1st annual “Monogram Early Career Excellence Award”

We seek to recognize outstanding young scientists and researchers in the field of small grain cereal and grass research in the UK. The excellence award will consider research contributions in both basic and more applied disciplines, as well as contributions of excellence to scientific outreach efforts.

Applicants Short List

We are pleased to announce that 6 applicants have been shortlisted for the first MECEA. They are listed below in alphabetical order and the winner will be announced at the Monogram Meeting in Dundee.

Ruth

Ruth_Bryant: Performance of wheat disease resistance can vary from year to year, causing concern to growers and therefore being of interest to breeders. Environmental effect on performance of wheat defence is thought to be the underlying cause, but it is poorly understood due to research complications. It is almost impossible to determine in the field because there are so many complicating factors, perhaps the most important being that it is difficult to distinguish between effect of environment on pathogen and effect of environment on resistance.

My PhD focuses on how climate change will affect plant disease which is conventionally studied by manipulating environment to see how the pathogen is affected and/or using historical disease records for disease forecasting. My feeling was that we have been trying to manage pathogens for many centuries, but have never managed to entirely control them however we can control the plant through breeding. For this reasons I chose to focus on how environment affects the plants ability to fight the disease. As a starting point I investigated how wheat defence to yellow rust is affected by temperature as several temperature sensitive yellow rust resistance genes have been identified.

I have shown that a change in day temperature can affect wheat resistance to Puccinia striiformis f. sp tritici (Pst), the causative agent of yellow rust. In wheat line UC1041 there was no significant difference in leaf pustule cover between plants kept in controlled environment facilities with day temperatures of 18°C and plants at 25°C. However when plants were shifted, following infection with Pst to the cooler day temperature, resistance was notably compromised. In contrast, plants shifted from 18°C to 25°C were effectively resistant. A comparable trend can be seen in young UC1041 plants and some UK varieties, yet leaf pustule cover in other varieties does not appear to follow this trend when plants undergo a temperature shift in either direction.

We hypothesise that temperature fluctuations in the field may compromise resistance in some wheat varieties, whereas resistance in other varieties is more stable. I wanted to use the system detailed above to see whether this was a general resistance phenomenon or whether it applied specifically to the yellow rust isolate I was using. However yellow rust appeared unique in being acceptant of both temperatures, other pathogens preferred one temperature to another, removing my control and making it difficult to draw conclusion from results. I am now in the process of developing a new system to confirm the hypothesis that some varieties have more temperature sensitive resistance than others and whether this is seen across infection from all pathogens types.

This research has potential to identify vulnerable periods for disease susceptibility in crops and to inform breeding to develop wheat varieties with more environmentally stable resistance. The work has been challenging in that there was very little literature out there to go on. Most effect of temperature on resistance has been on specific resistance genes, whereas I believe the effect observed is on innate immunity. I hope that by the end of my PhD I have shed some light on a complex, poorly investigated topic and opened up a new research area to help improve future crop yields and agricultural management.

Chris

Christopher Burt Eyespot is a stem base disease of wheat caused by two species of fungi, Oculimacula yallundae and Oculimacula acuformis, both of which are widespread in the UK, northern Europe and the Pacific north-west USA. However, there are only two main sources of resistance available to plant breeders; a potent resistance gene termed Pch1 previously introduced into wheat on a chromosome segment from the wild relative Aegilops ventricosa, and a moderate resistance exhibited by the variety Cappelle Desprez. Work conducted as part of my PhD provided new insights into the efficacy and genetic location of these resistances.

Pch1 has not been used widely in commercial varieties of wheat due to a linkage between Pch1 and yield limiting genes on the introgressed Ae. ventricosa chromosome segment. Breaking this linkage has proven difficult due to a low recombination frequency and also due to a lack of molecular markers that function across both wheat and Ae. ventricosa. Utilising the fully sequenced genome of Brachypodium distachyon, alongside wheat genomic resources, I targeted co-dominant Conserved Orthologous Sequence (COS) markers to the Pch1 region. Using these markers I identified recombinants within the Ae. ventricosa segment and to accurately locate Pch1 on chromosome 7DL. Furthermore, by exploiting co-linearity, I identified candidate gene regions in Brachypodium, rice and Sorghum as a prelude to the map-based cloning of the gene. More recently, I developed set of KASPar-SNP markers to the region surrounding Pch1 to enable highthroughput screening of large populations and to facilitate marker-assisted selection of the resistance by plant breeders.

The resistance in Cappelle Desprez has previously been attributed to the partial resistance gene Pch2 on chromosome 7A. Pch2 resistance to O. acuformis was mapped as a quantitative trait locus (QTL) on the distal end of chromosome 7AL and closely linked SSR markers were identified. However, it was not possible to map any resistance to O. yallundae on chromosome 7A, indicating that Pch2 has a reduced effect against this species. I confirmed this by testing a range of Pch2- containing wheat varieties, all of which demonstrated significantly lower levels of resistance to O. yallundae than to O. acuformis. Importantly, this suggests that Pch2 should not be relied upon as a stand-alone resistance in commercial varieties.

In addition to Pch2, an adult plant resistance has been identified on chromosome 5A of Cappelle Desprez. Importantly, I showed that resistance provides protection against both pathogen species, at both seedling and adult plant stages, indicating that it may provide useful disease control in wheat varieties. Two different chromosome 5A recombinant populations were tested in field trials and seedling bioassays, and the resistance was located as a major QTL on chromosome 5AL. SSR markers were identified to be associated with this QTL that will enable plant breeders to use marker-assisted selection to trace this resistance in their breeding programmes. Recently, I identified novel sources of resistance to O. yallundae and O. acuformis by screening 1,036 hexaploid genotypes from the AE Watkin’s Collection, maintained at the John Innes Centre. Repeat experiments identified accessions that consistently demonstrate high levels of resistance to both pathogens. Bi-parental populations are being developedfor three of these resistant genotypes, so that novel resistances can be genetically mapped and potentially introduced into commercial varieties by plant breeders.

In summary, my work has led to a greater understanding of the genetics controlling eyespot resistance in wheat, has provided plant breeders with markers to utilise these resistances and has

Phil

Phil Hands:Brachypodium distachyon is a proposed important model for temperate grasses, in particular the core-pooids, a significant group that includes major crop species such as wheat, barley, and oats. The significance of these crops as a food source is largely due to properties of their grain and endosperm storage capabilities but there is much about grain development and its regulation amongst the temperate grasses is poorly understood. We have produced a detailed physical and molecular level characterization of grain development B. distachyon(1). This is the first wild species to be examined in such a way and distinctive features of grain morphology and physiology are described in contrast to the perceived dogma of temperate grain morphology based largely upon analyses of domesticated wheat and barley. Direct comparison to wheat identifies key points of difference in Brachypodium grain morphology and physiology, most prominently;

  • lack of endosperm regional differentiation and distinct modified aleurone domain,
  • poorly defined peripheral aleurone layer of 1-4 cells depth
  • persistent nucellar epidermis
  • prominent endosperm cell walls and low endosperm starch levels

The modified aleurone domain has great importance in wheat grain filling and its absence, along with a persistent nucellar epidermis layer suggest a filling mechanism in Brachypodium more similar to that of rice. Low endosperm starch levels coupled with prominent cell walls indicate cell wall carbohydrates as the primary storage reserve in B. distachyon and may also reflect specific ecological adaptation against insect predation and possibly dispersal. We extended our comparative approach to a wider sample of both wild and cultivated corepooids, and other Brachypodium species, using B. distachyon as a reference point for surveying several characters of grain morphology and endosperm organization. The monogenic Brachypoideae is sister to the core-pooid tribes Poeae, Aveneae, Triticeae, and Bromeae, and Brachypodium offers a taxonomically relevant reference point for such comparison. Macroscopic, histological, and molecular analyses reveal distinct patterns of grain tissue organization and show Brachypodium to be distinct in both endosperm structure and organisation(2). The irregular peripheral aleurone is largely in contrast with the wider Pooideae species and results indicate aleurone organization is correlated with grain quality characters such as shape and starch content. Low endosperm starch reserves are generally linked to irregular peripheral aleurone, persistent nucellar epidermis and lack of distinctly creased grain morphology. Ultimately a grain’s final properties are a consequence of developmental processes generating its tissue organization. Preliminary expression analyses of Brachypodium candidate regulatory genes potentially underpinning variation reveals largely conserved patterns of expression in comparison to temperate counterparts. Brachypodium shows significant points of similarity and difference to many features commonly associated with temperate cereal grain morphology and highlights a potentially skewed knowledge of temperate grain morphology as a result of focus on wheat and barley(3). This research reveals new aspects of temperate cereal grain morphology offering insight into cereal evolution and domestication with findings valuable to both basic and applied fields of cereal research.

References

  1. Opanowicz, M., Hands, P., Betts, D., Parker, M. L., Toole, G. A., Mills, E. N., Doonan, J. H. & Drea, S. (2011) Endosperm development in Brachypodium distachyon. J Exp Bot, 62(2), 735-48.
  2. Hands, P., Kourmpetli, S., Sharples, D., Harris, R.G. & Drea, S. (2012) Analysis of grain characters in temperate grasses reveals distinctive patterns of endosperm organisation associated with grain shape. J Exp Bot, 63(17), 6253-6266
  3. Hands, P. & Drea, S. (2012) A comparative view of grain development in Brachypodium

Marta

Marta Maluk: - PhD Student from Claire Halpin Group, Division of Plant Sciences, College of Life Sciences, University of Dundee at JHI, Invergowrie, DUNDEE, DD2 5DA, UK.

My PhD study has extended work on the genetic manipulation of lignin biosynthesis in grasses using Agrobacterium–mediated transformation of barley in order to improve barley for second generation biofuels production. At the start of my PhD, I fully characterised the transformation system which was newly established at the JHI (FUNGEN, http://germinate.scri.ac.uk/planttransformation), and used it to generate transgenic barley where expression of the lignin biosynthesis gene, cinnamyl alcohol dehydrogenase (CAD), was suppressed using an RNAi construct for HvCAD2, the most highly expressed CAD gene.

Transgenic RNAiCAD plants showed reduced enzyme activity in the T0 generation and even greater reductions in the next two generations (homozygous lines). The CADRNAi barley lines had similar or slightly reduced Klason lignin content relative to the control plants but the lignin structure and composition were greatly altered. The frequency of resistant interunit bonds was increased and the yield of lignin monomers after thioacidolysis was also changed. The transgenic plants had lower amount of H and S units compared to control plants and aldehydes were incorporated into the lignin in the RNAi transformants, mainly sinapaldehyde. In conclusion, lignin structure was altered in all transformants with some differences between the various lines. The influence of down-regulation of CAD on saccharification (sugar release for biofuel production), culm strength and pathogen resistance was also investigated and will be reported in a paper in preparation.

My study demonstrated that it is possible to downregulate a gene from the lignin pathway in cereal straw in order to change lignin structure and improve sugar release without adverse impacts on plant growth, fertility and pathogen resistance. The knowledge generated from this study, can be transferred to wheat and other biomass crops in the future.

There is still one year for my PhD to be finished but the project has already achieved most of its objectives and results have been written up for submission to a good journal. My work to characterise the transformation system and collect, optimize and improve protocols and methods for characterization of transgenic barley plants, greatly helped other researchers in our group who have subsequently used the system in their own research.

Lee

Wing Sham Lee: Septoria tritici blotch disease is one of the most economically important diseases of wheat in the UK and Western Europe1, and a major recurrent problem in wheat-growing areas world-wide. The causal agent of this disease, Zymoseptoria tritici (Zt) (previously known as Mycosphaerella graminicola), infects leaves of wheat plants in which it induces host cell death after a period of symptomless growth, thus resulting in reduced photosynthesis and crop yield. However, the defence signalling networks that are activated or involved in responses to Mg infection are not yet known. Indeed, although defence signalling in dicot plant systems has been extensively studied, the molecular components of defence signalling pathways in cereal species in general are not well characterised. This has partly been due to the relative difficulty of generating knock-out or knock-down mutants of potential signalling components via stable transformation in wheat and barley.

The aim of my project has been to establish a cost-effective and moderately high-throughput functional genomics tool called Virus-induced gene silencing (VIGS)2 at Rothamsted which would allow us to identify wheat genes involved in the interaction with Zt. I screened a large number of wheat genotypes characterised for their reaction to Zt for their suitability as a host for Barley stripe mosaic virus (BSMV), the most commonly used vector for cereal VIGS. I have now identified a panel of genotypes that display consistently mild symptoms in response to wild-type BSMV and show uniform efficient silencing of control genes such as phytoene desaturase. I also showed that pre-infection of plants with BSMV does not alter their reaction to Zt. I subsequently developed a protocol that combines BSMV-VIGS and a Septoria disease attached leaf assay, allowing us to study the effect of silencing candidate defence genes on the Zt-wheat interaction in a variety of wheat genetic backgrounds.

As a proof of concept, I used BSMV-VIGS to silence the potential wheat homologues of the rice chitin receptors, CERK1 and CEBiP, which are known to be involved in the interaction with Magnaporthe oryzae. Zt encodes at least two LysM-containing effector proteins that are thought to shield fungal chitin from recognition by plant innate immunity systems. Mutants in one of these proteins, Zt3LysM, are unable to induce disease symptoms or sporulate on wheat leaves, presumably because they are unable to evade host recognition. When the wheat chitin receptors were silenced, Ztδ 3LysM was able to colonise wheat leaves (Figure 1). This suggests that chitin recognition via CERK1 and CEBiP has an important role in anti-fungal defence in wheat, and that Zt3LysM is indeed required for fungal evasion of plant innate immunity.

Using RNA sequencing data, we have now identified a large number of candidate plant genes that show altered expression levels during different stages of Zt infection of plant tissue. I will use BSMV-VIGS to determine which of these genes and gene signalling pathways are important in the Mg-wheat interaction, in order to further our understanding of the disease and ultimately, to provide insights into how this disease could be better controlled. As I am also using BSMV-VIGS to identify genes involved in the Fusarium graminearum-wheat interaction, we will try to identify common signalling pathways that are manipulated by both pathogens.

  1. Dean et al. (2012) The Top 10 fungal pathogens in molecular plant pathology. Mol. Plant Pathol. 13, 414-430.
  2. Lee et al. (2012) Barley stripe mosaic virus-mediated tools for investigating gene function in cereal plants and their pathogens. Plant Physiol. 160, 582-590.


Monika

Monika Zwirek: I am in the final year of my PhD in Claire Halpin’s lab at the University of Dundee. My thesis will highlight results obtained from the analysis of two genes of the lignin pathway in barley: 4-coumarate CoA ligase (4CL) and cinnamoyl CoA reducatse (CCR). Transgenic barley plants were produced using RNAi constructs designed with conserved regions of 4CL or CCR cDNA sequences. Despite the fact that the lignin biosynthesis pathway is being widely studied, the functions and role of Hv4CLs and HvCCRs in monocots are still poorly described. A better understanding of these would be an important step forward to effective manipulation of lignin structure, composition and content in barley biomass for improved biofuel production. One of the main objectives of my PhD work was to produce stable homozygous 4CL and CCR transgenic barley plants. I have tested these using biochemical, genetic and chemical approaches, including Klason lignin determinations. Klason is a widely used gravimetric method for wet chemistry determination of lignin. Results obtained indicate that downregulation of 4CL and CCR genes affects the ratio of two lignin fractions, acid soluble lignin (ASL) and acid insoluble lignin (AIL). Although the ratio of ASL to AIL contents is changed, the combined content of both fractions remains lower than in controls and the transgenic plants have reduced lignin overall. As the level of ASL in wild type plants is negligible, it is often overlooked within bioenergy applications. An increase in the proportion of ASL vs. AIL could be a huge advantage for industrial applications. These results will be a part of a publication that is currently in preparation.
Lately, my work has been given new broader relevance by a GWAS (Genome Wide Association Study) for biofuel-related traits in elite two-row spring barley. We screened for the yield of sugars that could be released from straw as an indicator of the amount of biofuel that might be produced. One of the discovered QTLs highlights a cluster of lignin biosynthesis genes. Thus my current work focuses on understanding what kinds of changes in these gene(s) are causing important changes in cell wall saccharification. I am exploring the variation that exists in these genes across the spring barley association panel. At the start of this study, the barley genome assembly was not available, so I employed bioinformatics to find rice genes spanning the QTL region and used these to search for the barley counterparts. I have identified over 100 new SNPs in this region and these are being mapped onto the association panel using Illumina’s VeraCode GoldenGate Genotyping Assay. At the same time as we try to fine map the region, we are also trying to understand how these new SNPs found in candidate genes could influence the phenotype. This work will further contribute to a high impact publication.

Further information about the workshop

Application Process

1) Eligibility: PhD students and early career postdocs (up to 3 year's post PhD graduation). We encourage UK-based academic and industrial applicants.

2) Nominations: Self-nominations; applicants will be required to write one page (A4, 11 Arial font, 2 cm margin, including any figures) describing a specific piece of work and detailing what they have achieved and the importance/relevance of this work. Nominations should be accompanied by a supporting statement from the PI (or a senior colleague) of no more than 300 words. Applicants should also attach an updated 2-page CV.

3) Criteria: The Monogram Steering Group will judge the applications based on the originality, excellence, and the significance of the work to the discipline of small grain cereal and grass research.

4) Prize: The award winner will receive a £300 cash prize and will have the opportunity to present their work at the 2013 Monogram meeting taking place in Dundee from April 17th to the 19th (“Monogram Early Career Excellence Award” lecture). They will also be expected to write a short piece/blog for the Monogram website.

All documents (1-page personal statement, CV, and PI supporting statement) should be combined as a single PDF file and emailed to monogram (info@monogram.ac.uk) by February 18th, 2013.