Genetic and biochemical analysis of essential enzymes in triacylglycerol synthesis in arabidopsis

Plant oils are used in food, fuel, and feedstocks for many consumer products, and so understanding the process by which they are made and modified will help us to make plant oils more healthy, useful, and sustainable. While some of the genes encoding the ER-localized enzymatic steps to triacylglycerol (TAG) have been well understood and documented, several are still in need of study. The glycerol-3-phosphate acyl transferase (GPAT) enzymatic activity is the first step in the pathway to TAG, and it acylates glycerol 3-phosphate to produce lysophosphatidic acid. GPAT9 (AT5G60620) is conserved across land plants and is homozygous lethal, indicating an essential function. Transcript level in knockdown mutants correlates with GPAT activity and with oil levels, and the protein interacts with other enzymes in the TAG biosynthesis pathway. These data suggest that GPAT9 encodes the main GPAT involved in membrane lipid and TAG synthesis. The phosphatidic acid phosphatase (PAP) step in TAG synthesis is responsible for the hydrolysis of inorganic phosphate from phosphatidic acid and creation of diacylglycerol (DAG). There are 13 putative PAPs in Arabidopsis which are homologous to known PAPs. Most of these are involved in other processes, including the plastidial lipid synthesis pathway and signaling pathways. The Arabidopsis gene LPPβ (At4g22550) is expressed in seed tissue, its protein product is localized to the ER, and it encodes PAP activity, indicating that it is a likely candidate for the PAP involved in oil synthesis. At the conclusion of this work, questions remain about the role of LPPβ in oil synthesis and which genes encode the major enzymes involved in the steps generating phosphatidylcholine and converting it back to DAG; but the main Kennedy Pathway enzymes generating TAG have been identified and characterized