Identification of genes associated with ricinoleic acid accumulation in Hiptage benghalensis via transcriptome analysis

Abstract

BackgroundRicinoleic acid is a high-value hydroxy fatty acid with broad industrial applications. Hiptage benghalensis seed oil contains a high amount of ricinoleic acid (~ 80%) and represents an emerging source of this unusual fatty acid. However, the mechanism of ricinoleic acid accumulation in H. benghalensis is yet to be explored at the molecular level, which hampers the exploration of its potential in ricinoleic acid production.ResultsTo explore the molecular mechanism of ricinoleic acid biosynthesis and regulation, H. benghalensis seeds were harvested at five developing stages (13, 16, 19, 22, and 25 days after pollination) for lipid analysis. The results revealed that the rapid accumulation of ricinoleic acid occurred at the early–mid-seed development stages (16–22 days after pollination). Subsequently, the gene transcription profiles of the developing seeds were characterized via a comprehensive transcriptome analysis with second-generation sequencing and single-molecule real-time sequencing. Differential expression patterns were identified in 12,555 transcripts, including 71 enzymes in lipid metabolic pathways, 246 putative transcription factors (TFs) and 124 long noncoding RNAs (lncRNAs). Twelve genes involved in diverse lipid metabolism pathways, including fatty acid biosynthesis and modification (hydroxylation), lipid traffic, triacylglycerol assembly, acyl editing and oil-body formation, displayed high expression levels and consistent expression patterns with ricinoleic acid accumulation in the developing seeds, suggesting their primary roles in ricinoleic acid production. Subsequent co-expression network analysis identified 57 TFs and 35 lncRNAs, which are putatively involved in the regulation of ricinoleic acid biosynthesis. The transcriptome data were further validated by analyzing the expression profiles of key enzyme-encoding genes, TFs and lncRNAs with quantitative real-time PCR. Finally, a network of genes associated with ricinoleic acid accumulation in H. benghalensis was established.ConclusionsThis study was the first step toward the understating of the molecular mechanisms of ricinoleic acid biosynthesis and oil accumulation in H. benghalensis seeds and identified a pool of novel genes regulating ricinoleic acid accumulation. The results set a foundation for developing H. benghalensis into a novel ricinoleic acid feedstock at the transcriptomic level and provided valuable candidate genes for improving ricinoleic acid production in other plants.Electronic supplementary materialThe online version of this article (10.1186/s13068-019-1358-2) contains supplementary material, which is available to authorized users.

Transcriptome analysis of developing castor bean seeds and identification of ricinoleic acid biosynthesis genes

“…Furthermore, in other plants which can produce ricinoleic acid, DGAT also possesses a signi cant affect in ricinoleic acid-enriched TAGs enrichment. In P. fendleri, DGAT1, DGAT2 and PDAT2 were all conducive primarily to hydroxy fatty acid enrichment [34], whereas in H. benghalensis, DGAT2 and PDAT2 were found to own high expression levels in the process of ricinoleic acid biosynthesis [31].…”

“…In this study, a LACS7gene (29844.t000211), had high expression levels during seed development, may contribute to ricinoleic acid accumulation (Table 3). Nevertheless, in H. benghalensis, LACS8 has been reported to have the same effect [31]. At the same time, LACS9 as the dominant LACS isoform was revealed to act an vital role in oil synthesis in R. communis and P. fendleri [34].…”

“…In this study, two OLE genes (29794.t000071 and 30147.t000162) showed the highest expression level among all selected candidate key DEGs. Tian et al reported that high expression of OLE3 during seed development in H. benghalensis was participated in the accumulation of ricinoleic acid in oil-body formation [31]. OLE genes had been reported to contribute to monoacylglycerol acyltransferase and PLA2 bifunctional enzyme activity in peanuts [37].…”

“…The ricinoleic acid content was measured by gas chromatography. Heptadecanoic acid (C17:0) (≥ 99%, GLPBIO, USA) was chosen as the internal standard, following the method of Pan et al with minor modi cation [31,43]. About 15 mg of seed samples were methylated to generate fatty acid methyl esters with 3 N methanolic HCl at 80 °C for 2 h, which were extracted with n-hexane, dried in nitrogen and suspended in 1.5 mL dichloromethane.…”

“…Han et al (2020) [30] found autophagy-related genes in castor bean by RNA-sEq. Tian et al (2019) revealed a series of genes related to biosynthesis via transcriptome analysis in Hiptage benghalensis [31].…”