WRINKLED1 Transcription Factor Research Articles

Possible crosstalk between the Arabidopsis TSPO-related protein and the transcription factor WRINKLED1

This study uncovers a regulatory interplay between WRINKLED1 (WRI1), a master transcription factor for glycolysis and lipid biosynthesis, and Translocator Protein (TSPO) expression in Arabidopsis thaliana seeds. We identified potential WRI1-responsive elements upstream of AtTSPO through bioinformatics, suggesting WRI1's involvement in regulating TSPO expression. Our analyses showed a significant reduction in AtTSPO levels in wri1 mutant seeds compared to wild type, establishing a functional link between WRI1 and TSPO. This connection extends to the coordination of seed development and lipid metabolism, with both WRI1 and AtTSPO levels decreasing post-imbibition, indicating their roles in seed physiology. Further investigations into TSPO's impact on fatty acid synthesis revealed that TSPO misexpression alters WRI1's post-translational modifications and significantly enhances seed oil content. Additionally, we noted a decrease in key reserve proteins, including 12 S globulin and oleosin 1, in seeds with TSPO misexpression, suggesting a novel energy storage strategy in these lines. Our findings reveal a sophisticated network involving WRI1 and AtTSPO, highlighting their crucial contributions to seed development, lipid metabolism, and the modulation of energy storage mechanisms in Arabidopsis.

Intron-mediated enhancement of DIACYLGLYCEROL ACYLTRANSFERASE1 expression in energycane promotes a step change for lipid accumulation in vegetative tissues

BackgroundMetabolic engineering for hyperaccumulation of lipids in vegetative tissues is a novel strategy for enhancing energy density and biofuel production from biomass crops. Energycane is a prime feedstock for this approach due to its high biomass production and resilience under marginal conditions. DIACYLGLYCEROL ACYLTRANSFERASE (DGAT) catalyzes the last and only committed step in the biosynthesis of triacylglycerol (TAG) and can be a rate-limiting enzyme for the production of TAG.ResultsIn this study, we explored the effect of intron-mediated enhancement (IME) on the expression of DGAT1 and resulting accumulation of TAG and total fatty acid (TFA) in leaf and stem tissues of energycane. To maximize lipid accumulation these evaluations were carried out by co-expressing the lipogenic transcription factor WRINKLED1 (WRI1) and the TAG protect factor oleosin (OLE1). Including an intron in the codon-optimized TmDGAT1 elevated the accumulation of its transcript in leaves by seven times on average based on 5 transgenic lines for each construct. Plants with WRI1 (W), DGAT1 with intron (Di), and OLE1 (O) expression (WDiO) accumulated TAG up to a 3.85% of leaf dry weight (DW), a 192-fold increase compared to non-modified energycane (WT) and a 3.8-fold increase compared to the highest accumulation under the intron-less gene combination (WDO). This corresponded to TFA accumulation of up to 8.4% of leaf dry weight, a 2.8-fold or 6.1-fold increase compared to WDO or WT, respectively. Co-expression of WDiO resulted in stem accumulations of TAG up to 1.14% of DW or TFA up to 2.08% of DW that exceeded WT by 57-fold or 12-fold and WDO more than twofold, respectively. Constitutive expression of these lipogenic “push pull and protect” factors correlated with biomass reduction.ConclusionsIntron-mediated enhancement (IME) of the expression of DGAT resulted in a step change in lipid accumulation of energycane and confirmed that under our experimental conditions it is rate limiting for lipid accumulation. IME should be applied to other lipogenic factors and metabolic engineering strategies. The findings from this study may be valuable in developing a high biomass feedstock for commercial production of lipids and advanced biofuels.Graphical abstract

The molecular mechanism of WRINKLED1 transcription factor regulating oil accumulation in developing seeds of castor bean

The transcription factor WRINKLED1 (WRI1), a member of AP2 gene family that contain typical AP2 domains, has been considered as a master regulator regulating oil biosynthesis in oilseeds. However, the regulatory mechanism of RcWRI1 in regulating oil accumulation during seed development has not been clearly addressed. Castor bean (Ricinus communis) is one of the most important non-edible oil crops and its seed oils are rich in hydroxy fatty acids, widely applied in industry. In this study, based on castor bean reference genome, three RcWRIs genes (RcWRI1, RcWRI2 and RcWRI3) were identified and the expressed association of RcWRI1 with oil accumulation were determined. Heterologous transformation of RcWRI1 significantly increased oil content in tobacco leaf, confirming that RcWRI1 activate lipid biosynthesis pathway. Using DNA Affinity Purification sequencing (DAP-seq) technology, we confirmed RcWRI1 binding with Transcription Start Site of genes and identified 7961 WRI1-binding candidate genes. Functionally, these identified genes were mainly involved in diverse metabolism pathways (including lipid biosynthesis). Three cis-elements AW-box ([CnTnG](n)7[CG]) and AW-boxes like ([GnAnC](n)6[GC]/[GnAnC](n)7[G]) bound with RcWRI1 were identified. Co-expression network analysis of RcWRI1 further found that RcWRI1 might be widely involved in biosynthesis of storage materials during seed development. In particular, yeast one hybrid experiments found that both AP2 domains within RcWRI1 were required in binding targeted genes. These results not only provide new evidence to understand the regulatory mechanism of RcWRI1 in regulation of oil accumulation during castor bean seed development, but also give candidate gene resource for subsequent genetic improvement toward increasing oil content in oilseed crops.

An expanded role for the transcription factor WRINKLED1 in the biosynthesis of triacylglycerols during seed development.

The transcription factor WRINKLED1 (WRI1) is known as a master regulator of fatty acid synthesis in developing oilseeds of Arabidopsis thaliana and other species. WRI1 is known to directly stimulate the expression of many fatty acid biosynthetic enzymes and a few targets in the lower part of the glycolytic pathway. However, it remains unclear to what extent and how the conversion of sugars into fatty acid biosynthetic precursors is controlled by WRI1. To shortlist possible gene targets for future in-planta experimental validation, here we present a strategy that combines phylogenetic foot printing of cis-regulatory elements with additional layers of evidence. Upstream regions of protein-encoding genes in A. thaliana were searched for the previously described DNA-binding consensus for WRI1, the ASML1/WRI1 (AW)-box. For about 900 genes, AW-box sites were found to be conserved across orthologous upstream regions in 11 related species of the crucifer family. For 145 select potential target genes identified this way, affinity of upstream AW-box sequences to WRI1 was assayed by Microscale Thermophoresis. This allowed definition of a refined WRI1 DNA-binding consensus. We find that known WRI1 gene targets are predictable with good confidence when upstream AW-sites are phylogenetically conserved, specifically binding WRI1 in the in vitro assay, positioned in proximity to the transcriptional start site, and if the gene is co-expressed with WRI1 during seed development. When targets predicted in this way are mapped to central metabolism, a conserved regulatory blueprint emerges that infers concerted control of contiguous pathway sections in glycolysis and fatty acid biosynthesis by WRI1. Several of the newly predicted targets are in the upper glycolysis pathway and the pentose phosphate pathway. Of these, plastidic isoforms of fructokinase (FRK3) and of phosphoglucose isomerase (PGI1) are particularly corroborated by previously reported seed phenotypes of respective null mutations.

The Sunflower WRINKLED1 Transcription Factor Regulates Fatty Acid Biosynthesis Genes through an AW Box Binding Sequence with a Particular Base Bias.

Sunflower is an important oilseed crop in which the biochemical pathways leading to seed oil synthesis and accumulation have been widely studied. However, how these pathways are regulated is less well understood. The WRINKLED1 (WRI1) transcription factor is considered a key regulator in the control of triacylglycerol biosynthesis, acting through the AW box binding element (CNTNG(N)7CG). Here, we identified the sunflower WRI1 gene and characterized its activity in electrophoretic mobility shift assays. We studied its role as a co-regulator of sunflower genes involved in plastidial fatty acid synthesis. Sunflower WRI1-targets included genes encoding the pyruvate dehydrogenase complex, the α-CT and BCCP genes, genes encoding ACPs and the fatty acid synthase complex, together with the FATA1 gene. As such, sunflower WRI1 regulates genes involved in seed plastidial fatty acid biosynthesis in a coordinated manner, establishing a WRI1 push and pull strategy that drives oleic acid synthesis for its export into the cytosol. We also determined the base bias at the N positions in the active sunflower AW box motif. The sunflower AW box is sequence-sensitive at the non-conserved positions, enabling WRI1-binding. Moreover, sunflower WRI1 could bind to a non-canonical AW-box motif, opening the possibility of searching for new target genes.

Analysis and Functional Verification of PoWRI1 Gene Associated with Oil Accumulation Process in Paeonia ostii.

The plant transcription factor WRINKLED1 (WRI1), a member of AP2/EREBP, is involved in the regulation of glycolysis and the expression of genes related to the de novo synthesis of fatty acids in plastids. In this study, the key regulator of seed oil synthesis and accumulation transcription factor gene PoWRI1 was identified and cloned, having a complete open reading frame of 1269 bp and encoding 422 amino acids. Subcellular localization analysis showed that PoWRI1 is located at the nucleus. After the expression vector of PoWRI1 was constructed and transformed into wild-type Arabidopsis thaliana, it was found that the overexpression of PoWRI1 increased the expression level of downstream target genes such as BCCP2, KAS1, and PKP-β1. As a result, the seeds of transgenic plants became larger, the oil content increased significantly, and the unsaturated fatty acid content increased, which provide a scientific theoretical basis for the subsequent use of genetic engineering methods to improve the fatty acid composition and content of plant seeds.

Differential Activation of Partially Redundant Δ9 Stearoyl-ACP Desaturase Genes Is Critical for Omega-9 Monounsaturated Fatty Acid Biosynthesis During Seed Development in Arabidopsis.

The spatiotemporal pattern of deposition, final amount, and relative abundance of oleic acid (cis-ω-9 C18:1) and its derivatives in the different lipid fractions of the seed of Arabidopsis (Arabidopsis thaliana) indicates that omega-9 monoenes are synthesized at high rates in this organ. Accordingly, we observed that four Δ9 stearoyl-ACP desaturase (SAD)-coding genes (FATTY ACID BIOSYNTHESIS2 [FAB2], ACYL-ACYL CARRIER PROTEIN5 [AAD5], AAD1, and AAD6) are transcriptionally induced in seeds. We established that the three most highly expressed ones are directly activated by the WRINKLED1 transcription factor. We characterized a collection of 30 simple, double, triple, and quadruple mutants affected in SAD-coding genes and thereby revealed the functions of these desaturases throughout seed development. Production of oleic acid by FAB2 and AAD5 appears to be critical at the onset of embryo morphogenesis. Double homozygous plants from crossing fab2 and aad5 could never be obtained, and further investigations revealed that the double mutation results in the arrest of embryo development before the globular stage. During later stages of seed development, these two SADs, together with AAD1, participate in the elaboration of the embryonic cuticle, a barrier essential for embryo-endosperm separation during the phase of invasive embryo growth through the endosperm. This study also demonstrates that the four desaturases redundantly contribute to storage lipid production during the maturation phase.

Molecular Basis of Plant Oil Biosynthesis: Insights Gained From Studying the WRINKLED1 Transcription Factor.

Most plant species generate and store triacylglycerol (TAG) in their seeds, serving as a core supply of carbon and energy to support seedling development. Plant seed oils have a wide variety of applications, from being essential for human diets to serving as industrial renewable feedstock. WRINKLED1 (WRI1) transcription factor plays a central role in the transcriptional regulation of plant fatty acid biosynthesis. Since the discovery of Arabidopsis WRI1 gene (AtWRI1) in 2004, the function of WRI1 in plant oil biosynthesis has been studied intensively. In recent years, the identification of WRI1 co-regulators and deeper investigations of the structural features and molecular functions of WRI1 have advanced our understanding of the mechanism of the transcriptional regulation of plant oil biosynthesis. These advances also help pave the way for novel approaches that will better utilize WRI1 for bioengineering oil production in crops.

Expression of the Arabidopsis WRINKLED 1 transcription factor leads to higher accumulation of palmitate in soybean seed.

SummarySoybean (Glycine max [L.] Merr.) is a commodity crop highly valued for its protein and oil content. The high percentage of polyunsaturated fatty acids in soybean oil results in low oxidative stability, which is a key parameter for usage in baking, high temperature frying applications, and affects shelf life of packaged products containing soybean oil. Introduction of a seed‐specific expression cassette carrying the Arabidopsis transcription factor WRINKLED1 (AtWRI1) into soybean, led to seed oil with levels of palmitate up to approximately 20%. Stacking of the AtWRI1 transgenic allele with a transgenic locus harbouring the mangosteen steroyl‐ACP thioesterase (GmFatA) resulted in oil with total saturates up to 30%. The creation of a triple stack in soybean, wherein the AtWRI1 and GmFatA alleles were combined with a FAD2‐1 silencing allele led to the synthesis of an oil with 28% saturates and approximately 60% oleate. Constructs were then assembled that carry a dual FAD2‐1 silencing element/GmFatA expression cassette, alone or combined with an AtWRI1 cassette. These plasmids are designated pPTN1289 and pPTN1301, respectively. Transgenic events carrying the T‐DNA of pPTN1289 displayed an oil with stearate levels between 18% and 25%, and oleate in the upper 60%, with reduced palmitate (<5%). While soybean events harboring transgenic alleles of pPTN1301 had similar levels of stearic and oleate levels as that of the pPTRN1289 events, but with levels of palmitate closer to wild type. The modified fatty acid composition results in an oil with higher oxidative stability, and functionality attributes for end use in baking applications.

Overexpression of a Transcription Factor Increases Lipid Content in a Woody Perennial Jatropha curcas.

Vegetable oil is an important renewable resource for dietary consumption for human and livestock, and more recently for biodiesel production. Lipid traits in crops are controlled by multiple quantitative trait loci (QTLs) and each of them has a small effect on lipid traits. So far, there is limited success to increase lipid yield and improve lipid quality in plants. Here, we reported the identification of a homolog of APETALA2 (AP2) transcription factor WRINKLED1 (JcWRI1) from an oleaginous plant Jatropha curcas and characterized its function in Jatropha and Arabidopsis thaliana. Using physical mapping data, we located JcWRI1 in a QTL region specifying high oleate and lipid content in Jatropha. Overexpression of JcWRI1 in Jatropha elevated seed lipid content and increased seed mass. Lipid profile in seeds of over-expression plants showed higher oleate content which will be beneficial to improve biodiesel quality. Overexpression of JcWRI1 activated lipid-related gene expression and JcWRI1 was shown to directly bind to the AW-box of promoters of some of these genes. In conclusion, we were able to increase seed lipid content and improve seed lipid quality in Jatropha by manipulating one key transcription factor JcWRI1.

Soybean (Glycine max) WRINKLED1 transcription factor, GmWRI1a, positively regulates seed oil accumulation.

Soybean is the world's most important leguminous crop producing high-quality protein and oil. Elevating oil accumulation in soybean seed is always many researchers' goal. WRINKLED1 (WRI1) encodes a transcription factor of the APETALA2/ethylene responsive element-binding protein (AP2/EREBP) family that plays important roles during plant seed oil accumulation. In this study, we isolated and characterized three distinct orthologues of WRI1 in soybean (Glycine max) that display different organ-specific expression patterns, among which GmWRI1a was highly expressed in maturing soybean seed. Electrophoretic mobility shift assays and yeast one-hybrid experiments demonstrated that the GmWRI1a protein was capable of binding to AW-box, a conserved sequence in the proximal upstream regions of many genes involved in various steps of oil biosynthesis. Transgenic soybean seeds overexpressing GmWRI1a under the control of the seed-specific napin promoter showed the increased total oil and fatty acid content and the changed fatty acid composition. Furthermore, basing on the activated expressions in transgenic soybean seeds and existence of AW-box element in the promoter regions, direct downstream genes of GmWRI1a were identified, and their products were responsible for fatty acid production, elongation, desaturation and export from plastid. We conclude that GmWRI1a transcription factor can positively regulate oil accumulation in soybean seed by a complex gene expression network related to fatty acid biosynthesis.

Sugar Potentiation of Fatty Acid and Triacylglycerol Accumulation.

Photosynthetically derived sugar provides carbon skeletons for lipid biosynthesis. We used mutants of Arabidopsis (Arabidopsis thaliana) and the expression of oleogenic factors to investigate relationships among sugar availability, lipid synthesis, and the accumulation of triacylglycerol (TAG) in leaf tissue. The adg1 mutation disables the small subunit of ADP-glucose pyrophosphorylase, the first step in starch synthesis, and the suc2 mutation disables a sucrose/proton symporter that facilitates sucrose loading from leaves into phloem. The adg1suc2 double mutant increases glucose plus sucrose content in leaves 80-fold relative to the wild type, total fatty acid (FA) content 1.8-fold to 8.3% dry weight, and TAG more than 10-fold to 1.2% dry weight. The WRINKLED1 transcription factor also accumulates to higher levels in these leaves, and the rate of FA synthesis increases by 58%. Adding tt4, which disables chalcone synthase, had little effect, but adding the tgd1 mutation, which disables an importer of lipids into plastids to create adg1suc2tt4tgd1, increased total leaf FA to 13.5% dry weight and TAG to 3.8% dry weight, demonstrating a synergistic effect upon combining these mutations. Combining adg1suc2 with the sdp1 mutation, deficient in the predominant TAG lipase, had little effect on total FA content but increased the TAG accumulation by 66% to 2% dry weight. Expression of the WRINKLED1 transcription factor, along with DIACYLGLYCEROL ACYLTRANSFERASE1 and the OLEOSIN1 oil body-associated protein, in the adg1suc2 mutant doubled leaf FA content and increased TAG content to 2.3% dry weight, a level 4.6-fold higher than that resulting from expression of the same factors in the wild type.

Auxin transport and conjugation caught together

Polar auxin transport together with local synthesis and turnover are crucial for establishing auxin gradients, which determine an array of plant developmental pathways. Transcription factor-mediated control of the genes involved in these processes is steadily receiving increased attention, including efforts to find regulators determining the expression of major auxin efflux carriers of the PIN family. Now, Kong et al. (2017) have provided evidence of a direct link between auxin metabolism and transport mediated by PINs that is controlled by the transcription factor WRINKLED1.

The Arabidopsis WRINKLED1 transcription factor affects auxin homeostasis in roots.

WRINKLED1 (WRI1) is a key transcriptional regulator of fatty acid biosynthesis genes in diverse oil-containing tissues. Loss of function of Arabidopsis WRI1 leads to a reduction in the expression of genes for fatty acid biosynthesis and glycolysis, and concomitant strong reduction of seed oil content. The wri1-1 loss-of-function mutant shows reduced primary root growth and decreased acidification of the growth medium. The content of a conjugated form of the plant growth hormone auxin, indole-3-acetic acid (IAA)-Asp, was higher in wri1-1 plants compared with the wild-type. GH3.3, a gene encoding an enzyme involved in auxin degradation, displayed higher expression in the wri1-1 mutant. EMSAs demonstrated that AtWRI1 bound to the promoter of GH3.3. Specific AtWRI1-binding motifs were identified in the promoter of GH3.3. In addition, wri1-1 displayed decreased auxin transport. Expression of some PIN genes, which encode IAA carrier proteins, was reduced in wri1-1 plants as well. Correspondingly, AtWRI1 bound to the promoter regions of some PIN genes. It is well known that auxin exerts its maximum effects at a specific, optimal concentration in roots requiring a finely balanced auxin homeostasis. This process appears to be disrupted when the expression of WRI1 and in turn a subset of its target genes are misregulated, highlighting a role for WRI1 in root auxin homeostasis.

Comparative transcriptomics reveals molecular components associated with differential lipid accumulation between microalgal sp., Scenedesmus dimorphus and Scenedesmus quadricauda

Scenedesmus is considered as a potential oil-producing green microalgae having higher lipid content with suitable fatty acid profile (high oleic acid) for biodiesel production. Comparative transcriptome analysis of two Scenedesmus sp., viz. Scenedesmus dimorphus (26%) and Scenedesmus quadricauda (14%) having equivalent biomass and variable lipid content was performed to uncover molecular mechanisms controlling differential lipid production. 76,969 and 40,979 CDSs were predicted from the transcriptomes of S. dimorphus and S. quadricauda respectively, which were subsequently mapped to metabolic pathways. Overall up-regulation of metabolic pathways contributing precursors to storage lipid biosynthesis was observed in S. dimorphus. Glyceraldehyde 3-phosphate dehydrogenase, enolase, acetyl-CoA synthetase, pyruvate dehydrogenase, ATP citrate lyase, glycerol kinase, citrate synthase were identified as major regulators of high lipid content in S. dimorphus. Further, WRINKLED1 transcription factor significantly correlated with high lipid accumulation as revealed by reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) in three growth stages of two species. This study provides a broad view of storage lipid production in Scenedesmus species with potential implications in designing suitable genetic interventions towards increase in lipid content vis-à-vis central carbon metabolism.

Cumulative effect of heterologous AtWRI1 gene expression and endogenous BjAGPase gene silencing increases seed lipid content in Indian mustard Brassica juncea

The production of vegetable oil in many countries of the world, including India has not been able to keep pace with the increasing requirement, leading to a very large gap in the demand-supply chain. Thus, there is an urgent need to increase the yield potential of the oilseed crops so as to enhance the storage lipid productivity. The present study describes a novel metabolic engineering ploy involving the constitutive down-regulation of endogenous ADP-glucose pyrophosphorylase (BjAGPase) enzyme and the seed-specific expression of WRINKLED1 transcription factor (AtWRI1) from Arabidopsis thaliana in Indian mustard (Brassica juncea) with an aim to divert the photosynthetically fixed carbon pool from starch to lipid synthesis in the seeds for the enhanced production of storage lipids in the seeds of transgenic mustard plants. The starch content, in both the vegetative leaf and developing seed tissues of the transgenic B. juncea lines exhibited a reduction by about 45–53% compared to the untransformed control, whereas the soluble sugar content was increased by 2.4 and 1.3-fold in the leaf and developing seed tissues, respectively. Consequently, the transgenic lines showed a significant enhancement in total seed lipid content ranging between 7.5 and 16.9%. The results indicate that the adopted metabolic engineering strategy was successful in significantly increasing the seed oil content. Therefore, findings of our research suggest that the metabolic engineering strategy adopted in this study for shifting the anabolic carbon flux from starch synthesis to lipid biosynthesis can be employed for increasing the storage lipid content of seeds in other plant species.

The Mediator Complex MED15 Subunit Mediates Activation of Downstream Lipid-Related Genes by the WRINKLED1 Transcription Factor.

The Mediator complex is known to be a master coordinator of transcription by RNA polymerase II, and this complex is recruited by transcription factors (TFs) to target promoters for gene activation or repression. The plant-specific TF WRINKLED1 (WRI1) activates glycolysis-related and fatty acid biosynthetic genes during embryogenesis. However, no Mediator subunit has yet been identified that mediates WRI1 transcriptional activity. Promoter-β-glucuronidase fusion experiments showed that MEDIATOR15 (MED15) is expressed in the same cells in the embryo as WRI1. We found that the Arabidopsis (Arabidopsis thaliana) MED15 subunit of the Mediator complex interacts directly with WRI1 in the nucleus. Overexpression of MED15 or WRI1 increased transcript levels of WRI1 target genes involved in glycolysis and fatty acid biosynthesis; these genes were down-regulated in wild-type or WRI1-overexpressing plants by silencing of MED15 However, overexpression of MED15 in the wri1 mutant also increased transcript levels of WRI1 target genes, suggesting that MED15 also may act with other TFs to activate downstream lipid-related genes. Chromatin immunoprecipitation assays confirmed the association of MED15 with six WRI1 target gene promoters. Additionally, silencing of MED15 resulted in reduced fatty acid content in seedlings and mature seeds, whereas MED15 overexpression increased fatty acid content in both developmental stages. Similar results were found in wri1 mutant and WRI1 overexpression lines. Together, our results indicate that the WRI1/MED15 complex transcriptionally regulates glycolysis-related and fatty acid biosynthetic genes during embryogenesis.

WRINKLED1 Rescues Feedback Inhibition of Fatty Acid Synthesis in Hydroxylase-Expressing Seeds.

Previous attempts at engineering Arabidopsis (Arabidopsis thaliana) to produce seed oils containing hydroxy fatty acids (HFA) have resulted in low yields of HFA compared with the native castor (Ricinus communis) plant and caused undesirable effects, including reduced total oil content. Recent studies have led to an understanding of problems involved in the accumulation of HFA in oils of transgenic plants, which include metabolic bottlenecks and a decrease in the rate of fatty acid synthesis. Focusing on engineering the triacylglycerol assembly mechanisms led to modest increases in the HFA content of seed oil, but much room for improvement still remains. We hypothesized that engineering fatty acid synthesis in the plastids to increase flux would facilitate enhanced total incorporation of fatty acids, including HFA, into seed oil. The transcription factor WRINKLED1 (WRI1) positively regulates the expression of genes involved in fatty acid synthesis and controls seed oil levels. We overexpressed Arabidopsis WRI1 in seeds of a transgenic line expressing the castor fatty acid hydroxylase. The proportion of HFA in the oil, the total HFA per seed, and the total oil content of seeds increased to an average of 20.9%, 1.26 µg, and 32.2%, respectively, across five independent lines, compared with 17.6%, 0.83 µg, and 27.9%, respectively, for isogenic segregants. WRI1 and WRI1-regulated genes involved in fatty acid synthesis were up-regulated, providing for a corresponding increase in the rate of fatty acid synthesis.

Potato tuber expression of Arabidopsis WRINKLED1 increase triacylglycerol and membrane lipids while affecting central carbohydrate metabolism.

SummaryTuber and root crops virtually exclusively accumulate storage products in the form of carbohydrates. An exception is yellow nutsedge (Cyperus esculentus) in which tubers have the capacity to store starch and triacylglycerols (TAG) in roughly equal amounts. This suggests that a tuber crop can efficiently handle accumulation of energy dense oil. From a nutritional as well as economic aspect, it would be of interest to utilize the high yield capacity of tuber or root crops for oil accumulation similar to yellow nutsedge. The transcription factor WRINKLED1 from Arabidopsis thaliana, which in seed embryos induce fatty acid synthesis, has been shown to be a major factor for oil accumulation. WRINKLED1 was expressed in potato (Solanum tuberosum) tubers to explore whether this factor could impact tuber metabolism. This study shows that a WRINKLED1 transcription factor could induce triacylglycerol accumulation in tubers of transformed potato plants grown in field (up to 12 nmol TAG/mg dry weight, 1% of dry weight) together with a large increase in polar membrane lipids. The changes in metabolism further affected starch accumulation and composition concomitant with massive increases in sugar content.

Transcriptional transitions in Nicotiana benthamiana leaves upon induction of oil synthesis by WRINKLED1 homologs from diverse species and tissues.

BackgroundCarbon accumulation and remobilization are essential mechanisms in plants to ensure energy transfer between plant tissues with different functions or metabolic needs and to support new generations. Knowledge about the regulation of carbon allocation into oil (triacylglycerol) in plant storage tissue can be of great economic and environmental importance for developing new high-yielding oil crops. Here, the effect on global gene expression as well as on physiological changes in leaves transiently expressing five homologs of the transcription factor WRINKLED1 (WRI1) originating from diverse species and tissues; Arabidopsis thaliana and potato (Solanum tuberosum) seed embryo, poplar (Populus trichocarpa) stem cambium, oat (Avena sativa) grain endosperm, and nutsedge (Cyperus esculentus) tuber parenchyma, were studied by agroinfiltration in Nicotiana benthamiana.ResultsAll WRI1 homologs induced oil accumulation when expressed in leaf tissue. Transcriptome sequencing revealed that all homologs induced the same general patterns with a drastic shift in gene expression profiles of leaves from that of a typical source tissue to a source-limited sink-like tissue: Transcripts encoding enzymes for plastid uptake and metabolism of phosphoenolpyruvate, fatty acid and oil biosynthesis were up-regulated, as were also transcripts encoding starch degradation. Transcripts encoding enzymes in photosynthesis and starch synthesis were instead down-regulated. Moreover, transcripts representing fatty acid degradation were up-regulated indicating that fatty acids might be degraded to feed the increased need to channel carbons into fatty acid synthesis creating a futile cycle. RT-qPCR analysis of leaves expressing Arabidopsis WRI1 showed the temporal trends of transcripts selected as ‘markers’ for key metabolic pathways one to five days after agroinfiltration. Chlorophyll fluorescence measurements of leaves expressing Arabidopsis WRI1 showed a significant decrease in photosynthesis, even though effect on starch content could not be observed.ConclusionsThis data gives for the first time a general view on the transcriptional transitions in leaf tissue upon induction of oil synthesis by WRI1. This yields important information about what effects WRI1 may exert on global gene expression during seed and embryo development. The results suggest why high oil content in leaf tissue cannot be achieved by solely transcriptional activation by WRI1, which can be essential knowledge in the development of new high-yielding oil crops.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-015-0579-1) contains supplementary material, which is available to authorized users.