The biotrophic fungal pathogen Blumeria graminis causes the powdery mildew disease of cereals and grasses. We present the first crystal structure of a B. graminis effector of pathogenicity ...(CSEP0064/BEC1054), demonstrating it has a ribonuclease (RNase)-like fold. This effector is part of a group of RNase-like proteins (termed RALPHs) which comprise the largest set of secreted effector candidates within the B. graminis genomes. Their exceptional abundance suggests they play crucial functions during pathogenesis. We show that transgenic expression of RALPH CSEP0064/BEC1054 increases susceptibility to infection in both monocotyledonous and dicotyledonous plants. CSEP0064/BEC1054 interacts in planta with the pathogenesis-related protein PR10. The effector protein associates with total RNA and weakly with DNA. Methyl jasmonate (MeJA) levels modulate susceptibility to aniline-induced host RNA fragmentation. In planta expression of CSEP0064/BEC1054 reduces the formation of this RNA fragment. We propose CSEP0064/BEC1054 is a pseudoenzyme that binds to host ribosomes, thereby inhibiting the action of plant ribosome-inactivating proteins (RIPs) that would otherwise lead to host cell death, an unviable interaction and demise of the fungus.
The use of CRISPR/Cas9 systems could prove to be a valuable tool in crop research, providing the ability to fully knockout gene function in complex genomes or to precisely adjust gene function by ...knockout of individual alleles.
We compare gene editing in hexaploid wheat (Triticum aestivum) with diploid barley (Hordeum vulgare), using a combination of single genome and tri-genome targeting. High efficiency gene editing, 11-17% for single genome targeted guides and 5% for tri-genome targeted guides, was achieved in wheat using stable Agrobacterium-mediated transformation. Gene editing in wheat was shown to be predominantly heterozygous, edits were inherited in a Mendelian fashion over multiple generations and no off-target effects were observed. Comparison of editing between the two species demonstrated that more stable, heritable edits were produced in wheat, whilst barley exhibited continued and somatic editing.
Our work shows the potential to obtain stable edited transgene-free wheat lines in 36 weeks through only two generations and that targeted mutagenesis of individual homeologues within the wheat genome is achievable with a modest amount of effort, and without off-target mutations or the need for lengthy crossing strategies.
Summary
Wheat is the most widely grown crop globally, providing 20% of all human calories and protein. Achieving step changes in genetic yield potential is crucial to ensure food security, but ...efforts are thwarted by an apparent trade‐off between grain size and number. Expansins are proteins that play important roles in plant growth by enhancing stress relaxation in the cell wall, which constrains cell expansion.
Here, we describe how targeted overexpression of an α‐expansin in early developing wheat seeds leads to a significant increase in grain size without a negative effect on grain number, resulting in a yield boost under field conditions.
The best‐performing transgenic line yielded 12.3% higher average grain weight than the control, and this translated to an increase in grain yield of 11.3% in field experiments using an agronomically appropriate plant density.
This targeted transgenic approach provides an opportunity to overcome a common bottleneck to yield improvement across many crops.
See also the Commentary on this article by Cosgrove, 230: 403‐405.
Most plants associate with beneficial arbuscular mycorrhizal (AM) fungi that facilitate soil nutrient acquisition. Prior to contact, partner recognition triggers reciprocal genetic remodelling to ...enable colonisation. The plant Dwarf14-Like (D14L) receptor conditions pre-symbiotic perception of AM fungi, and also detects the smoke constituent karrikin. D14L-dependent signalling mechanisms, underpinning AM symbiosis are unknown. Here, we present the identification of a negative regulator from rice, which operates downstream of the D14L receptor, corresponding to the homologue of the Arabidopsis thaliana Suppressor of MAX2-1 (AtSMAX1) that functions in karrikin signalling. We demonstrate that rice SMAX1 is a suppressor of AM symbiosis, negatively regulating fungal colonisation and transcription of crucial signalling components and conserved symbiosis genes. Similarly, rice SMAX1 negatively controls strigolactone biosynthesis, demonstrating an unexpected crosstalk between the strigolactone and karrikin signalling pathways. We conclude that removal of SMAX1, resulting from D14L signalling activation, de-represses essential symbiotic programmes and increases strigolactone hormone production.
Wheat is a staple crop, frequently cultivated in water-restricted environments. Improving crop water-use efficiency would be desirable if grain yield can be maintained. We investigated whether a ...decrease in wheat stomatal density via the manipulation of epidermal patterning factor (EPF) gene expression could improve water-use efficiency. Our results show that severe reductions in stomatal density in EPF-overexpressing wheat plants have a detrimental outcome on yields. However, wheat plants with a more moderate reduction in stomatal density (i.e. <50% reduction in stomatal density on leaves prior to tillering) had yields indistinguishable from controls, coupled with an increase in intrinsic water-use efficiency. Yields of these moderately reduced stomatal density plants were also comparable with those of control plants under conditions of drought and elevated CO2. Our data demonstrate that EPF-mediated control of wheat stomatal development follows that observed in other grasses, and we identify the potential of stomatal density as a tool for breeding wheat plants that are better able to withstand water-restricted environments without yield loss.
In the coming decades, larger genetic gains in yield will be necessary to meet projected demand, and this must be achieved despite the destabilizing impacts of climate change on crop production. The ...root systems of crops capture the water and nutrients needed to support crop growth, and improved root systems tailored to the challenges of specific agricultural environments could improve climate resiliency. Each component of root initiation, growth and development is controlled genetically and responds to the environment, which translates to a complex quantitative system to navigate for the breeder, but also a world of opportunity given the right tools. In this review, we argue that it is important to know more about the ‘hidden half’ of crop plants and hypothesize that crop improvement could be further enhanced using approaches that directly target selection for root system architecture. To explore these issues, we focus predominantly on bread wheat (
Triticum aestivum
L.), a staple crop that plays a major role in underpinning global food security. We review the tools available for root phenotyping under controlled and field conditions and the use of these platforms alongside modern genetics and genomics resources to dissect the genetic architecture controlling the wheat root system. To contextualize these advances for applied wheat breeding, we explore questions surrounding which root system architectures should be selected for, which agricultural environments and genetic trait configurations of breeding populations are these best suited to, and how might direct selection for these root ideotypes be implemented in practice.
Wheat grains contain gluten proteins, which harbour immunogenic epitopes that trigger Coeliac disease in 1-2% of the human population. Wheat varieties or accessions containing only safe gluten have ...not been identified and conventional breeding alone struggles to achieve such a goal, as the epitopes occur in gluten proteins encoded by five multigene families, these genes are partly located in tandem arrays, and bread wheat is allohexaploid. Gluten immunogenicity can be reduced by modification or deletion of epitopes. Mutagenesis technologies, including CRISPR/Cas9, provide a route to obtain bread wheat containing gluten proteins with fewer immunogenic epitopes.
In this study, we analysed the genetic diversity of over 600 α- and γ-gliadin gene sequences to design six sgRNA sequences on relatively conserved domains that we identified near coeliac disease epitopes. They were combined in four CRISPR/Cas9 constructs to target the α- or γ-gliadins, or both simultaneously, in the hexaploid bread wheat cultivar Fielder. We compared the results with those obtained with random mutagenesis in cultivar Paragon by γ-irradiation. For this, Acid-PAGE was used to identify T1 grains with altered gliadin protein profiles compared to the wild-type endosperm. We first optimised the interpretation of Acid-PAGE gels using Chinese Spring deletion lines. We then analysed the changes generated in 360 Paragon γ-irradiated lines and in 117 Fielder CRISPR/Cas9 lines. Similar gliadin profile alterations, with missing protein bands, could be observed in grains produced by both methods.
The results demonstrate the feasibility and efficacy of using CRISPR/Cas9 to simultaneously edit multiple genes in the large α- and γ-gliadin gene families in polyploid bread wheat. Additional methods, generating genomics and proteomics data, will be necessary to determine the exact nature of the mutations generated with both methods.
Grain size is thought to be a major component of yield in many plant species. Here we set out to understand if knowledge from other cereals such as rice could translate to increased yield gains in ...wheat and lead to increased nitrogen use efficiency. Previous findings that the overexpression of OsBG1 in rice increased yields while increasing seed size suggest translating gains from rice to other cereals may help to increase yields.
The orthologous genes of OsBG1 were identified in wheat. One homoeologous wheat gene was cloned and overexpressed in wheat to understand its role in controlling seed size. Potential alteration in the nutritional profile of the grains were also analyzed in wheat overexpressing TaBG1. It was found that increased TaBG1-A expression could indeed lead to larger seed size but was linked to a reduction in seed number per plant leading to no significant overall increase in yield. Other important components of yield such as biomass or tillering did not change significantly with increased TaBG1-A expression. The nutritional profile of the grain was altered, with a significant decrease in the Zn levels in the grain associated with increased seed size, but Fe and Mn concentrations were unchanged. Protein content of the wheat grain also fell under moderate N fertilization levels but not under deficient or adequate levels of N.
TaBG1 does control seed size in wheat but increasing the seed size per se does not increase yield and may come at the cost of lower concentrations of essential elements as well as potentially lower protein content. Nevertheless, TaBG1 could be a useful target for further breeding efforts in combination with other genes for increased biomass.
The application of CRISPR/Cas9 technologies has transformed our ability to target and edit designated regions of a genome. It's broad adaptability to any organism has led to countless advancements in ...our understanding of many biological processes. Many current tools are designed for simple plant systems such as diploid species, however, efficient deployment in crop species requires a greater efficiency of editing as these often contain polyploid genomes. Here, we examined the role of temperature to understand if CRISPR/Cas9 editing efficiency can be improved in wheat. The recent finding that plant growth under higher temperatures could increase mutation rates was tested with Cas9 expressed from two different promoters in wheat. Increasing the temperature of the tissue culture or of the seed germination and early growth phase increases the frequency of mutation in wheat when the Cas9 enzyme is driven by the
promoter but not
. In contrast, Cas9 expression driven by the
promoter did not increase the mutations detected in either transformed lines or during the transformation process itself. These results demonstrate that CRISPR/Cas9 editing efficiency can be significantly increased in a polyploid cereal species with a simple change in growth conditions to facilitate increased mutations for the creation of homozygous or null knock-outs.
There is a large demand to reduce inputs for current crop production, particularly phosphate and nitrogen inputs which are the two most frequently added supplements to agricultural production. Gene ...characterization is often limited to the native species from which it was identified, but may offer benefits to other species. To understand if the rice gene Phosphate Starvation Tolerance 1 (PSTOL)
, a gene identified from rice which improves tolerance to low P growth conditions, might improve performance and provide the same benefit in wheat,
was transformed into wheat and expressed from a constitutive promoter. The ability of
to improve nutrient acquisition under low phosphate or low nitrogen was evaluated. Here we show that
works through a conserved pathway in wheat and rice to improve yields under both low phosphate and low nitrogen. This increase is yield is mainly driven by improved uptake from the soil driving increased biomass and ultimately increased seed number, but does not change the concentration of N in the straw or grain. Overexpression of
in wheat modifies N regulated genes to aid in this uptake whereas the putative homolog
does not suggesting that expression of
in wheat can help to improve yields under low input agriculture.
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