Abstract
BackgroundStorage triacylglycerols in castor bean seeds are enriched in the hydroxylated fatty acid ricinoleate. Extensive tissue-specific RNA-Seq transcriptome and lipid analysis will help identify components important for its biosynthesis.Methodology/FindingsStorage triacylglycerols (TAGs) in the endosperm of developing castor (Ricinus communis) seeds are highly enriched in ricinoleic acid (18:1-OH). We have analysed neutral lipid fractions from other castor tissues using TLC, GLC and mass spectrometry. Cotyledons, like the endosperm, contain high levels of 18:1-OH in TAG. Pollen and male developing flowers accumulate TAG but do not contain 18:1-OH and leaves do not contain TAG or 18:1-OH. Analysis of acyl-CoAs in developing endosperm shows that ricinoleoyl-CoA is not the dominant acyl-CoA, indicating that either metabolic channelling or enzyme substrate selectivity are important in the synthesis of tri-ricinolein in this tissue. RNA-Seq transcriptomic analysis, using Illumina sequencing by synthesis technology, has been performed on mRNA isolated from two stages of developing seeds, germinating seeds, leaf and pollen-producing male flowers in order to identify differences in lipid-metabolic pathways and enzyme isoforms which could be important in the biosynthesis of TAG enriched in 18:1-OH. This study gives comprehensive coverage of gene expression in a variety of different castor tissues. The potential role of differentially expressed genes is discussed against a background of proteins identified in the endoplasmic reticulum, which is the site of TAG biosynthesis, and transgenic studies aimed at increasing the ricinoleic acid content of TAG.Conclusions/SignificanceSeveral of the genes identified in this tissue-specific whole transcriptome study have been used in transgenic plant research aimed at increasing the level of ricinoleic acid in TAG. New candidate genes have been identified which might further improve the level of ricinoleic acid in transgenic crops.
Highlights
Plant seeds contain a diverse range of fatty acids which have uses in the food, health and industrial sectors
Triacylglycerol (TAG) accumulates in both of these tissues, but its fatty acid composition is different to TAG synthesised in developing castor seeds
This is consistent with high levels of 18:1-OH accumulation in TAG being driven by one, or a combination of, the following: [a] restricted access to different in vivo pools of acyl-CoAs, [b] strict metabolic channelling and [c] substrate selectivity of enzymes involved in their biosynthesis
Summary
Plant seeds contain a diverse range of fatty acids which have uses in the food, health and industrial sectors. Ricinoleic acid (12hydroxyoctadec-cis-9-enoic acid; 18:1-OH) is the major fatty acid in Ricinus communis (castor bean) seeds where it constitutes up to 90% of the fatty acids found in TAGs. One of the principle uses of 18:1-OH is as a raw material for the production of Nylon-11 (N11) which is resistant to organic solvents at high temperature. An added problem is the inconsistent price of castor oil in the commercial market which can greatly affect the economics of N-11 production To overcome these problems, research has been directed at understanding the metabolic pathways of tri-ricinolein (TAG with 18:1-OH at all three positions on the glycerol backbone) biosynthesis from oleic acid in developing castor seeds with a view to engineering its synthesis in alternative oilseed crops. Extensive tissue-specific RNA-Seq transcriptome and lipid analysis will help identify components important for its biosynthesis
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