14 ERA-CAPS_Simultaneous manipulation of source and sink metabolism for improved crop yield

The capacity of the metabolic networks of different plant tissues is a key determinant of the yield of crop plants. Of particular importance is the capacity to assimilate environmental carbon (CO2) and nitrogen (NO3), the capacity to transport the resultant sugars and amino acids to the sink tissues (such as tubers, fruits and seeds) and the capacity of the sink tissues to convert the incoming sugars and amino acids into storage compounds. There is a great deal of interest in increasing the capacity or efficiency of these metabolic and transport processes by genetic engineering. Many of the current research consortia working in this area are focussing on the initial processes responsible for carbon and nitrogen assimilation in the source tissues. However, it is clear both on theoretical grounds and from experimental evidence that whole plant fluxes of carbon and nitrogen are co-limited by the metabolic capacities of both source and sink tissues. This is especially true if the source capacity is increased: control will inevitably shift to the sink tissues, the metabolism of which will therefore severely limit the yield potential of an engineered crop.

In this project, we will implement a metabolic engineering strategy of unprecedented scale in plants. Not only will we engineer both source and sink tissues, but we will target multiple metabolic and transport processes in each in an attempt to remove flux bottlenecks from across the metabolic network. The project will exploit the new technique of biolistic combinatorial co-transformation which allows the stable integration of an unlimited number of transgenes into a single locus in any plant amenable to biolistic transformation of the nuclear genome. Based on prior knowledge, we have identified 18 transgene targets which will be introduced into tomato plants. We will generate a large library of up to 200 transgenic lines and these will be screened for fruit yield, with the expectation of achieving a step-change in yield in comparison to the introduction of small numbers of transgenes modifying just source or sink. In addition, the project will undertake extensive research to identify additional metabolic bottlenecks (by comparison of metabolic network fluxes and enzyme activities), to identify transporters involved in fruit nitrogen allocationand to identify strong genetic alleles for harvest index and fruit nitrogen content (based on analysis of tomato introgression populations). This research will provide additional targets which will be super-transformed into the best performing transgenic line to assess the scope for even further yield increases.

The capacity of the metabolic networks of different plant tissues is a key determinant of the yield of crop plants. Of particular importance is the capacity to assimilate environmental carbon (CO2) and nitrogen (NO3), the capacity to transport the resultant sugars and amino acids to the sink tissues (such as tubers, fruits and seeds) and the capacity of the sink tissues to convert the incoming sugars and amino acids into storage compounds. In this project, we will implement a metabolic engineering strategy of unprecedented scale in plants. Not only will we engineer both source and sink tissues, but we will target multiple metabolic and transport processes in each in an attempt to remove flux bottlenecks from across the metabolic network. The project will exploit the new technique of biolistic combinatorial co-transformation which allows the stable integration of an unlimited number of transgenes into a single locus in any plant amenable to biolistic transformation of the nuclear genome. Based on prior knowledge, we have identified 18 transgene targets which will be introduced into tomato plants. We will generate a large library of up to 200 transgenic lines and these will be screened for fruit yield, with the expectation of achieving a step-change in yield in comparison to the introduction of small numbers of transgenes modifying just source or sink. In addition, the project will undertake extensive research to identify additional metabolic bottlenecks (by comparison of metabolic network fluxes and enzyme activities), to identify transporters involved in fruit nitrogen allocation and to identify strong genetic alleles for harvest index and fruit nitrogen content (based on analysis of tomato introgression populations). This research will provide additional targets which will be super-transformed into the best performing transgenic line to assess the scope for even further yield increases.

All knowledge generated in this project will be disseminated by timely publication in high profile peer-review journals and once published, all data will be made available in complete form, both through journal websites (supplemental files) and via a dedicated project website. The website will serve two purposes: to facilitate data and information exchange between the partners and to provide a publically accessible data and information portal. The preference will be to publish in journals with an open access policy. In addition, all members of the consortium will be active in presenting the work at international conferences.
A consortium agreement (CA) will be drawn up and signed by all partner institutions prior to the start of the project using the template provided through the ERA-CAPS call, subject to modifications by the legal offices of the partner institutions. The consortium will manage intellectual property rights (IPR) following the principle that a result shall be owned by a partner generating the result. Results may be shared if generated by more than one partner and this will lead to co-ownership wherein a separate joint ownership agreement will set out allocation and terms. All partners will endeavour to exploit any IP arising from the work. All three institutions have exceptional track records of exploitation and translation of their research and have dedicated personnel and departments for this purpose. All partners will be required to check with the project coordinator prior to any public dissemination of results (includes conference presentations and posters as well as published manuscripts) and a 3 month delay may be requested to allow protection of IPR.

Improving crop yield is of crucial importance to the European agro-economy and for food security. Tomato is a valuable crop (as reflected by the involvement of Syngenta in this project) and, after China, Europe is the most important market for tomatoes, with an annual production of around 16 million tonnes for both the fresh and processed fruit markets. This project has the potential to dramatically and rapidly increase the yield of the crop and is a timely exploration of the potential of new genetic-engineering technologies. Ultimately, the immediate economic benefit of this research is limited by current GM legislation, but it is possible that current European regulatory requirements will have been relaxed by the time this project is completed. Even were this not to be the case, Syngenta routinely uses transgenic experiments to rapidly assess the effect of specific genes and alleles and then introduces these genetic changes into their elite varieties using molecular breeding techniques to provide new seed varieties for the non-GM markets. This route to exploitation would be slower because most of our transgenic changes are gain-of-function and this is more difficult to introduce with molecular breeding or mutational approaches. Most likely, additional work would be required to find negative regulators of the target genes which could be modified by mutation. Ultimately, it would be desirable to translate this research into the main broad-acre field crops such as wheat and rice. We view the current project as a means of demonstrating the potential for multi-point manipulation of source-to-sink nutrient flows for increasing yield in an experimentally tractable and commercially important crop as well as providing gene targets that may be used to apply the same approach in cereals.