Dissecting the diverse development programmes in different tissues during the development of a nitrogen fixing nodule

Plants must acquire the elemental nutrient nitrogen from their surrounding environment and its availability is often a major limitation to plant growth. Legumes (peas and beans) have a unique strategy to deal with nitrogen limitation. They enter symbiotic interactions with soil bacteria that are able to fix an atmospheric form of nitrogen that plants cannot take up, and convert it into the biologically active form ammonia. This interaction involves the entry of the bacteria into the roots of the legume plant together with development of a 'nodule', a root outgrowth that houses the bacteria. Co-ordination of bacterial entry with cell division that leads to the formation of the nodule is essential to ensure a bacterially infected and nitrogen fixing nodule. We have uncovered some of the key genes that control nodulation responses in the outermost layer of the root, the epidermis, where bacteria enter the plant. We have also identified genes that regulate responses several layers beneath the epidermis, in the cortical cell layer where the nodule structure develops. Intriguingly there appears to be genetic overlap between these two response systems, suggesting that the function of a gene is defined by the cell type where it is active. In this proposal we will assess the mechanisms of nodulation signaling in the root epidermis and the root cortex and in particular will focus on the role of transcription factors that regulate gene expression. We will combine genetic approaches with state of the art genomic approaches to develop a systems level understanding of this process. In particular we will attempt to define the roles of the transcription factors in the different tissues and the mechanisms by which these transcription factors are able to coordinate diverse developmental programmes in different tissues. This will enable us to define the mechanisms that allow coordinated plant development leading to the formation of the nitrogen fixing nodule.

Nodulation requires the coordinated activation of two diverse developmental processes: infection thread formation in root epidermal cells to allow rhizobial invasion and nodule primordium development in root cortical cells. The developmental processes associated with nodulation are initiated following recognition of bacterially made Nod factor. Perception of Nod factor leads to oscillations in calcium that are restricted to epidermal root cells, implying that Nod factor signaling is limited to the root epidermis. However, Nod factor can activate the initiation of nodule organogenesis in the inner root cortex. Thus a diffusible signal must exist that links Nod factor perception in root epidermal cells to the activation of cell division in the root cortex. Cytokinin alone is sufficient to activate the nodule meristem, suggesting that this diffusible signal leads to localised changes in cytokinin signaling in the root cortex. We have identified a number of transcriptional regulators NSP1, NSP2, NIN and ERN1, that are necessary for nodulation signaling and that play diverse roles in the root cortex and the root epidermis. We have evidence to suggest that NIN is the diffusible signal. In this proposal we will assess the mechanisms of nodulation signaling in the root epidermis and the root cortex and define the specific roles of the transcription factors in these two different tissues. We will characterize the tissue specific gene expression regulated by the transcription factors, the promoters bound by the transcription factors and assess whether these transcription factors directly regulate different suites of gene profiles in the different tissues. We will combine this systems approach with a more focused analysis of the mode of action of these transcription factors. We will test the relevance of transcription factor movement for the coordinated development in different tissues that is a necessity of nodule formation.

Plants must acquire the elemental nutrients nitrogen and phosphorus from their surrounding environment and the availability of these nutrients is often a major limitation to plant growth. To overcome this limitation farmers apply these nutrients at high concentrations to their crop plants through the application of fertilisers, the major ingredients of which are phosphates and nitrates. The production of nitrate fertiliser is a highly energy demanding process and currently accounts for approximately 2% of the worlds energy usage. The primary energy source for fertiliser production is fossil fuels and as such fertilisers not only account for a significant cost in food production, but also almost 50% of carbon dioxide emissions from agriculture. They also account for the majority of global nitrous oxide emissions, a potent class of greenhouse gases. As energy prices rise the cost of food production increases, mostly due to the link between energy usage and fertiliser production. Reducing agricultural reliance on inorganic fertilisers will dramatically enhance sustainable and affordable food production. This is crucial if we are going to meet the increasing demands for food from an expanding and increasingly discerning global population. The major natural contributor to biologically available nitrogen in terrestrial systems are nitrogen fixing bacteria in symbiosis with plants. This symbiosis is restricted to a subset of plant species, including legumes such as peas and beans, but absent in many of our major crop plants such as wheat, rice and maize. There is no 'quick fix' to the global nitrogen challenge and biological nitrogen fixation likely holds the key to solving this problem. Our work will use state-of-the-art technologies to uncover how the symbiosis in plants is controlled in individual cells. We need this detailed understanding of the mechanisms that allow this symbiosis to form if we are to use this process to tackle agricultural addiction to nitrogen fertilisers.