The Phosphatidylcholine Diacylglycerol Cholinephosphotransferase Is Required for Efficient Hydroxy Fatty Acid Accumulation in Transgenic Arabidopsis    

Abstract

We previously identified an enzyme, phosphatidylcholine diacylglycerol cholinephosphotransferase (PDCT), that plays an important role in directing fatty acyl fluxes during triacylglycerol (TAG) biosynthesis. The PDCT mediates a symmetrical interconversion between phosphatidylcholine (PC) and diacylglycerol (DAG), thus enriching PC-modified fatty acids in the DAG pool prior to forming TAG. We show here that PDCT is required for the efficient metabolism of engineered hydroxy fatty acids in Arabidopsis (Arabidopsis thaliana) seeds. When a fatty acid hydroxylase (FAH12) from castor (Ricinus communis) was expressed in Arabidopsis seeds, the PDCT-deficient mutant accumulated only about half the amount of hydroxy fatty acids compared with that in the wild-type seeds. We also isolated a PDCT from castor encoded by the RcROD1 (Reduced Oleate Desaturation1) gene. Seed-specific coexpression of this enzyme significantly increased hydroxy fatty acid accumulation in wild type-FAH12 and in a previously produced transgenic Arabidopsis line coexpressing a castor diacylglycerol acyltransferase 2. Analyzing the TAG molecular species and regiochemistry, along with analysis of fatty acid composition in TAG and PC during seed development, indicate that PDCT acts in planta to enhance the fluxes of fatty acids through PC and enrich the hydroxy fatty acids in DAG, and thus in TAG. In addition, PDCT partially restores the oil content that is decreased in FAH12-expressing seeds. Our results add a new gene in the genetic toolbox for efficiently engineering unusual fatty acids in transgenic oilseeds.

Coexpressing Escherichia coli Cyclopropane Synthase with Sterculia foetida Lysophosphatidic Acid Acyltransferase Enhances Cyclopropane Fatty Acid Accumulation

“…Acyltransferase enzymes facilitate the conversion of membrane lipids to TAG, the sink for fatty acid. Coexpression of castor seed DGAT2, PDCT, or PDAT along with RcFAH12 increased ricinoleic acid accumulation in TAG and decreased ricinoleate levels in membrane lipids along with partial reversion of the inhibition of FAS (Burgal et al, 2008;van Erp et al, 2011;Hu et al, 2012). We hypothesize that elevated levels of CPA in membrane lipids could result in a wrinkled phenotype by a similar feedback inhibition of FAS to that observed upon the accumulation of ricinoleate in membrane lipids (i.e.…”

“…The de novo biosynthesis from glycerol-3-phosphate and acyl-CoA occurs via the Kennedy pathway and includes three acyltransferases: glycerol-2-phosphate acyltransferase, acyl-CoA:lysophosphatidic acid acyltransferase (LPAT), and acyl-CoA:diacylglycerol acyltransferase (DGAT; Kennedy, 1961). Alternatively, acyl-CoAs can be redirected from phosphatidylcholine (PC) via the action of a phospholipase C, choline phosphotransferase, phosphatidylcholine:diacylglycerol cholinephosphotransferase (PDCT; Hu et al, 2012;Lu et al, 2009), or phospholipid: diacylglycerol acyltransferase (PDAT; Dahlqvist et al, 2000). An acyl group can be released from PC to generate lysophosphatidylcholine (LPC) by the back reaction of acyl-CoA:LPC acyltransferase (Stymne and Stobart, 1984;Wang et al, 2012) or a phospholipase A/acyl-CoA synthase (Chen et al, 2011).…”

Carbon and Acyl Chain Flux during Stress-induced Triglyceride Accumulation by Stable Isotopic Labeling of the Polar Microalga Coccomyxa subellipsoidea C169

“…These alterations could also be the result of increased activity of phosphatidylcholine:diacylglycerol cholinephosphotransferase (PDCT), which catalyzes the removal of the choline head group from intact PC allowing the conversion of the resulting DAG molecule to TG through diacylglycerol acyltransferase (15). PDCT has been found to be mainly responsible for the interconversion of PC and DAG and shown to be required for about half of the incorporation of hydroxylated FA into the TG pool in transgenic A. thaliana (16). Overexpression of PDCT in A. thaliana seeds also results in a 20% increase in polyunsaturated fatty acid (PUFA) content (17).…”

Acyl Editing and Headgroup Exchange Are the Major Mechanisms That Direct Polyunsaturated Fatty Acid Flux into Triacylglycerols

“…However, our recent discovery indicates that the main PC-to-DAG conversion is catalyzed by a phosphatidylcholine:diacylglycerol cholinephosphotransferase (PDCT) through the phosphocholine headgroup exchange between PC and DAG ( Fig. 1C; Lu et al, 2009;Hu et al, 2012). The PDCT is encoded by the REDUCED OLEATE DESATURA-TION1 (ROD1) gene (At3g15820) in Arabidopsis, which is responsible for about 40% of the flux of PUFA from PC through DAG into TAG synthesis (Lu et al, 2009).…”