Acyl Lipid Metabolism Research Articles

Identification of transcription factor BnHDG4-A08 as a novel candidate associated with the accumulation of oleic, linoleic, linolenic, and erucic acid in Brassica napus.

We screened 47 significantly associated haplotype blocks for oleic, linoleic, linolenic, and erucic acid, with 17 blocks influencing multiple traits. A novel candidate of transcription factor BnHDG4 A08 influencing oleic, linoleic, linolenic, and erucic acid was identified, by a joint strategy of haplotype-based genome-wide association study, genomic resequencing, gene cloning, and co-expression network Fatty acid (FA) composition determines the quality and economic value of rapeseed oil (Brassica napus). However, the molecular network of FAs is unclear. In the current study, multi-strategies of haplotype-based genome-wide association study (GWAS), genomic resequencing, gene cloning, and co-expression network were joint to reveal novel genetic factors influencing FA accumulation in rapeseed. We identified 47 significantly associated haplotype blocks for oleic, linoleic, linolenic, and erucic acid, with 17 blocks influencing multiple traits, using a haplotype-based GWAS with phenotype data from 203 Chinese semi-winter accessions. A total of 61 rapeseed orthologs involved in acyl-lipid metabolism, carbohydrate metabolism, or photosynthesis were identified in these 17 blocks. Among these genes, BnHDG4-A08, encoding a class IV homeodomain leucine-zipper transcription factor, exhibited two single-nucleotide polymorphisms (SNPs) in the exon and intron, with significant associations with oleic, linoleic, linolenic, and erucic acid. Gene cloning further validated two SNPs in the exon of BnHDG4-A08 in a population with 75 accessions, leading to two amino acid changes (T372A and P366L) and significant variation of oleic, linoleic, linolenic, and erucic acid. A competitive allele-specific PCR (KASP) marker based on the SNPs was successfully developed and validated. Moreover, 98 genes exhibiting direct interconnections and high weight values with BnHDG4-A08 were identified through co-expression network analysis using transcriptome data from 13 accessions. Our study identified a novel FA candidate of transcription factor BnHDG4-A08 influencing oleic, linoleic, linolenic, and erucic acid. This gene provides a potential promising gene resource for the novel mechanistic understanding of transcription factors regulating FA accumulation.

Comparative transcriptomic analysis provides insights into the genetic networks regulating oil differential production in oil crops

BackgroundPlants differ more than threefold in seed oil contents (SOCs). Soybean (Glycine max), cotton (Gossypium hirsutum), rapeseed (Brassica napus), and sesame (Sesamum indicum) are four important oil crops with markedly different SOCs and fatty acid compositions.ResultsCompared to grain crops like maize and rice, expanded acyl-lipid metabolism genes and relatively higher expression levels of genes involved in seed oil synthesis (SOS) in the oil crops contributed to the oil accumulation in seeds. Here, we conducted comparative transcriptomics on oil crops with two different SOC materials. In common, DIHYDROLIPOAMIDE DEHYDROGENASE, STEAROYL-ACYL CARRIER PROTEIN DESATURASE, PHOSPHOLIPID:DIACYLGLYCEROL ACYLTRANSFERASE, and oil-body protein genes were both differentially expressed between the high- and low-oil materials of each crop. By comparing functional components of SOS networks, we found that the strong correlations between genes in “glycolysis/gluconeogenesis” and “fatty acid synthesis” were conserved in both grain and oil crops, with PYRUVATE KINASE being the common factor affecting starch and lipid accumulation. Network alignment also found a conserved clique among oil crops affecting seed oil accumulation, which has been validated in Arabidopsis. Differently, secondary and protein metabolism affected oil synthesis to different degrees in different crops, and high SOC was due to less competition of the same precursors. The comparison of Arabidopsis mutants and wild type showed that CINNAMYL ALCOHOL DEHYDROGENASE 9, the conserved regulator we identified, was a factor resulting in different relative contents of lignins to oil in seeds. The interconnection of lipids and proteins was common but in different ways among crops, which partly led to differential oil production.ConclusionsThis study goes beyond the observations made in studies of individual species to provide new insights into which genes and networks may be fundamental to seed oil accumulation from a multispecies perspective.

Transgenic manipulation of triacylglycerol biosynthetic enzymes in B. napus alters lipid-associated gene expression and lipid metabolism

Oilseed rape (Brassica napus) is an important crop that is cultivated for the oil (mainly triacylglycerol; TAG) it produces in its seeds. TAG synthesis is controlled mainly by key enzymes in the Kennedy pathway, such as glycerol 3-phosphate acyltransferase (GPAT), lysophosphatidate acyltransferase (LPAT) and diacylglycerol acyltransferase (DGAT) but can also be produced from phosphoglycerides such as phosphatidylcholine (PC) by the activity of the enzyme phospholipid: diacylglycerol acyltransferase (PDAT). To evaluate the potential for these enzymes to alter oil yields or composition, we analysed transgenic B. napus lines which overexpressed GPAT, LPAT or PDAT using heterologous transgenes from Arabidopsis and Nasturtium and examined lipid profiles and changes in gene expression in these lines compared to WT. Distinct changes in PC and TAG abundance and spatial distribution in embryonic tissues were observed in some of the transgenic lines, together with altered expression of genes involved generally in acyl-lipid metabolism. Overall our results show that up-regulation of these key enzymes differentially affects lipid composition and distribution as well as lipid-associated gene expression, providing important information which could be used to improve crop properties by metabolic engineering.

In Vitro Growth Conditions Boost Plant Lipid Remodelling and Influence Their Composition.

Acyl-lipids are vital components for all life functions of plants. They are widely studied using often in vitro conditions to determine inter alia the impact of genetic modifications and the description of biochemical and physiological functions of enzymes responsible for acyl-lipid metabolism. What is currently lacking is knowledge of if these results also hold in real environments—in in vivo conditions. Our study focused on the comparative analysis of both in vitro and in vivo growth conditions and their impact on the acyl-lipid metabolism of Camelina sativa leaves. The results indicate that in vitro conditions significantly decreased the lipid contents and influenced their composition. In in vitro conditions, galactolipid and trienoic acid (16:3 and 18:3) contents significantly declined, indicating the impairment of the prokaryotic pathway. Discrepancies also exist in the case of acyl-CoA:lysophospholipid acyltransferases (LPLATs). Their activity increased about 2–7 times in in vitro conditions compared to in vivo. In vitro conditions also substantially changed LPLATs’ preferences towards acyl-CoA. Additionally, the acyl editing process was three times more efficient in in vitro leaves. The provided evidence suggests that the results of acyl-lipid research from in vitro conditions may not completely reflect and be directly applicable in real growth environments.

Combining quantitative trait locus and co-expression analysis allowed identification of new candidates for oil accumulation in rapeseed.

In crops there are quantitative trait loci (QTLs) in which some of the causal quantitative trait genes (QTGs) have not been functionally characterized even in the model plant Arabidopsis. We propose an approach to delineate QTGs in rapeseed by coordinating expression of genes located within QTLs and known orthologs related to traits from Arabidopsis. Using this method in developing siliques 15 d after pollination in 71 lines of rapeseed, we established an acyl-lipid metabolism co-expression network with 21 modules composed of 270 known acyl-lipid genes and 3503 new genes. The core module harbored 76 known genes involved in fatty acid and triacylglycerol biosynthesis and 671 new genes involved in sucrose transport, carbon metabolism, amino acid metabolism, seed storage protein processes, seed maturation, and phytohormone metabolism. Moreover, the core module closely associated with the modules of photosynthesis and carbon metabolism. From the co-expression network, we selected 12 hub genes to identify their putative Arabidopsis orthologs. These putative orthologs were functionally analysed using Arabidopsis knockout and overexpression lines. Four knockout mutants exhibited lower seed oil content, while the seed oil content in 10 overexpression lines was significantly increased. Therefore, combining gene co-expression network analysis and QTL mapping, this study provides new insights into the detection of QTGs and into acyl-lipid metabolism in rapeseed.

Hybrid genome assembly and functional annotation reveals insights on lipid biosynthesis of oleaginous native isolate Parachlorella kessleri, a potential industrial strain for production of biofuel precursors

The need for mass production of biofuels has been pushed up by rising energy demand in the world population. In this context, microalgae have emerged as a potential source for biodiesel production because of its high lipid content and ability to grow on minimal media sources. However, the knowledge of the metabolic processes in these systems is fairly naive and therefore hinders the genetic progress that can be proven to improve the development of biofuel precursors. In the present study, a comprehensive analysis of an indigenous oleaginous microalgae P. kessleri has been performed, using hybrid assembly of both short Illumina and long Nanopore read, leading to assembly of a highly contiguous 64.90 Mb genome with 8371 functional genes and 874 putative transcription factors. The genome reconstruction is complemented using phylogenetic analysis of acyl lipid metabolism revealed the evolutionary conservation of these proteins among various microalgal species. Henceforth, understanding and reconstruction of global metabolic pathways will provide clues towards engineering of algal strains for enhanced production of biomass, biofuels and bioproducts (B3).

A combination of genome-wide association study and transcriptome analysis in leaf epidermis identifies candidate genes involved in cuticular wax biosynthesis in Brassica napus

BackgroundBrassica napus L. is one of the most important oil crops in the world. However, climate-change-induced environmental stresses negatively impact on its yield and quality. Cuticular waxes are known to protect plants from various abiotic/biotic stresses. Dissecting the genetic and biochemical basis underlying cuticular waxes is important to breed cultivars with improved stress tolerance.ResultsHere a genome-wide association study (GWAS) of 192 B. napus cultivars and inbred lines was used to identify single-nucleotide polymorphisms (SNPs) associated with leaf waxes. A total of 202 SNPs was found to be significantly associated with 31 wax traits including total wax coverage and the amounts of wax classes and wax compounds. Next, epidermal peels from leaves of both high-wax load (HW) and low-wax load (LW) lines were isolated and used to analyze transcript profiles of all GWAS-identified genes. Consequently, 147 SNPs were revealed to have differential expressions between HW and LW lines, among which 344 SNP corresponding genes exhibited up-regulated while 448 exhibited down-regulated expressions in LW when compared to those in HW. According to the gene annotation information, some differentially expressed genes were classified into plant acyl lipid metabolism, including fatty acid-related pathways, wax and cutin biosynthesis pathway and wax secretion. Some genes involved in cell wall formation and stress responses have also been identified.ConclusionsCombination of GWAS with transcriptomic analysis revealed a number of directly or indirectly wax-related genes and their associated SNPs. These results could provide clues for further validation of SNPs for marker-assisted breeding and provide new insights into the genetic control of wax biosynthesis and improving stress tolerance of B. napus.

Genome-wide association study identifies acyl-lipid metabolism candidate genes involved in the genetic control of natural variation for seed fatty acid traits in Brassica napus L.

Brassica napus L. represents a potential plant feedstock for the sustainable production of hydrotreated renewable fuels needed to support carbon-based energy production. However, to increase the use of plant-derived oils for energy needs, breeding efforts are required to optimize the amount and profile of fatty acids (FAs) contained in the oil extracted from B. napus seed to meet demands of the various market categories. To this end, we analyzed the genetic basis of FA content and composition of seed from a diverse panel of spring-type B. napus accessions evaluated at four US locations across multiple years. The extent of phenotypic variations for total oil content, nine FA compounds, and 14 derivative traits were found, in general, to be highly heritable. A genome-wide association study (GWAS) was conducted that detected 53 SNPs significantly associated with one or more of the 24 FA seed traits, resulting in the implicated genetic role of 12 candidate genes, four of which had two homologs each, from the acyl-lipid pathway. To our knowledge, the two detected homologs of 3-Ketoacyl-CoA thiolase (KAT), have never been associated with seed oil traits in B. napus. Through the application of whole-genome prediction, the 24 FA seed traits were generally found to have moderately high predictive abilities (70% of traits with abilities > 0.70), suggesting that these traits are highly amenable to genomic selection. Overall, our results contribute to the expanding body of knowledge regarding key enzymes in the acyl-lipid pathway at the quantitative genetic level and illustrate how genomics-assisted breeding could be leveraged to genetically improve FA seed traits in B. napus.

Reorganization of Acyl Flux through the Lipid Metabolic Network in Oil-Accumulating Tobacco Leaves.

The triacylglycerols (TAGs; i.e. oils) that accumulate in plants represent the most energy-dense form of biological carbon storage, and are used for food, fuels, and chemicals. The increasing human population and decreasing amount of arable land have amplified the need to produce plant oil more efficiently. Engineering plants to accumulate oils in vegetative tissues is a novel strategy, because most plants only accumulate large amounts of lipids in the seeds. Recently, tobacco (Nicotiana tabacum) leaves were engineered to accumulate oil at 15% of dry weight due to a push (increased fatty acid synthesis)-and-pull (increased final step of TAG biosynthesis) engineering strategy. However, to accumulate both TAG and essential membrane lipids, fatty acid flux through nonengineered reactions of the endogenous metabolic network must also adapt, which is not evident from total oil analysis. To increase our understanding of endogenous leaf lipid metabolism and its ability to adapt to metabolic engineering, we utilized a series of in vitro and in vivo experiments to characterize the path of acyl flux in wild-type and transgenic oil-accumulating tobacco leaves. Acyl flux around the phosphatidylcholine acyl editing cycle was the largest acyl flux reaction in wild-type and engineered tobacco leaves. In oil-accumulating leaves, acyl flux into the eukaryotic pathway of glycerolipid assembly was enhanced at the expense of the prokaryotic pathway. However, a direct Kennedy pathway of TAG biosynthesis was not detected, as acyl flux through phosphatidylcholine preceded the incorporation into TAG. These results provide insight into the plasticity and control of acyl lipid metabolism in leaves.

Comparative Transcriptome Analysis Reveals an Efficient Mechanism of α-Linolenic Acid in Tree Peony Seeds.

Tree peony (Paeonia section Moutan DC.) species are woody oil crops with high unsaturated fatty acid content, including α-linolenic acid (ALA/18:3; >40% of the total fatty acid). Comparative transcriptome analyses were carried out to uncover the underlying mechanisms responsible for high and low ALA content in the developing seeds of P. rockii and P. lutea, respectively. Expression analysis of acyl lipid metabolism genes revealed upregulation of select genes involved in plastidial fatty acid synthesis, acyl editing, desaturation, and triacylglycerol assembly in seeds of P. rockii relative to P. lutea. Also, in association with ALA content in seeds, transcript levels for fatty acid desaturases (SAD, FAD2, and FAD3), which encode enzymes necessary for polyunsaturated fatty acid synthesis, were higher in P. rockii compared to P. lutea. Furthermore, the overexpression of PrFAD2 and PrFAD3 in Arabidopsis increased linoleic and ALA content, respectively, and modulated the final ratio 18:2/18:3 in the seed oil. In conclusion, we identified the key steps and validated the necessary desaturases that contribute to efficient ALA synthesis in a woody oil crop. Together, these results will aid to increase essential fatty acid content in seeds of tree peonies and other crops of agronomic interest.

An integrated omics analysis reveals molecular mechanisms that are associated with differences in seed oil content between Glycine max and Brassica napus

BackgroundRapeseed (Brassica napus L.) and soybean (Glycine max L.) seeds are rich in both protein and oil, which are major sources of biofuels and nutrition. Although the difference in seed oil content between soybean (~ 20%) and rapeseed (~ 40%) exists, little is known about its underlying molecular mechanism.ResultsAn integrated omics analysis was performed in soybean, rapeseed, Arabidopsis (Arabidopsis thaliana L. Heynh), and sesame (Sesamum indicum L.), based on Arabidopsis acyl-lipid metabolism- and carbon metabolism-related genes. As a result, candidate genes and their transcription factors and microRNAs, along with phylogenetic analysis and co-expression network analysis of the PEPC gene family, were found to be largely associated with the difference between the two species. First, three soybean genes (Glyma.13G148600, Glyma.13G207900 and Glyma.12G122900) co-expressed with GmPEPC1 are specifically enriched during seed storage protein accumulation stages, while the expression of BnPEPC1 is putatively inhibited by bna-miR169, and two genes BnSTKA and BnCKII are co-expressed with BnPEPC1 and are specifically associated with plant circadian rhythm, which are related to seed oil biosynthesis. Then, in de novo fatty acid synthesis there are rapeseed-specific genes encoding subunits β-CT (BnaC05g37990D) and BCCP1 (BnaA03g06000D) of heterogeneous ACCase, which could interfere with synthesis rate, and β-CT is positively regulated by four transcription factors (BnaA01g37250D, BnaA02g26190D, BnaC01g01040D and BnaC07g21470D). In triglyceride synthesis, GmLPAAT2 is putatively inhibited by three miRNAs (gma-miR171, gma-miR1516 and gma-miR5775). Finally, in rapeseed there was evidence for the expansion of gene families, CALO, OBO and STERO, related to lipid storage, and the contraction of gene families, LOX, LAH and HSI2, related to oil degradation.ConclusionsThe molecular mechanisms associated with differences in seed oil content provide the basis for future breeding efforts to improve seed oil content.

In silico identification and characterization of a diverse subset of conserved microRNAs in bioenergy crop Arundo donax L.

MicroRNAs (miRNAs) are small non-coding RNA molecules involved in the post-transcriptional regulation of gene expression in plants. Arundo donax L. is a perennial C3 grass considered one of the most promising bioenergy crops. Despite its relevance, many fundamental aspects of its biology still remain to be elucidated. In the present study we carried out the first in silico mining and tissue-specific characterization of microRNAs and their putative targets in A. donax. We identified a total of 141 miRNAs belonging to 14 families along with the corresponding primary miRNAs, precursor miRNAs and a total of 462 high-confidence predicted targets and novel target sites were validated by 5′-race. Gene Ontology functional annotation showed that miRNA targets are constituted mainly by transcription factors, but three of the newly validated targets are enzymes involved in novel functions like RNA editing, acyl lipid metabolism and post-Golgi trafficking. Folding variability of pre-miRNA loops and phylogenetic analyses indicate variable selective pressure acting on the different miRNA families. The set of miRNAs identified in this study will pave the road to further miRNA research in Arundo donax and contribute towards a better understanding of miRNA-mediated gene regulatory processes in other bioenergy crops.

Stable, Environmental Specific and Novel QTL Identification as Well as Genetic Dissection of Fatty Acid Metabolism in Brassica napus.

Fatty acid (FA) composition is the typical quantitative trait in oil seed crops, of which study is not only closely related to oil content, but is also more critical for the quality improvement of seed oil. The double haploid (DH) population named KN with a high density SNP linkage map was applied for quantitative trait loci (QTL) analysis of FA composition in this study. A total of 406 identified QTL were detected for eight FA components with an average confidence interval (CI) of 2.92 cM, the explained phenotypic variation (PV) value ranged from 1.49 to 45.05%. Totally, 204 consensus and 91 unique QTL were further obtained via meta-analysis method for the purpose of detecting multiple environment expressed and pleiotropic QTL, respectively. Of which, 74 stable expressed and 22 environmental specific QTL were also revealed, respectively. In order to make clear the genetic mechanism of FA metabolism at individual QTL level, conditional QTL analysis was also conducted and more than two thousand conditional QTL which could not be detected under the unconditional mapping were detected, which indicated the complex interrelationship of the QTL controlling FA content in rapeseed. Through comparative genomic analysis and homologous gene annotation, 61 candidates related to acyl lipid metabolism were identified underlying the CI of FA QTL. To further visualize the genetic mechanism of FA metabolism, an intuitive and meticulous network about acyl lipid metabolism was constructed and some closely related candidates were positioned. This study provided a more accurate localization for stable and pleiotropic QTL, and a deeper dissection of the molecular regulatory mechanism of FA metabolism in rapeseed.

Homoeologous exchange is a major cause of gene presence/absence variation in the amphidiploid Brassica napus.

SummaryHomoeologous exchanges (HEs) have been shown to generate novel gene combinations and phenotypes in a range of polyploid species. Gene presence/absence variation (PAV) is also a major contributor to genetic diversity. In this study, we show that there is an association between these two events, particularly in recent Brassica napus synthetic accessions, and that these represent a novel source of genetic diversity, which can be captured for the improvement of this important crop species. By assembling the pangenome of B. napus, we show that 38% of the genes display PAV behaviour, with some of these variable genes predicted to be involved in important agronomic traits including flowering time, disease resistance, acyl lipid metabolism and glucosinolate metabolism. This study is a first and provides a detailed characterization of the association between HEs and PAVs in B. napus at the pangenome level.

Reconstructing the suberin pathway in poplar by chemical and transcriptomic analysis of bark tissues.

The tree bark periderm confers the first line of protection against pathogen invasion and abiotic stresses. The phellogen (cork cambium) externally produces cork (phellem) cells that are dead at maturity; while metabolically active, these tissues synthesize cell walls, as well as cell wall modifications, namely suberin and waxes. Suberin is a heteropolymer with aliphatic and aromatic domains, composed of acylglycerols, cross-linked polyphenolics and solvent-extractable waxes. Although suberin is essentially ubiquitous in vascular plants, the biochemical functions of many enzymes and the genetic regulation of its synthesis are poorly understood. We have studied suberin and wax composition in four developmental stages of hybrid poplar (Populus tremula x Populus alba) stem periderm. The amounts of extracellular ester-linked acyl lipids per unit area increased with tissue age, a trend not observed with waxes. We used RNA-Seq deep-sequencing technology to investigate the cork transcriptome at two developmental stages. The transcript analysis yielded 455 candidates for the biosynthesis and regulation of poplar suberin, including genes with proven functions in suberin metabolism, genes highlighted as candidates in other plant species and novel candidates. Among these, a gene encoding a putative lipase/acyltransferase of the GDSL-motif family emerged as a suberin polyester synthase candidate, and specific isoforms of peroxidase and laccase genes were preferentially expressed in cork, suggesting that their corresponding proteins may be involved in cross-linking aromatics to form lignin-like polyphenolics. Many transcriptional regulators with possible roles in meristem identity, cork differentiation and acyl-lipid metabolism were also identified. Our work provides the first large-scale transcriptomic dataset on the suberin-synthesizing tissue of poplar bark, contributing to our understanding of tree bark development at the molecular level. Based on these data, we have proposed a number of hypotheses that can be used in future research leading to novel biological insights into suberin biosynthesis and its physiological function.

Genome-wide association mapping of QTL underlying seed oil and protein contents of a diverse panel of soybean accessions

To investigate the genetic basis of variation in oil and protein contents in soybean seeds, a diverse collection of 421 mainly Chinese soybean cultivars was genotyped using 1536 SNPs, mostly from candidate genes related to acyl-lipid metabolism and from regions harboring known QTL. Six significant associations were identified for each of seed oil and protein contents which individually explained 2.7–5.9% of the phenotypic variance. Six associations occurred in or near known QTL and the remaining are putative novel QTL. Ten significant associations influenced the oil content without decreasing protein content, and vice versa. One SNP was pleiotropic, with opposite effects on oil and protein contents. The genetic region covering Map-6076 and-6077 was shown to be involved in controlling oil content in soybean by integrating the results of association mapping with information on known QTL and tissue-specific expression data. This region was subject to strong selection during the genetic improvement of soybean. Our results not only confirm and refine the map positions of known QTL but also contribute to a further elucidation of the genetic architecture of protein and oil contents in soybean seeds by identifying new associations exhibiting pleiotropic effects on seed protein and oil contents.

Three-dimensional visualization of membrane phospholipid distributions in Arabidopsis thaliana seeds: A spatial perspective of molecular heterogeneity

Arabidopsis thaliana has been widely used as a model plant to study acyl lipid metabolism. Seeds of A. thaliana are quite small (approximately 500×300μm and weigh ~20μg), making lipid compositional analyses of single seeds difficult to achieve. Here we have used matrix assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) to map and visualize the three-dimensional spatial distributions of two common membrane phospholipid classes, phosphatidylcholine (PC) and phosphatidylinositol (PI), in single A. thaliana seeds. The 3D images revealed distinct differences in distribution of several molecular species of both phospholipids among different seed tissues. Using data from these 3D reconstructions, the PC and PI mol% lipid profiles were calculated for the embryonic axis, cotyledons, and peripheral endosperm, and these data agreed well with overall quantification of these lipids in bulk seed extracts analyzed by conventional electrospray ionization-mass spectrometry (ESI-MS). In addition, MALDI-MSI was used to profile PC and PI molecular species in seeds of wild type, fad2–1, fad3–2, fad6–1, and fae1–1 acyl lipid mutants. The resulting distributions revealed previously unobserved changes in spatial distribution of several lipid molecular species, and were used to suggest new insights into biochemical heterogeneity of seed lipid metabolism. These studies highlight the value of mass spectrometry imaging to provide unprecedented spatial and chemical resolution of metabolites directly in samples even as small as a single A. thaliana seeds, and allow for expanded imaging of plant metabolites to improve our understanding of plant lipid metabolism from a spatial perspective.

Comparative Transcriptomic Analysis of Two Brassica napus Near-Isogenic Lines Reveals a Network of Genes That Influences Seed Oil Accumulation.

Rapeseed (Brassica napus) is an important oil seed crop, providing more than 13% of the world’s supply of edible oils. An in-depth knowledge of the gene network involved in biosynthesis and accumulation of seed oil is critical for the improvement of B. napus. Using available genomic and transcriptomic resources, we identified 1,750 acyl-lipid metabolism (ALM) genes that are distributed over 19 chromosomes in the B. napus genome. B. rapa and B. oleracea, two diploid progenitors of B. napus, contributed almost equally to the ALM genes. Genome collinearity analysis demonstrated that the majority of the ALM genes have arisen due to genome duplication or segmental duplication events. In addition, we profiled the expression patterns of the ALM genes in four different developmental stages. Furthermore, we developed two B. napus near isogenic lines (NILs). The high oil NIL, YC13-559, accumulates significantly higher (∼10%) seed oil compared to the other, YC13-554. Comparative gene expression analysis revealed upregulation of lipid biosynthesis-related regulatory genes in YC13-559, including SHOOTMERISTEMLESS, LEAFY COTYLEDON 1 (LEC1), LEC2, FUSCA3, ABSCISIC ACID INSENSITIVE 3 (ABI3), ABI4, ABI5, and WRINKLED1, as well as structural genes, such as ACETYL-CoA CARBOXYLASE, ACYL-CoA DIACYLGLYCEROL ACYLTRANSFERASE, and LONG-CHAIN ACYL-CoA SYNTHETASES. We observed that several genes related to the phytohormones, gibberellins, jasmonate, and indole acetic acid, were differentially expressed in the NILs. Our findings provide a broad account of the numbers, distribution, and expression profiles of acyl-lipid metabolism genes, as well as gene networks that potentially control oil accumulation in B. napus seeds. The upregulation of key regulatory and structural genes related to lipid biosynthesis likely plays a major role for the increased seed oil in YC13-559.

Transcriptome analysis and identification of genes associated with ω-3 fatty acid biosynthesis in Perilla frutescens (L.) var. frutescens.

BackgroundPerilla (Perilla frutescens (L.) var frutescens) produces high levels of α-linolenic acid (ALA), a ω-3 fatty acid important to health and development. To uncover key genes involved in fatty acid (FA) and triacylglycerol (TAG) synthesis in perilla, we conducted deep sequencing of cDNAs from developing seeds and leaves for understanding the mechanism underlying ALA and seed TAG biosynthesis.ResultsPerilla cultivar Dayudeulkkae contains 66.0 and 56.2 % ALA in seeds and leaves, respectively. Using Illumina HiSeq 2000, we have generated a total of 392 megabases of raw sequences from four mRNA samples of seeds at different developmental stages and one mature leaf sample of Dayudeulkkae. De novo assembly of these sequences revealed 54,079 unique transcripts, of which 32,237 belong to previously annotated genes. Among the annotated genes, 66.5 % (21,429 out of 32,237) showed highest sequences homology with the genes from Mimulus guttatus, a species placed under the same Lamiales order as perilla. Using Arabidopsis acyl-lipid genes as queries, we searched the transcriptome and identified 540 unique perilla genes involved in all known pathways of acyl-lipid metabolism. We characterized the expression profiles of 43 genes involved in FA and TAG synthesis using quantitative PCR. Key genes were identified through sequence and gene expression analyses.ConclusionsThis work is the first report on building transcriptomes from perilla seeds. The work also provides the first comprehensive expression profiles for genes involved in seed oil biosynthesis. Bioinformatic analysis indicated that our sequence collection represented a major transcriptomic resource for perilla that added valuable genetic information in order Lamiales. Our results provide critical information not only for studies of the mechanisms involved in ALA synthesis, but also for biotechnological production of ALA in other oilseeds.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2805-0) contains supplementary material, which is available to authorized users.

Primary Fatty Alcohols Are Major Components of Suberized Root Tissues of Arabidopsis in the Form of Alkyl Hydroxycinnamates.

Suberin is a complex hydrophobic polymer that acts as a barrier controlling water and solute fluxes and restricting pathogen infections. Suberin is deposited immediately outside of the plasmalemma in the cell wall of certain tissues such as endodermis of roots, aerial and underground periderms, and seed coats. Suberin consists of a variety of fatty acid derivatives polymerized with glycerol and phenolics. In this study, we show using liquid chromatography-tandem mass spectrometry and gas chromatography-mass spectrometry techniques that most of the fatty alcohols not covalently linked to the suberin polymer are in the form of alkyl hydroxycinnamates (AHCs), with alkyl caffeates predominating. Such compounds are not restricted to the periderm of mature roots but also are present in the endodermis of younger roots, where they are not extracted by rapid dipping in chloroform. Analysis of several mutants affected in key enzymes involved in the biosynthesis and export of suberin monomers suggests that the formation of the suberin polymer and associated waxes involves common pathways and occurs concomitantly in Arabidopsis (Arabidopsis thaliana) roots. Although fatty alcohols represent only minor components of the suberin polymer in Arabidopsis roots, this study demonstrates that they constitute the major aliphatics of suberin-associated waxes in the form of AHCs. Therefore, our results indicate that esterified fatty alcohols, both soluble and polymerized forms, represent major constituents of Arabidopsis root suberized barriers, being as abundant as α,ω-dicarboxylic and unsubstituted fatty acids. In addition, our results show that suberized layers represent a major sink for acyl-lipid metabolism in Arabidopsis roots.