Abstract

It is now possible to produce economically important oils by genetic engineering of major EU crops such as oilseed rape. However, two major technical hurdles have to be overcome in order to market such crops. Firstly, futile cycling (breakdown of the foreign oil by the plant) represents a major generic barrier to the successful commercialisation of designer oils in crop plants. Secondly, oilseeds cannot use these foreign oils as an efficient source of fuel for young seedlings to grow, resulting in non-viable seed. CONFAB aims to overcome hurdle 1 through genetic engineering to increase the rate of oil synthesis and decrease or block the rate of breakdown of the unusual fatty acid components of the oil. CONFAB aims to overcome hurdle 2 by identifying key genes and enzymatic steps in novel oil breakdown and use these to promote seedling growth in crops that have been engineered to accumulate high levels of these valuable products.

Objectives

CONFAB will use a problem-solving approach to remove the barriers to the production of a high yield of novel oils in EU crop plants. This work is highly relevant to the objectives of the Cell Factory Key Action. Major objectives will be to:

• Develop a strategy to increase the flux of unusual fatty acids into triacylglycerol, resulting in increased yield of novel oils. A dual approach will be adopted in which we aim to: i) increase the flux of fatty acids into oil by overexpression of key genes involved in the production of triacylglycerol from diacylglycerol and fatty acid and ii) decrease the flux through the fatty acid breakdown pathway by blocking key steps in this process.
• Identify key genes and enzymatic steps in the breakdown of novel oil and unusual fatty acids. These genes will then be used to promote germination and seedling establishment in crops such as oilseed rape that have been engineered to accumulate high levels of an unusual fatty acid.

Description of the work

A dual strategy will be adopted to increase the flux of unusual fatty acids into triacylglycerol thus increasing the yield of novel oils, specifically:

• Over-expression of key genes involved in the production of oil from diacylglcerol and fatty acid to increase the flux of fatty acids into triacylglycerols (oil).
• Blocking key steps in the breakdown process responsible for futile cycling to increase the amount of fatty acid available for incorporation into oil.

This work will involve identification and characterisation of key genes and their manipulation in transgenic plants. Arabidopsis will be used as the model oilseed plant for identification and evaluation of target genes, enabling rapid progress to be made using state-of-the-art molecular genetic tools and methodologies. Technology transfer into oilseed rape will be performed upon confirmation of the effectiveness of target genes in Arabidopsis. CONFAB partners have developed novel methodologies that allow high sensitivity detection of metabolites such as fatty acyl-CoA compounds and beta-oxidation intermediates. These will be used to monitor the relative flux of unusual fatty acids into oil synthesis versus breakdown in Arabidopsis and oilseed rape.

Transgenic plants that accumulate a foreign oil exhibit poor germination and seedling growth. This will be addressed by CONFAB through characterisation of unusual fatty acid mobilisation in transgenic seed germination. The rate of mobilisation of different fatty acids as well as potential intermediates in beta-oxidation will be monitored using novel methodologies. This approach should identify the bottlenecks in the breakdown of unusual fatty acids and provide the first information necessary to allow the cloning of essential genes.

Deliverables

• Identify and manipulate key genes that control yield of novel oils: to increase the yield of novel oils a number of target genes will have to be evaluated to achieve increased fatty acid flux into oil and a decreased rate of break-down.
• Identify bottlenecks causing poor seedling growth in seeds containing new oils and genes that can alleviate this effect.

Contacts

Coordinator

• Ian GRAHAM / Centre for Novel Agricultural Products (CNAP), Dpt Biology area 7, University of York / UNITED KINGDOM

Participant

• Jens LERCHL / BASF AG Plant Biotechnology / GERMANY
• Vincent ARONDEL / CNRS UPR 9025 Laboratoire de Lipolyse Enzymatique / FRANCE
• Luigi de BELLIS / Dipartimento di Biologia Universitŕ di Lecce / ITALY
• Sten STYMNE / Swedish University of Agricultural Sciences Department of Plant Crop Science / SWEDEN
• Alison POIRIER / University of Leeds Centre for Plant Biochemistry and Biotechnology / UNITED KINGDOM
• Yves POIRIER / Université de Lausanne Institut de Biologie et Physiologie Végétales / SWITZERLAND