Castor LPCAT and PDAT1A Act in Concert to Promote Transacylation of Hydroxy-Fatty Acid onto Triacylglycerol

Abstract

Oilseeds produce abundant triacylglycerol (TAG) during seed maturation, to fuel the establishment of photoautotrophism in the subsequent generation. Commonly, TAG contains 18carbon polyunsaturated fatty acids (FA), but plants also produce oils with unique chemical properties highly desirable for industrial processes. Unfortunately, plants that produce such oils are poorly suited to agronomic exploitation, leading to a desire to reconstitute novel oil biosynthesis in crop plants. Here, we studied the production and incorporation of hydroxy-FA (HFA) onto TAG in Arabidopsis (Arabidopsis thaliana) plants expressing the castor (Ricinus communis) FAH12 hydroxylase. One factor limiting HFA accumulation in these plants is the inefficient removal of HFA from the site of synthesis on phosphatidylcholine (PC). In Arabidopsis, lysophosphatidic acid acyltransferase (LPCAT) cycles FA to and from PC for modification. We reasoned that the castor LPCAT (RcLPCAT) would preferentially remove HFA from PC resulting in greater incorporation onto TAG. However, expressing RcLPCAT in Arabidopsis expressing FAH12 alone (line CL37) or together with castor acyl:CoA:diacylglycerol acyltransferase2 reduced HFA and total oil yield. Detailed analysis indicated that RcLPCAT reduced the removal of HFA from PC, possibly by competing with the endogenous LPCAT isozymes. Significantly, co-expressing RcLPCAT with castor phospholipid:diacylglycerol acyltransferase increased novel FA and total oil content, by transferring HFA from PC to diacylglycerol. Our results demonstrate that a detailed understanding is required to engineer modified FA production in oilseeds and suggest that phospholipase A2 enzymes rather than LPCAT mediate the highly efficient removal of HFA from PC in castor seeds.

Distinct Physiological Roles of Three Phospholipid:Diacylglycerol Acyltransferase Genes in Olive Fruit with Respect to Oil Accumulation and the Response to Abiotic Stress

“…Regarding biotechnological applications, castor bean PDAT has been employed to obtain Arabidopsis transgenic plants with increased amounts (up to 25%) of ricinoleic acid in the seed oil by co-expression with castor bean fatty acid hydroxylase ( Kim et al, 2011 ; van Erp et al, 2011 , 2015 ) or lysophosphatidic acid acyltransferase ( Lunn et al, 2020 ). In addition, the α-linolenic acid content of Arabidopsis seed oil was increased up to 10% by seed-specific expression of a flax PDAT ( Pan et al, 2013 ).…”

Engineering Yarrowia lipolytica for Sustainable Production of the Pomegranate Seed Oil-Derived Punicic Acid

“…communis significantly decreased the production of ricinoleic acid in A. thaliana . Analysis of the fatty acid composition indicated that expression of RcLPCAT resulted in a reduced flux of ricinoleic acid from the PC pool to the acyl-CoA pool and an increased flux to TAGs catalyzed by Pdat.…”

“…Coexpression of RcLPCAT and RcPDAT from R. communis effectively increased the accumulation of ricinoleic acid and total lipids, while coexpression of RcLPCAT and RcDGAT from R. communis significantly decreased the production of ricinoleic acid in A. thaliana. 43 Analysis of the fatty acid composition indicated that expression of RcLPCAT resulted in a reduced flux of ricinoleic acid from the PC pool to the acyl-CoA pool and an increased flux to TAGs catalyzed by Pdat. Therefore, coexpression of Lpcat and PDAT from different sources can strengthen the exchange of acyl group between the PC and acyl-CoA pools, enhancing the flux of C18:3Δ 9,11,13 from CoA to TAGs and driving the accumulation of punicic acid and total lipids.…”

Functional analysis of alternative castor bean DGAT enzymes

“…However, it was still distant from the 90 % found in castor bean seeds. Therefore, other enzymes might be related to the accumulation of the unusual FA in lipid droplets and their removal from cell membranes ( Lunn et al , 2020 ).…”