Metabolic engineering of biomass for high energy density: oilseed-like triacylglycerol yields from plant leaves.
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ABSTRACT:
SUBMITTER: Vanhercke T
PROVIDER: S-EPMC4285938 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The synthesis of fatty acids and glycerolipids in wild-type Arabidopsis leaves does not typically lead to strong triacylglycerol (TAG) accumulation. LEAFY COTYLEDON2 (LEC2) is a master regulator of seed maturation and oil accumulation in seeds. Constitutive ectopic LEC2 expression causes somatic embryogenesis and defects in seedling growth. Here, we report that senescence-inducible LEC2 expression caused a threefold increase in TAG levels in transgenic leaves compared with that in the leaves of wild-type plants. Plant growth was not severely affected by the accumulation the TAG in response to LEC2 expression. The levels of plastid-synthesized lipids, mono- and di-galactosyldiacylglycerol and phosphatidylglycerol were reduced more in senescence-induced LEC2 than in endoplasmic reticulum-synthesized lipids, including phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol. Senescence-induced LEC2 up-regulated the expression of many genes involved in fatty acid and TAG biosynthesis at precise times in senescent leaves, including WRINKLED1 (WRI1), which encodes a fatty acid transcription factor. The expressions of glycerol-3-phosphate dehydrogenase 1 and phospholipid:diacylglycerol 2 were increased in the transgenic leaves. Five seed-type oleosin-encoding genes, expressed during oil-body formation, and the seed-specific FAE1 gene, which encodes the enzyme responsible for the synthesis of C20:1 and C22:1 fatty acids, were also expressed at higher levels in senescing transgenic leaves than in wild-type leaves. Senescence-inducible LEC2 triggers the key metabolic steps that increase TAG accumulation in vegetative tissues.
| S-EPMC5448714 | biostudies-literature
Project description:To evaluate the relative contribution of individual sugar catabolic pathways to the overall physiology of A. niger during growth on plant biomass, the A. niger sugar catabolic pathways involved in converting the most common monomers of plant biomass polysaccharides were blocked, and the phenotype as well as the transcriptomic response of these single-pathway deletion mutants was analysed on two distinct plant biomass substrates: wheat bran (WB) and sugar beet pulp (SBP). Deletion of all the above catabolic pathways were also combined in a single strain (?all), to evaluate whether other compounds of plant biomass could still support growth of A. niger when utilization of the major sugar components is blocked. On both WB and SBP, the strongest affected single-pathway deletion mutants were the PCP and glycolysis deficient mutants, delta-xkiA and delta-hxkA/delta-glkA, respectively. Especially on WB, which is a particularly rich substrate in pentose sugars, blocking PCP caused a significant effect in delta-xkiA with respect to both growth and transcriptomic response. However, delta-hxkA/delta-glkA was shown to be the most critically affected strain. On both substrates, combined deletion of hxkA and glkA resulted in the strongest growth reduction of all the single-pathway mutants, very similar to the multi-pathway deletion mutant delta-all. This result clearly pointed to glycolysis as the most essential and indispensable pathway for the survival of A. niger on these substrates. Since cellulose is not a preferred carbon source for A. niger, it seems that the double deletion of hxkA and glkA mostly affected the utilization of starch. (Note: A portion of this research was performed on a project award (10.46936/expl.proj.2019.51072/60006675) from the Environmental Molecular Sciences Laboratory, a DOE Office of Science User Facility sponsored by the Biological and Environmental Research program under Contract No. DE-AC05-76RL01830)
2022-08-31 | GSE196397 | GEO
Project description:The great potential of pharmacologically active secondary plant metabolites is often limited by low yield and availability of the producing plant. Chemical synthesis of these complex compounds is often too expensive. Plant cell fermentation offers an alternative strategy to overcome these limitations. However, production in batch cell cultures remains often inefficient. One reason might be the fact that different cell types have to interact for metabolite maturation, which is poorly mimicked in suspension cell lines. Using alkaloid metabolism of tobacco, we explore an alternative strategy, where the metabolic interactions of different cell types in a plant tissue are technically mimicked based on different plant-cell based metabolic modules. In this study, we simulate the interaction found between the nicotine secreting cells of the root and the nicotine-converting cells of the senescent leaf, generating the target compound nornicotine in the model cell line tobacco BY-2. When the nicotine demethylase NtomCYP82E4 was overexpressed in tobacco BY-2 cells, nornicotine synthesis was triggered, but only to a minor extent. However, we show here that we can improve the production of nornicotine in this cell line by feeding the precursor, nicotine. Engineering of another cell line overexpressing the key enzyme NtabMPO1 allows to stimulate accumulation and secretion of this precursor. We show that the nornicotine production of NtomCYP82E4 cells can be significantly stimulated by feeding conditioned medium from NtabMPO1 overexpressors without any negative effect on the physiology of the cells. Co-cultivation of NtomCYP82E4 with NtabMPO1 stimulated nornicotine accumulation even further, demonstrating that the physical presence of cells was superior to just feeding the conditioned medium collected from the same cells. These results provide a proof of concept that combination of different metabolic modules can improve the productivity for target compounds in plant cell fermentation.
| S-EPMC5231347 | biostudies-literature
Project description:Feedstocks for industrial applications ranging from polymers to lubricants are largely derived from petroleum, a non-renewable resource. Vegetable oils with fatty acid structures and storage forms tailored for specific industrial uses offer renewable and potentially sustainable sources of petrochemical-type functionalities. A wide array of industrial vegetable oils can be generated through biotechnology, but will likely require non-commodity oilseed platforms dedicated to specialty oil production for commercial acceptance. Here we show the feasibility of three Brassicaceae oilseeds crambe, camelina, and carinata, none of which are widely cultivated for food use, as hosts for complex metabolic engineering of wax esters for lubricant applications. Lines producing wax esters >20% of total seed oil were generated for each crop and further improved for high temperature oxidative stability by down-regulation of fatty acid polyunsaturation. Field cultivation of optimized wax ester-producing crambe demonstrated commercial utility of these engineered crops and a path for sustainable production of other industrial oils in dedicated specialty oilseeds.
| S-EPMC4768164 | biostudies-other
Project description:Background and aimsPlant biomass-density relationships during self-thinning are determined mainly by allometry. Both allometry and biomass-density relationship have been shown to vary with abiotic conditions, but the effects of biotic interactions have not been investigated. Arbuscular mycorrhizal fungi (AMF) can promote plant growth and affect plant form. Here experiments were carried out to test whether AMF affect plant allometry and the self-thinning trajectory.MethodsTwo experiments were conducted on Medicago sativa L., a leguminous species known to be highly dependent on mycorrhiza. Two mycorrhizal levels were obtained by applying benomyl (low AMF) or not (high AMF). Experiment 1 investigated the effects of AMF on plant growth in the absence of competition. Experiment 2 was a factorial design with two mycorrhizal levels and two plant densities (6000 and 17 500 seeds m(-2)). Shoot biomass, root biomass and canopy radius were measured 30, 60, 90 and 120 d after sowing. The allometric relationships among these aspects of size were estimated by standardized major axis regression on log-transformed data.Key resultsShoot biomass in the absence of competition was lower under low AMF treatment. In self-thinning populations, the slope of the log (mean shoot biomass) vs. log density relationship was significantly steeper for the high AMF treatment (slope = -1·480) than for the low AMF treatment (-1·133). The canopy radius-biomass allometric exponents were not significantly affected by AMF level, but the root-shoot allometric exponent was higher in the low AMF treatment. With a high level of AMF, the biomass-density exponent can be predicted from the above-ground allometric model of self-thinning, while this was not the case when AMF were reduced by fungicide.ConclusionsAMF affected the importance of below-ground relative to above-ground interactions and changed root vs. shoot allocation. This changed allometric allocation of biomass and altered the self-thinning trajectory.
| S-EPMC3043928 | biostudies-literature
Project description:Background:Many microbes used for the rapid discovery and development of metabolic pathways have sensitivities to final products and process reagents. Isopentenol (3-methyl-3-buten-1-ol), a biogasoline candidate, has an established heterologous gene pathway but is toxic to several microbial hosts. Reagents used in the pretreatment of plant biomass, such as ionic liquids, also inhibit growth of many host strains. We explored the use of Corynebacterium glutamicum as an alternative host to address these constraints. Results:We found C. glutamicum ATCC 13032 to be tolerant to both the final product, isopentenol, as well to three classes of ionic liquids. A heterologous mevalonate-based isopentenol pathway was engineered in C. glutamicum. Targeted proteomics for the heterologous pathway proteins indicated that the 3-hydroxy-3-methylglutaryl-coenzyme A reductase protein, HmgR, is a potential rate-limiting enzyme in this synthetic pathway. Isopentenol titers were improved from undetectable to 1.25 g/L by combining three approaches: media optimization; substitution of an NADH-dependent HmgR homolog from Silicibacter pomeroyi; and development of a C. glutamicum ∆poxB ∆ldhA host chassis. Conclusions:We describe the successful expression of a heterologous mevalonate-based pathway in the Gram-positive industrial microorganism, C. glutamicum, for the production of the biogasoline candidate, isopentenol. We identified critical genetic factors to harness the isopentenol pathway in C. glutamicum. Further media and cultivation optimization enabled isopentenol production from sorghum biomass hydrolysates.
| S-EPMC6391826 | biostudies-other
Project description:The ability to enhance photosynthetic capacity remains a recognized bottleneck to improving plant productivity. Phototropin blue light receptors (phot1 and phot2) optimize photosynthetic efficiency in Arabidopsis thaliana by coordinating multiple light-capturing processes. In this study, we explore the potential of using protein engineering to improve photoreceptor performance and thereby plant growth. We demonstrate that targeted mutagenesis can decrease or increase the photocycle lifetime of Arabidopsis phototropins in vitro and show that these variants can be used to reduce or extend the duration of photoreceptor activation in planta Our findings show that slowing the phototropin photocycle enhanced several light-capturing responses, while accelerating it reduced phototropin's sensitivity for chloroplast accumulation movement. Moreover, plants engineered to have a slow-photocycling variant of phot1 or phot2 displayed increased biomass production under low-light conditions as a consequence of their improved sensitivity. Together, these findings demonstrate the feasibility of engineering photoreceptors to manipulate plant growth and offer additional opportunities to enhance photosynthetic competence, particularly under suboptimal light regimes.
| S-EPMC6589663 | biostudies-literature
Project description:Sesame is considered as the queen of oil seeds owing to its high oil content of about 56-62% and good quality oil. Sesame oil alone or in combination with other vegetable oils can yield good quality biodiesel. Sesame biodiesel blends up to 20% yields fuel efficiency and power output on par with mineral diesel but superior in environmental performance. Though biodiesel from edible oil is highly criticized, the demand for renewable energy compels the development of high-performance sesame plants. Triacylglycerol synthesis in flowering plants follows an acyl CoA-dependent and independent manner. This study envisages transgenic approaches to enhance oil production in sesame biomass. The genes of choice for oil enhancement includes DGAT1, PDAT1, FAD3 and cytochrome b5F. Diacylglycerol acyltransferase (DGAT) and phospholipid diacylglycerol acyltransferases (PDAT) are key enzymes in TAG synthesis. Fatty acid desaturases (FAD) has the ability to enhance specific fatty acids, whereas cytochrome b5 genes augment the process by donating electrons. A combination of the above categories of genes which performed well in terms of oil content in the yeast expression system from our earlier studies is used in Agrobacterium-mediated sesame transformation experiments to evaluate the biodiesel potential of transgenic sesame plants. The transgenic construct with PDAT1 and FAD3 combination yielded a 10% increase in TAG content. The possibility of transgenic sesame as a biodiesel plant is discussed.
| S-EPMC8449027 | biostudies-literature
Project description:Triacylglycerol (TAG), typically represents <1% of leaf glycerolipids but can accumulate under stress and other conditions or if leaves are supplied with fatty acids, or in plants transformed with regulators or enzymes of lipid metabolism. To better understand the metabolism of TAG in leaves, pulse-chase radiolabelling experiments were designed to probe its synthesis and turnover. When Arabidopsis leaves were incubated with [(14)C]lauric acid (12:0), a major initial product was [(14)C]TAG. Thus, despite low steady-state levels, leaves possess substantial TAG biosynthetic capacity. The contributions of diacylglycerol acyltransferase1 and phospholipid:diacylglycerol acyltransferase1 to leaf TAG synthesis were examined by labelling of dgat1 and pdat1 mutants. The dgat1 mutant displayed a major (76%) reduction in [(14)C]TAG accumulation whereas pdat1 TAG labelling was only slightly reduced. Thus, DGAT1 has a principal role in TAG biosynthesis in young leaves. During a 4h chase period, radioactivity in TAG declined 70%, whereas the turnover of [(14)C]acyl chains of phosphatidylcholine (PC) and other polar lipids was much lower. Sixty percent of [(14)C]12:0 was directly incorporated into glycerolipids without modification, whereas 40% was elongated and desaturated to 16:0 and 18:1 by plastids. The unmodified [(14)C]12:0 and the plastid products of [(14)C]12:0 metabolism entered different pathways. Although plastid-modified (14)C-labelled products accumulated in monogalactosyldiacylglycerol, PC, phosphatidylethanolamine, and diacylglcerol (DAG), there was almost no accumulation of [(14)C]16:0 and [(14)C]18:1 in TAG. Because DAG and acyl-CoA are direct precursors of TAG, the differential labelling of polar glycerolipids and TAG by [(14)C]12:0 and its plastid-modified products provides evidence for multiple subcellular pools of both acyl-CoA and DAG.
| S-EPMC4339603 | biostudies-literature
Project description:Camelina sativa (L.) Crantz is an important Brassicaceae oil crop with a number of excellent agronomic traits including low water and fertilizer input, strong adaptation and resistance. Furthermore, its short life cycle and easy genetic transformation, combined with available data of genome and other "-omics" have enabled camelina as a model oil plant to study lipid metabolism regulation and genetic improvement. Particularly, camelina is capable of rapid metabolic engineering to synthesize and accumulate high levels of unusual fatty acids and modified oils in seeds, which are more stable and environmentally friendly. Such engineered camelina oils have been increasingly used as the super resource for edible oil, health-promoting food and medicine, biofuel oil and high-valued chemical production. In this review, we mainly highlight the latest advance in metabolic engineering towards the predictive manipulation of metabolism for commercial production of desirable bio-based products using camelina as an ideal platform. Moreover, we deeply analysis camelina seed metabolic engineering strategy and its promising achievements by describing the metabolic assembly of biosynthesis pathways for acetyl glycerides, hydroxylated fatty acids, medium-chain fatty acids, ?-3 long-chain polyunsaturated fatty acids, palmitoleic acid (?-7) and other high-value oils. Future prospects are discussed, with a focus on the cutting-edge techniques in camelina such as genome editing application, fine directed manipulation of metabolism and future outlook for camelina industry development.
| S-EPMC7028685 | biostudies-literature