Spatiotemporal transcriptomics and metabolic profiling provide insights into gene regulatory networks during lentil seed development.

Lentil (Lens culinaris Medik.) is a nutritious legume with seeds rich in protein, minerals and an array of diverse specialized metabolites. The formation of a seed requires regulation and tight coordination of developmental programs to form the embryo, endosperm and seed coat compartments, which determines the structure and composition of mature seed and thus its end-use quality. Understanding the molecular and cellular events and metabolic processes of seed development is essential for improving lentil yield and seed nutritional value. However, such information remains largely unknown, especially at the seed compartment level. In this study, we generated high-resolution spatiotemporal gene expression profiles in lentil embryo, seed coat and whole seeds from fertilization through maturation. Apart from anatomic differences between the embryo and seed coat, comparative transcriptomics and weighted gene co-expression network analysis revealed embryo- and seed coat-specific genes and gene modules predominant in specific tissues and stages, which highlights distinct genetic programming. Furthermore, we investigated the dynamic profiles of flavonoid, isoflavone, phytic acid and saponin in seed compartments across seed development. Coupled with transcriptome data, we identified sets of candidate genes involved in the biosynthesis of these metabolites. The global view of the transcriptional and metabolic changes of lentil seed tissues throughout development provides a valuable resource for dissecting the genetic control of secondary metabolism and development of molecular tools for improving seed nutritional quality.

Comprehensive compositional assessment of bioactive compounds in diverse pea accessions.

Pea (Pisum sativum L.) is an important legume crop providing a good source of protein, vitamins, minerals and bioactive compounds with health benefits for humans. In this study, an improved method for simultaneous analysis of multiple phytoestrogens among 100 pea accessions was developed. Ipriflavone, (a synthetic isoflavone), was used as an internal standard for the semiquantitative analysis of 17 phytoestrogens including isoflavone aglycones and conjugates, allowing direct analysis of isoflavones in their naturally occurring forms. This comprehensive dataset demonstrated that the isoflavones varied greatly and some accessions tended to have high levels of multiple phytoestrogens among the 100 accessions analyzed. Isoliquiritigenin followed by glycitein were the predominant compounds detected in the accessions and showed the highest correlation with the total phytoestrogens content. Secoisolariciresinol content was consistently higher in yellow cotyledon peas than in green cotyledon peas, whereas the contents of coumestrol, genestein and secoisolariciresinol were significantly correlated with seed coat color. The total phenolics and saponins showed a wide range of variability among the accessions with higher concentrations of total phenolics observed in seeds with pigmented seed coat or yellow cotyledon seeds, suggesting the synthesis of saponins and phenolics are significantly affected by metabolic pathway genes controlling cotyledon color or seed coat color. This study profiled the variability of bioactive compounds of pea seed quality traits in diverse pea accessions and provides an immense resource for continued research, breeding and selection of genotypes for a wide range of applications.

Fine-mapping of a putative glutathione S-transferase (GST) gene responsible for yellow seed colour in flax (Linum usitatissimum).

OBJECTIVE: The brown seed coat colour of flax (Linum ustiatissimum) results from proanthocyanidin synthesis and accumulation. Glutathione S-transferases (GSTs), such as the TT19 protein in Arabidopsis, have been implicated in the transport of anthocyanidins during the synthesis of the brown proanthocyanidins. This study fine mapped the g allele responsible for yellow seed colour in S95407 and identified it as a putative mutated GST. RESULTS: We developed a Recombinant Inbred Line population with 320 lines descended from a cross between CDC Bethune (brown seed coat) and S95407 (yellow seed) and used molecular markers to fine map the G gene on Chromosome 6 (Chr 6). We used Next Generation Sequencing (NGS) to identify a putative GST was identified in this region and Sanger sequenced the gene from CDC Bethune, S95407 and other yellow seeded genotypes. The putative GST from S95407 had 13 SNPs encoding, including four non-synonymous amino acid changes, compared to the CDC Bethune reference sequence and the other genotypes. The GST encoded by Lus10019895 is a lambda-GST in contrast to the Arabidopsis TT19 which is a phi-GST.

Publisher Correction: QTL mapping and molecular characterization of the classical D locus controlling seed and flower color in Linum usitatissimum (flax).

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Contrasting Root and Photosynthesis Traits in a Large-Acreage Canadian Durum Variety and Its Distant Parent of Algerian Origin for Assembling Drought/Heat Tolerance Attributes.

In Canada, the world's top exporter of high-protein durum, varietal development over its nearly six-decade history has been driven by a quest for yield improvement without compromise on grain protein content and other quality aspects. Pelissier, a landrace selection from Algeria that was introduced into North America more than a century ago and the variety Strongfield that was released in 2004 are notable. Pelissier, known to elaborate more roots and considered as drought tolerant, has been cultivated commercially and thus deemed adapted. Strongfield has Pelissier in its pedigree, and it remains a high-acreage variety. Strongfield was found to elaborate only about half of the root biomass of Pelissier at maturity in greenhouse trials under well-watered conditions. Extended drought stress caused a significant reduction in the root biomass of both lines. However, Pelissier under drought maintained at least as much root biomass as that of Strongfield under well-watered conditions. In comparison to Pelissier, it had a superior photosynthesis rate (27.16 µmol CO2 m-2 s-1), capacity for carboxylation (Vcmax: 132.83 µmol CO2 m-2 s-1) and electron transport/ribulose-1,5-bisphosphate (RuBP) regeneration (Jmax: 265.40 µmol CO2 m-2 s-1); the corresponding values for Pelissier were 19.62 µmol CO2 m-2 s-1, 91.87 µmol CO2 m-2 s-1, and 163.83 µmol CO2 m-2 s-1, respectively, under well-watered conditions. Under short-term/mild drought conditions, the carbon assimilation rate remained stable in Pelissier while it declined in Strongfield to the Pelissier level. However, Strongfield succumbed to extended drought sooner than Pelissier. Photosynthesis in Strongfield but not Pelissier was found to be sensitive to high temperature stress. These results provide encouraging prospects for further exploitation of beneficial physiological traits from Pelissier in constructing climate-resilient, agronomically favorable wheat ideotypes.

QTL mapping and molecular characterization of the classical D locus controlling seed and flower color in Linum usitatissimum (flax).

The flowers of flax (linseed) are blue-hued, ephemeral and self-pollinating, and the seeds are typically brown. A century-old interest in natural yellow seed variants and a historical model point to recessive alleles in B1, D and G loci being responsible, but the functional aspects had remained unknown. Here, we characterized the "D" locus by quantitative trait loci (QTL) mapping and identified a FLAVONOID 3'5' HYDROXYLASE (F3'5'H) gene therein. It does not belong to the F3'5'H clade, but resembles biochemically characterized F3'Hs (flavonoid 3' hydroxylase) but without F3'H activity. The genome lacks other F3'H or F3'H-like genes. The apparent neo-functionalization from F3'H is associated with a Thr498 → Ser498 substitution in a substrate recognition site (SRS). The yellow seed and white flower phenotypes of the classical d mutation was found to be due to one nucleotide deletion that would truncate the deduced product and remove three of the six potential SRS, negatively impacting delphinidin synthesis. Delphinidin is sporadic in angiosperms, and flax has no known pollination syndrome(s) with functional pollinator group(s) that are attracted to blue flowers, raising questions on the acquisition of F3'5'H. The appearance of d allele is suggestive of the beginning of the loss of F3'5'H in this species.

Induction of antioxidant enzymes by curcumin and its analogues in human islets: implications in transplantation.

OBJECTIVES: The survival of transplanted human islets is hampered by the quality of islets, which is affected by oxidative stress during isolation. The objective of this study was to determine if curcumin and its analogues could induce antioxidant enzymes in beta cells of human islets. METHODS: The expression of antioxidant enzymes in isolated human islets exposed to curcuminoids was determined at the messenger RNA levels by real-time quantitative reverse transcription-polymerase chain reaction using Taqman probes and at the protein level by Western blot analysis. Double immunofluorescent staining of islets was carried out to determine the induction of antioxidant enzymes in beta cells. RESULTS: Curcuminoids induced the expression of heme oxygenase 1; modulatory subunit of gamma-glutamyl-cysteine ligase; and NAD(P)H:quinone oxidoreductase 1 at the messenger RNA levels by 2- to 12-fold and at the protein levels by 2- to 6-fold in human islets. Increased expression of antioxidant enzymes was seen in beta cells of islets as shown by immunofluorescent staining. Curcuminoids also increased the islet content of glutathione (a product of the modulatory subunit of gamma-glutamyl-cysteine ligase) and the basal insulin secretion and protected them from oxidative stress. CONCLUSIONS: Our observations suggest that curcumin or its analogues could be used to induce cellular defense against oxidative stress and improve islet transplantation outcomes.

Regulation of heme oxygenase-1 expression by demethoxy curcuminoids through Nrf2 by a PI3-kinase/Akt-mediated pathway in mouse beta-cells.

Curcumin (diferuloylmethane), a component of turmeric, has been shown to have therapeutic properties. Induction of phase 2 detoxifying enzymes is a potential mechanism through which some of the actions of curcumin could proceed. Heme oxygenase-1 (HO-1), an antioxidant phase 2 enzyme, has been reported to have cytoprotective effects in pancreatic beta-cells. Curcumin on further purification yields demethoxy curcumin (DMC) and bisdemethoxy curcumin (BDMC). The objective of the present study was to determine the mechanism by which these purified curcuminoids induce HO-1 in MIN6 cells, a mouse beta-cell line. Demethoxy curcuminoids induced HO-1 promoter linked to the luciferase reporter gene more effectively than curcumin. The induction was dependent on the presence of antioxidant response element (ARE) sites containing enhancer regions (E1 and E2) in HO-1 promoter and nuclear translocation of nuclear factor-E2-related factor (Nrf2), the transcription factor that binds to ARE. Curcuminoids stimulated multiple signaling pathways that are known to induce HO-1. Inhibition of specific signaling pathways with pharmacological inhibitors and cotransfection experiments suggested the involvement of phosphotidylinositol 3-kinase and Akt. Real-time quantitative RT-PCR analysis showed significant elevation in the mRNA levels of HO-1 and two other phase 2 enzymes, the regulatory subunit of glutamyl cysteine ligase, which is needed for the synthesis of glutathione, and NAD(P)H:quinone oxidoreductase, which detoxifies quinones. DMC and BDMC induced the expression of HO-1 and translocated Nrf2 to nucleus in beta-cells of mouse islets. Our observations suggest that demethoxy curcuminoids could be used to induce a cellular defense mechanism in beta-cells under conditions of stress as seen in diabetes.