Oil-producing metabolons containing DGAT1 use separate substrate pools from those containing DGAT2 or PDAT

Authors: REGMI, ANUSHOBHA - University Of Southern Mississippi; Shockey, Jay; KOTAPATI, HARI - Washington State University; BATES, PHILIP - Washington State University

Submitted to: Plant Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/23/2020
Publication Date: 7/30/2020
Citation: Regmi, A., Shockey, J., Kotapati, H.K., Bates, P.D. 2020. Oil-producing metabolons containing DGAT1 use separate substrate pools from those containing DGAT2 or PDAT. Plant Physiology. 184(2):720-737. https://doi.org/10.1104/pp.20.00461.
DOI: https://doi.org/10.1104/pp.20.00461

Interpretive Summary: Many attempts have been made to biologically engineer oilseeds to alter their fatty acid composition, as a means of trying to produce new vegetable oils that could act as new foods and feeds, as well as feedstocks for the chemical industry. Most such attempts have not been fully successful, most likely due to incompatible integration of the foreign enzymes into the proper chemical contexts available in the cells of the developing seeds. Here we examine how each of four different oil metabolizing enzymes fit into the known pathways, seeing how well carbon flux is controlled and redistributed by each. The findings confirm that not all oil enzymes behave the same way, and that a good deal of empirical testing still remains to be done to reach a level of predictable engineering strategies. The outcome is a developing model that will help to further refine our knowledge and direct our next steps.

Technical Abstract: Seed triacylglycerol (TAG) biosynthesis involves a metabolic network containing multiple different diacylglycerol (DAG) and acyl donor substrate pools. This network of pathways overlaps with those of essential membrane lipid synthesis, and utilizes multiple TAG biosynthetic enzymes. Acyl flux through this network ultimately dictates the final oil fatty acid composition. Most strategies to alter seed oil composition involve the overexpression of lipid biosynthetic enzymes, but how these enzymes are assembled into metabolons and which substrate pools are used by each is still not well understood. To understand the roles of different classes of TAG biosynthetic acyltransferases in seed oil biosynthesis, we utilized the Arabidopsis thaliana dgat1-1 mutant (in which phosphotidylcholine:diacylglycerol acyltransferase (PDAT) is the major TAG biosynthetic enzyme), and enhanced TAG biosynthesis by expression of two diacylglycerol acyltransferases from Arabidopsis (e.g. AtDGAT1 and AtDGAT2), as well as the DGAT2 enzymes from soybean (Glycine max), and castor (Ricinus communis), followed by isotopic tracing of glycerol flux through the lipid metabolic network in developing seeds. The results indicate each acyltransferase has a unique effect on seed oil composition. AtDGAT1 produces TAG from a rapidly produced phosphatidylcholine (PC)-derived DAG pool. However, AtPDAT1 and plant DGAT2 enzymes utilize a different and larger bulk PC-derived DAG pool that is more slowly turned over for TAG biosynthesis. Based on the metabolic flux results and protein:protein interaction analyses, we present a model of TAG synthesis suggesting that substrate channeling and spatial separation of metabolic reactions in the ER membrane affect efficient TAG production and oil fatty acid composition.