Lipid remodelling: unravelling the response to cold stress in Arabidopsis and its extremophile relative Eutrema salsugineum

This is an Open Access article distributed under the terms of the Creative Commons Attribution 4.0 International License https://creativecommons.org/licenses/by/4.0/ Crown Copyright © 2017 Published by Elsevier Ireland Ltd.Environmental constraints limit the geographic distribution of many economically important crops. Cold stress is an important abiotic stress that affects plant growth and development, resulting in loss of vigour and surface lesions. These symptoms are caused by, among other metabolic processes, the altered physical and chemical composition of cell membranes. As a major component of cell membranes lipids have been recognized as having a significant role in cold stress, both as a mechanical defence through leaf surface protection and plasma membrane remodelling, and as signal transduction molecules. We present an overview integrating gene expression and lipidomic data published so far in Arabidopsis and its relative the extremophile Eutrema salsugineum. This data enables a better understanding of the contribution of the lipidome in determining the ability to tolerate suboptimal temperature conditions. Collectively this information will allow us to identify the key lipids and pathways responsible for resilience, enabling the development of new approaches for crop tolerance to stress.Peer reviewe

Modifying the lipid content and composition of plant seeds: engineering the production of LC-PUFA

Omega-3 fatty acids are characterized by a double bond at the third carbon atom from the end of the carbon chain. Latterly, long chain polyunsaturated omega-3 fatty acids such as eicosapentaenoic acid (EPA; 20:5Δ5,8,11,14,17) and docosahexanoic acid (DHA; 22:6 Δ4,7,10,13,16,19), which typically only enter the human diet via the consumption of oily fish, have attracted much attention. The health benefits of the omega-3 LC-PUFAs EPA and DHA are now well established. Given the desire for a sustainable supply of omega-LC-PUFA, efforts have focused on enhancing the composition of vegetable oils to include these important fatty acids. Specifically, EPA and DHA have been the focus of much study, with the ultimate goal of producing a terrestrial plant-based source of these so-called fish oils. Over the last decade, many genes encoding the primary LC-PUFA biosynthetic activities have been identified and characterized. This has allowed the reconstitution of the LC-PUFA biosynthetic pathway in oilseed crops, producing transgenic plants engineered to accumulate omega-3 LC-PUFA to levels similar to that found in fish oil. In this review, we will describe the most recent developments in this field and the challenges of overwriting endogenous seed lipid metabolism to maximize the accumulation of these important fatty acids

Oat and lipolysis: food matrix effect

Oat is rich in a wide range of phytochemicals with various physico-chemical, colloidal and interfacial properties. These characteristics are likely to influence human lipid metabolism and the subsequent effect on health following oat consumption. The aim of this work was to investigate the impact of oat materials varying in complexity on the lipolysis process. The composition, structure and digestibility of different lipid systems (emulsions, oil bodies and oil enriched in phytosterols) were determined. The surface activities of phytosterols were examined using the pendant drop technique. Differences in lipid digestibility of the oat oil emulsions and the oil bodies were clearly seen. Also, the digestion of sunflower oil was reduced proportionally to the concentration of phytosterols present. This may be due to their interfacial properties as demonstrated by the pendant drop experiments. This work highlights the importance of considering the overall structure of the system studied and not only its composition

Using lipidomics to reveal details of lipid accumulation in developing seeds from oilseed rape (Brassica napus L.)

With dwindling available agricultural land, concurrent with increased demand for oil, there is much current interest in raising oil crop productivity. We have been addressing this issue by studying the regulation of oil accumulation in oilseed rape (Brassica napus L). As part of this research we have carried out a detailed lipidomic analysis of developing seeds. The molecular species distribution in individual lipid classes revealed quite distinct patterns and showed where metabolic connections were important. As the seeds developed, the molecular species distributions changed, especially in the period of early (20 days after flowering, DAF) to mid phase (27DAF) of oil accumulation. The patterns of molecular species of diacylglycerol, phosphatidylcholine and acyl-CoAs were used to predict the possible relative contributions of diacylglycerol acyltransferase (DGAT) and phospholipid:diacylglycerol acyltransferase to triacylglycerol production. Our calculations suggest that DGAT may hold a more important role in influencing the molecular composition of TAG. Enzyme selectivity had an important influence on the final molecular species patterns. Our data contribute significantly to our understanding of lipid accumulation in the world’s third most important oil crop

Functional Characterization of the Arabidopsis β-Ketoacyl-Coenzyme A Reductase Candidates of the Fatty Acid Elongase

In plants, very-long-chain fatty acids (VLCFAs; \u3e18 carbon) are precursors of sphingolipids, triacylglycerols, cuticular waxes, and suberin. VLCFAs are synthesized by a multiprotein membrane-bound fatty acid elongation system that catalyzes four successive enzymatic reactions: condensation, reduction, dehydration, and a second reduction. A bioinformatics survey of the Arabidopsis (Arabidopsis thaliana) genome has revealed two sequences homologous to YBR159w encoding a Saccharomyces cerevisiae β-ketoacyl reductase (KCR), which catalyzes the first reduction during VLCFA elongation. Expression analyses showed that both AtKCR1 and AtKCR2 genes were transcribed in siliques, flowers, inflorescence stems, leaves, as well as developing embryos, but only AtKCR1 transcript was detected in roots. Fluorescent protein-tagged AtKCR1 and AtKCR2 were localized to the endoplasmic reticulum, the site of fatty acid elongation. Complementation of the yeast ybr159Δ mutant demonstrated that the two KCR proteins are divergent and that only AtKCR1 can restore heterologous elongase activity similar to the native yeast KCR gene. Analyses of insertional mutants in AtKCR1 and AtKCR2 revealed that loss of AtKCR1 function results in embryo lethality, which cannot be rescued by AtKCR2 expression using the AtKCR1 promoter. In contrast, a disruption of the AtKCR2 gene had no obvious phenotypic effect. Taken together, these results indicate that only AtKCR1 is a functional KCR isoform involved in microsomal fatty acid elongation. To investigate the roles of AtKCR1 in postembryonic development, transgenic lines expressing RNA interference and overexpression constructs targeted against AtKCR1 were generated. Morphological and biochemical characterization of these lines confirmed that suppressed KCR activity results in a reduction of cuticular wax load and affects VLCFA composition of sphingolipids, seed triacylglycerols, and root glycerolipids, demonstrating in planta that KCR is involved in elongation reactions supplying VLCFA for all these diverse classes of lipids

Overexpression of an endogenous type 2 diacylglycerol acyltransferase in the marine diatom Phaeodactylum tricornutum enhances lipid production and omega-3 long-chain polyunsaturated fatty acid content

Background: Oleaginous microalgae represent a valuable resource for the production of high-value molecules. Considering the importance of omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs) for human health and nutrition the yields of high-value eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) require significant improvement to meet demand; however, the current cost of production remains high. A promising approach is to metabolically engineer strains with enhanced levels of triacylglycerols (TAGs) enriched in EPA and DHA. Results: Recently, we have engineered the marine diatom Phaeodactylum tricornutum to accumulate enhanced levels of DHA in TAG. To further improve the incorporation of omega-3 LC-PUFAs in TAG, we focused our effort on the identification of a type 2 acyl-CoA:diacylglycerol acyltransferase (DGAT) capable of improving lipid production and the incorporation of DHA in TAG. DGAT is a key enzyme in lipid synthesis. Following a diatom based in vivo screen of candidate DGATs, a native P. tricornutum DGAT2B was taken forward for detailed characterisation. Overexpression of the endogenous P. tricornutum DGAT2B was confirmed by qRT-PCR and the transgenic strain grew successfully in comparison to wildtype. PtDGAT2B has broad substrate specificity with preferences for C16 and LC-PUFAs acyl groups. Moreover, the overexpression of an endogenous DGAT2B resulted in higher lipid yields and enhanced levels of DHA in TAG. Furthermore, a combined overexpression of the endogenous DGAT2B and ectopic expression of a Δ5-elongase showed how iterative metabolic engineering can be used to increase DHA and TAG content, irrespective of nitrogen treatment. Conclusion: This study provides further insight into lipid metabolism in P. tricornutum and suggests a metabolic engineering approach for the efficient production of EPA and DHA in microalgae

Tailoring seed oil composition in the real world: optimising omega-3 long chain polyunsaturated fatty acid accumulation in transgenic Camelina sativa

There is considerable interest in the de novo production of omega-3 long chain polyunsaturated fatty acids such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), not least of all given the importance of these fatty acids in both aquaculture and human nutrition. Previously we have demonstrated the feasibility of using metabolic engineering in transgenic plants (Camelina sativa) to modify the seed oil composition to now include EPA and/or DHA. In this study, we further tailored the seed oil profile to reduce the omega-6 content, and evaluated the performance of such GM plants under field conditions (i.e. environmental releases), in terms of agronomic performance and also the lipidomic profile of seed oil. We used MALDI- mass spectrometry imaging to identify discrete tissue-types in the seed in which these non-native fatty acids preferentially accumulated. Collectively, these data provide new insights into the complexity of plant lipid metabolism and the challenges associated with predictive manipulation of these pathways. However, this study identified the likely dispensable nature of a Δ12-desturase activity in our omega-3 metabolic engineering rationales for Camelina

Agriculture can help aquaculture become greener

Aquaculture, the farming of fish and seafood, is recognized as a highly efficient system for producing protein for human consumption. In contrast, many terrestrial animal protein production systems are inefficient, impacting land use and exacerbating climate change. Humankind needs to adopt a more plant-centric diet, the only exception being fish consumed as both a source of protein and essential dietary nutrients such as omega-3 fatty acids. Here we consider the implications of such a transition, and the challenges that aquaculture must overcome to increase productivity within planetary boundaries. We consider how agriculture, specifically crops, can provide solutions for aquaculture, especially the sectors that are dependent on marine ingredients. For example, agriculture can provide experience with managing monocultures and new technologies such as genetically modified crops tailored specifically for use in aquaculture. We propose that a closer connection between agriculture and aquaculture will create a resilient food system capable of meeting increasing dietary and nutritional demands without exhausting planetary resources

Estimating the bispectrum of the Very Small Array data

We estimate the bispectrum of the Very Small Array data from the compact and extended configuration observations released in December 2002, and compare our results to those obtained from Gaussian simulations. There is a slight excess of large bispectrum values for two individual fields, but this does not appear when the fields are combined. Given our expected level of residual point sources, we do not expect these to be the source of the discrepancy. Using the compact configuration data, we put an upper limit of 5400 on the value of f_NL, the non-linear coupling parameter, at 95 per cent confidence. We test our bispectrum estimator using non-Gaussian simulations with a known bispectrum, and recover the input values.Comment: 17 pages, 16 figures, replaced with version accepted by MNRAS. Primordial bispectrum recalculated and figure 11 change

High sensitivity measurements of the CMB power spectrum with the extended Very Small Array

We present deep Ka-band ($\nu \approx 33$ GHz) observations of the CMB made with the extended Very Small Array (VSA). This configuration produces a naturally weighted synthesized FWHM beamwidth of $\sim 11$ arcmin which covers an $\ell$-range of 300 to 1500. On these scales, foreground extragalactic sources can be a major source of contamination to the CMB anisotropy. This problem has been alleviated by identifying sources at 15 GHz with the Ryle Telescope and then monitoring these sources at 33 GHz using a single baseline interferometer co-located with the VSA. Sources with flux densities \gtsim 20 mJy at 33 GHz are subtracted from the data. In addition, we calculate a statistical correction for the small residual contribution from weaker sources that are below the detection limit of the survey. The CMB power spectrum corrected for Galactic foregrounds and extragalactic point sources is presented. A total $\ell$-range of 150-1500 is achieved by combining the complete extended array data with earlier VSA data in a compact configuration. Our resolution of $\Delta \ell \approx 60$ allows the first 3 acoustic peaks to be clearly delineated. The is achieved by using mosaiced observations in 7 regions covering a total area of 82 sq. degrees. There is good agreement with WMAP data up to $\ell=700$ where WMAP data run out of resolution. For higher $\ell$-values out to $\ell = 1500$, the agreement in power spectrum amplitudes with other experiments is also very good despite differences in frequency and observing technique.Comment: 16 pages. Accepted in MNRAS (minor revisions