(6di) Applications of Flux Analysis to Plant Oilseeds

Allen, D. K., Michigan State University

For most crops the component of agronomic importance is the seed. Plant seeds store oil, protein and carbohydrate in a dense form and offer great potential to produce novel biochemicals and petroleum replacements. Nevertheless, much of the biochemistry of seed metabolism is poorly understood. Elucidating fluxes through the network of carbon metabolism in developing seeds is needed to provide a framework for rational metabolic engineering of plants as ?green factories?. Soybean (Glycine max) seeds are an attractive target for analysis because they provide the world's largest supply of biological oil for food and industrial applications and are the highest producer of protein of any major crop. The complexity of plant metabolism compared to prokaryotes requires more information for successful flux analysis and a greater investment in accurately mimicking in planta growth conditions. To address these challenges we have increased the number of measurements using a variety of stable- and radio-labeling experiments with subsequent analysis by NMR, GCMS, and scintillation counting. To ensure that the conditions of labeling experiments accurately mimic in planta conditions we have optimized medium composition and light levels. Embryos cultured in defined medium containing sugars and amino acids under 30 µmol m-2 s-1 green light accumulate storage reserves very close to in planta levels (39% protein and 19% oil in culture compared to 40% and 20% in planta, respectively). In addition, plants maintain an ability to refix carbon released as carbon dioxide. Oil biosynthesis is an expensive process from a carbon standpoint, with one carbon being lost to carbon dioxide for every two that go into oil production. Brassica napus seeds that produce large amounts of biomass as oil have evolved to recapture this carbon, but an understanding of the generality and regulation of this process is lacking. Furthermore, a number of parallel pathways occur in different subcellular compartments of plant cells, resulting in some less identifiable intracellular fluxes. Through the use of flux models these issues can be explored more rigorously.