnRK1α. Down-regulation of two TaSnRK1αs homoeolog groups using virus induced gene silencing (VIGS) increased the DON-induced damage of wheat spikelets. Thus, we demonstrate that TaSnRK1αs contribute positively to wheat tolerance of DON and conclude that this gene family may provide useful tools for the improvement of crop biotic stress resistance.

A wheat cytochrome P450 enhances both resistance to deoxynivalenol and grain yield.

The mycotoxin deoxynivalenol (DON) serves as a plant disease virulence factor for the fungi Fusarium graminearum and F. culmorum during the development of Fusarium head blight (FHB) disease on wheat. A wheat cytochrome P450 gene from the subfamily CYP72A, TaCYP72A, was cloned from wheat cultivar CM82036. TaCYP72A was located on chromosome 3A with homeologs present on 3B and 3D of the wheat genome. Using gene expression studies, we showed that TaCYP72A variants were activated in wheat spikelets as an early response to F. graminearum, and this activation was in response to the mycotoxic Fusarium virulence factor deoxynivalenol (DON). Virus induced gene silencing (VIGS) studies in wheat heads revealed that this gene family contributes to DON resistance. VIGS resulted in more DON-induced discoloration of spikelets, as compared to mock VIGS treatment. In addition to positively affecting DON resistance, TaCYP72A also had a positive effect on grain number. VIGS of TaCYP72A genes reduced grain number by more than 59%. Thus, we provide evidence that TaCYP72A contributes to host resistance to DON and conclude that this gene family warrants further assessment as positive contributors to both biotic stress resistance and grain development in wheat.