Comparative Grain Development in the Temperate Grasses.

Despite being a vital food source, grain development has been poorly characterised compared to other organs. While much work has been done of the evolution and development of floral organ identity and inflorescence architecture in analyses across cereal phylogeny that extend into uncultivated and wild relatives, less is known about the evolution of grain form and function, particularly with regard to its tissue organisation and the ability of the endosperm to store such rich reserves. Grain morphology and organisation can have important practical implications - for example, the wheat grain processes a distinct crease that is missing in rice, with profound implications for processing. This proposal, therefore, aims to identify regulators of grain development focusing on temperate cereals and using a comparative cellular developmental and transcriptomics approach. There has been less interest in non-crop grasses despite the potential insights that could come of such a comparative approach. Although clearly related to our domesticated cereals, grasses (like Brachypodium) have structurally and functionally distinct grains. A greater understanding of what underlies the significant differences between the grains of related non-crop species to the nutritional storehouses of cultivated cereals - which are subject to breeding and domestication - should provide knowledge to drive efforts to ensure greater yields and food security. We will exploit a comparative approach to understanding grain development in the grasses, focusing on the pooideae subfamily and selected taxa therein. Within the grass family, the pooideae subfamily has diversified in cooler climates and contains the cereals of most value to the UK economy: wheat, barley, oats and rye. In addition there are genera of interest for translational reasons including Brachypodium (particularly the distachyon species and its accompanying genomic tools), invasive weed genera such as Elymus and Bromus and important forage grasses such as Lolium. We propose that the striking organisational differences between Brachypodium, wheat and barley grains is underpinned by distinctive gene expression patterns through grain development and it is this gene expression profile that we will characterise. This project aims to use previously generated wheat gene expression data and publicly available data in order to compare developing grain gene expression profiles between wild (Brachypodium) and cultivated temperate cereals (wheat, barley). We will use the analyses to build a shortlist of genes that could be involved in grain variation, particularly with regard to grain morphology and endosperm organisation. The focus will be on Transcription Factors becuase Transcription Factors have been shown to be key players in the genetic differences distinguishing wild and cultivated species. Of ten domestication genes cloned so far in grasses (rice, maize, wheat and barley), eight are Transcription Factors.

The research proposes to provide a consummate characterisation of grain development in the temperate grasses using a comparative approach. This will be done as follows: 1. Performing a cytological survey of grains inselected taxa to determine the arrangement of key tissue types and cell contents within the various grains: We will focus on key maternal and endosperm tissues already identified as showing significant variation in preliminary analyses to date, variation that underpins the contrast between cultivated and non-crop species and features that have implications in nutritional profiles and post-harvest processing. 2. Using our detailed knowledge of grain development in Brachypodium distachyon in a developmental timecourse microarray analysis of grain development: We will to use our knowledge of the grain's development in Brachypodium distachyon Bd21 to select 5 key stages for a developmental timecouse microarray analysis. We will use this data to identify gene clusters that coincide with the initiation of key developmental stages and the activation of specific metabolic pathways - initiation of endosperm proliferation, syncitial endosperm development, cellularisation, aleurone layer differentiation and the early stages of grain filling processes.These stages also incorporate the striking grain elongation early in Brachypodium grain development and establishment of a persistent nucellar epidermis. 3. We will extend analyses of the microarray timecourse results to a cross-species comparison with barley and wheat using publically available and published datasets. 4. Finely-tuned definition of expression patterns of genes selected from the microarray analyses will be done using high-throughput mRNA ISH (in situ hybridisation). Genes with highest and lowest expression levels at these timepoints will be selected and expression validated by qPCR. We will focus on characterising the expression profiles of transcription factors.

Overall impact - The project aims to contribute significantly to the pool of basic knowledge that can be used to enhance the production of abundant and affordable food. Through our analysis of grain development we will enhance understanding of the genetic control of yield-potential, and of traits relevant to post-harvest processing. This will deliver the knowledge, tools and trained crop scientists necessary to contribute to the next generation of plant research and breeding programmes, which will be genomics-led and predictive. We will elucidate the nature of genetic mechanisms controlling key aspects of grain development to enable the production of crop varieties optimally adapted to the farming environment of today and the future. Outputs - Specific outputs include integrated knowledge of gene identity (specifically transcription factors) underlying the key agronomic traits of yield and end-use quality for crops invaluable to the UK and global economies accompanied by the molecular markers necessary for the application of this knowledge in national and international breeding programmes. This knowledge will be communicated through detailed and practical knowledge of genetics and genomics, publications (in high impact journals), trained scientific personnel (postdoctoral researchers, who will fill senior technical and managerial roles in industry and academia across the UK and worldwide), MSc and PhD students, advanced technologies (e.g. markers for breeding), public databases, patents, training, software, and genome sequences. End users - The project is relevant a whole range of end-users that includes breeders, farmers, growers, food producers, academics (in universities and other institutes working on both basic and applied agronomic biology), genomic technology researchers, biotech industry, policy makers and consumers. Enhancing our impact -We will proactively ensure that the outputs of our research are provided to end users in a format that they can utilise effectively. Mechanisms to facilitate this include: - Networking: partaking in key networks and user groups, including MONOGRAM. We will also develop databases and bioinformatics tools for distributing genomic and other data and knowledge for scientists world-wide. These organisations allow us to interact with the larger scientific community and associated organisations such as NIAB, TGAC as well as breeders, physiologists and millers. Associations such as the latter should encourage possible collaborations on LINK or HGCA projects. - Training: Key objectives include training the next generation of plant biologists who will have a keen appreciation of how their work impacts on crop research in applied ways as well as through basic research. We will also exploit the wealth of biological information available in model plants by ensuring that the new generation of scientists can translate this into crop improvement. The provision of BBSRC funding will enable training of a Postdoctoral Researcher in a range of valuable molecular and cell biology techniques as well as key bioinformatics applications: microarray analysis, phylogenetic analysis and database design and construction. The Postdoc will get further opportunities to help train PhD, MSc. and undergraduate students and to collaborate with top scientists involved in complementary but more applied areas of grain research. I currently contribute to interactive teaching resources designed to encourage A level students to undertake scientific careers through the Nuffield Foundation by providing a summer research project for rising A-level students and is a tutor at the annual Gatsby Plants Summer School. In direct terms I lecture to undergraduates and postgraduates in the University of Leicester and supervise project students in my lab - this provides an opportunity to inform and train the researchers and policy-makers of tomorrow.