Glucosinolate Production Research Articles

Effects of long light exposure and drought stress on plant growth and glucosinolate production in pak choi (Brassica rapa subsp. chinensis)

Glucosinolates (GLs), found in Brassicaceae family, are precursor metabolites with anti-cancer properties. Increased GLs have been studied under various environmental growth conditions. Pak choi (Brassica rapa subsp. chinensis) is a GL-rich vegetable. We hypothesize that long exposure to light and drought will increase the biomass of, and GL production in, pak choi. The experiment was conducted for 6 weeks. Long light exposure (20 h/day) increased, whilst drought exposure (12 h/week) decreased the plant growth. The plants exposed to a combination of drought and long light conditions showed similar growth pattern as control plants. GL production increased at week 6 in plants exposed to long light, while drought exposure had no impact on GL production, with the exception of glucoraphanin. Significant positive correlations were observed between plant growth and GL yield with accumulated light exposure time. Our findings suggest that long exposure to light can be used to increase both the biomass and GL production in pak choi.

Cytochrome P450s CYP380C6 and CYP380C9 in green peach aphid facilitate its adaptation to indole glucosinolate-mediated plant defense.

Overexpressing CIRCADIAN CLOCK ASSOCIATED1 in Arabidopsis thaliana (CCA1-ox) increases indole glucosinolate production and resistance to green peach aphid (Myzus persicae). Little is known of how aphids respond to this group of plant defense compounds or of the underlying molecular mechanism. Aphids reared on CCA1-ox for over 40 generations (namely the CCA population) became less susceptible to CCA1-ox than aphids maintained on the wild-type Col-0 (namely the COL population). This elevated tolerance was transgenerational as it remained for at least eight generations after the CCA population was transferred to Col-0. Intriguingly, transcriptome analysis indicated that all differential cytochrome P450 monooxygenase genes (MpCYPs), primarily MpCYP4s, MpCYP380s and MpCYP6s, were more highly expressed in the CCA population. Application of a P450 inhibitor to the CCA population resulted in decreased aphid reproduction on CCA1-ox, which was not observed if aphids were reared on Col-0. When indole glucosinolate biosynthesis in CCA1-ox was blocked using virus-induced gene silencing, the effect of the P450 inhibitor on the CCA population was attenuated, affirming the essential role played by MpCYPs in counteracting the defense mechanism in CCA1-ox that is low or absent in Col-0. Furthermore, we used host-induced gene silencing to identify MpCYP380C6 and MpCYP380C9 that specifically facilitated the CCA population to cope with CCA1-mediated plant defense. Expression profiles revealed their possible contribution to the transgenerational tolerance observed in aphids. MpCYP380C6 and MpCYP380C9 in aphids play a crucial role in mitigating indole glucosinolate-mediated plant defense, and this effect is transgenerational.

A Comprehensive Gene Inventory for Glucosinolate Biosynthetic Pathway in Arabidopsis thaliana.

Glucosinolates (GSLs) are plant secondary metabolites comprising sulfur and nitrogen mainly found in plants from the order of Brassicales, such as broccoli, cabbage, and Arabidopsis thaliana. The activated forms of GSL play important roles in fighting against pathogens and have health benefits to humans. The increasing amount of data on A. thaliana generated from various omics technologies can be investigated more deeply in search of new genes or compounds involved in GSL biosynthesis and metabolism. This review describes a comprehensive inventory of A. thaliana GSLs identified from published literature and databases such as KNApSAcK, KEGG, and AraCyc. A total of 113 GSL genes encoding for 23 transcription components, 85 enzymes, and five protein transporters were experimentally characterized in the past two decades. Continuous efforts are still on going to identify all molecules related to the production of GSLs. A manually curated database known as SuCCombase (http://plant-scc.org) was developed to serve as a comprehensive GSL inventory. Realizing lack of information on the regulation of GSL biosynthesis and degradation mechanisms, this review also includes relevant information and their connections with crosstalk among various factors, such as light, sulfur metabolism, and nitrogen metabolism, not only in A. thaliana but also in other crucifers.

<strong>From the frying pan: an unusual dwarf shrub from Namibia turns out to be a new brassicalean family</strong>

Tiganophyton karasense, an evergreen dwarf shrub, is described as a new species. A new genus and family are also proposed for it in the order Brassicales. Phylogenetic analysis of DNA sequence data indicate that Tiganophyton is sister to Bataceae/Salvadoraceae, and all three sister to Koeberliniaceae. First realized to be undescribed in 2010, T. karasense is a rare species known only from three localities in the arid Karas Region, southern Namibia. These small shrubs grow near the edges of seasonal pans on calcareous substrate underlaid by shales and mudstones of the Prince Albert Formation of the Karoo Supergroup. Morphological characters diagnostic of the new family include: a marked differentiation into long and short shoots; dimorphic, spirally arranged leaves; glucosinolate production; bisexual laterally flattened flowers borne singly in bract axils on short shoots only; tetramerous calyx, corolla and androecium with fused sepals and free, non-clawed petals; a staminal disc, but no nectary glands; deeply bilobed ovary with a gynobasic style; S-shaped gynophore supporting a bilocular, horizontally orientated or inverted ovary; two ovules per locule; and a dry, persistent fruit, provisionally interpreted as a one-seeded nutlet. Based on IUCN Red List categories and criteria, a conservation assessment of Vulnerable (VU D1) is recommended for Tiganophyton karasense.

Variation in Below-to Aboveground Systemic Induction of Glucosinolates Mediates Plant Fitness Consequences under Herbivore Attack.

Plants defend themselves against herbivore attack by constitutively producing toxic secondary metabolites, as well as by inducing them in response to herbivore feeding. Induction of secondary metabolites can cross plant tissue boundaries, such as from root to shoot. However, whether the potential for plants to systemically induce secondary metabolites from roots to shoots shows genetic variability, and thus, potentially, is under selection conferring fitness benefits to the plants is an open question. To address this question, we induced 26 maternal plant families of the wild species Cardamine hirsuta belowground (BG) using the wound-mimicking phytohormone jasmonic acid (JA). We measured resistance against a generalist (Spodoptera littoralis) and a specialist (Pieris brassicae) herbivore species, as well as the production of glucosinolates (GSLs) in plants. We showed that BG induction increased AG resistance against the generalist but not against the specialist, and found substantial plant family-level variation for resistance and GSL induction. We further found that the systemic induction of several GSLs tempered the negative effects of herbivory on total seed set production. Using a widespread natural system, we thus confirm that BG to AG induction has a strong genetic component, and can be under positive selection by increasing plant fitness. We suggest that natural variation in systemic induction is in part dictated by allocation trade-offs between constitutive and inducible GSL production, as well as natural variation in AG and BG herbivore attack in nature.

Harvest strategies to maximize the annual production of bioactive compounds, glucosinolates, and total antioxidant activities of kale in plant factories

Since kale (Brassica oleracea L. var. acephala) is one of the healthiest vegetables, its cultivation is increasing for either fresh consumption or as a source for functional foods and nutraceuticals. Plant factories are able to control the environment and trigger the accumulation of bioactive compounds with a stable supply by systematic cultivation methods. The objectives of this study were to evaluate the changes in the total phenolic compounds (TPCs), total flavonoid compounds (TFCs), glucosinolates (GLSs), and antioxidant capacity of kale in a plant factory and to determine an optimal harvest time for the maximum annual production. Two cultivars, namely ‘Manchoo collard’ and ‘Jangsoo collard’, were cultivated in a plant factory and thinned to avoid mutual shading. Both cultivars were harvested every week from 14 to 49 days after transplanting (DAT). The fresh weight (FW), dry weight (DW), projected leaf area (PLA), TPCs, TFCs, GLSs, and antioxidant capacity of both plants were measured every week. The annual production was calculated as follows: DW × the concentration × planting density × cultivation cycles per year. The optimal harvest time was determined based on the continuous phase of the production by modeling. The FW and DW of both cultivars exponentially increased, but the PLA hardly increased at 35 DAT. The TPCs, TFCs, and antioxidant capacity fluctuated or slightly changed, but the amount of substance per plant gradually increased. Their annual production increased with increasing harvest time, and only the production of TPCs in ‘Manchoo collard’ showed a local maximum when harvested at 35–42 DAT. Glucoiberin, sinigrin, and glucobrassicin were the major components of GLSs in both cultivars, and their contents fluctuated. The concentration of total GLSs was the highest at 42 DAT. Additionally, the annual production of the total and major GLSs showed the same results as the TPCs, TFCs, and antioxidant capacity. From the results, the optimum harvest time for production was determined to be 42 DAT.

Selenium Application During Radish (Raphanus sativus) Plant Development Alters Glucosinolate Metabolic Gene Expression and Results in the Production of 4-(methylseleno)but-3-enyl glucosinolate.

Selenium (Se) is an essential micronutrient for human health, entering the diet mainly through the consumption of plant material. Members of the Brassicaceae are Se-accumulators that can accumulate up to 1g Se kg−1 dry weight (DW) from the environment without apparent ill effect. The Brassicaceae also produce glucosinolates (GSLs), sulfur (S)-rich compounds that benefit human health. Radish (Raphanus sativus) has a unique GSL profile and is a Se-accumulating species that is part of the human diet as sprouts, greens and roots. In this report we describe the effects of Se-fertilisation on GSL production in radish during five stages of early development (from seed to mature salad greens) and on the transcript abundance of eight genes encoding enzymes involved in GSL metabolism. We tentatively identified (by tandem mass spectrometry) the selenium-containing glucosinolate, 4-(methylseleno)but-3-enyl glucosinolate, with the double bond geometry not resolved. Two related isothiocyanates were tentatively identified by Gas Chromatography-Mass Spectrometry as (E/Z?) isomers of 4-(methylseleno)but-3-enyl isothiocyanate. Se fertilisation of mature radish led to the presence of selenoglucosinolates in the seed. While GSL concentration generally reduced during radish development, GSL content was generally not affected by Se fertilisation, aside from the indole GSL, indol-3-ylmethyl glucosinolate, which increased on Se treatment, and the Se-GSLs, which significantly increased during development. The transcript abundance of genes involved in aliphatic GSL biosynthesis declined with Se treatment while that of genes involved in indole GSL biosynthesis tended to increase. APS kinase transcript abundance increased significantly in three of the four developmental stages following Se treatment. The remaining genes investigated were not significantly changed following Se treatment. We hypothesise that increased APS kinase expression in response to Se treatment is part of a general protection mechanism controlling the uptake of S and the production of S-containing compounds such as GSLs. The upregulation of genes encoding enzymes involved in indole GSL biosynthesis and a decrease in those involved in aliphatic GSL biosynthesis may be part of a similar mechanism protecting the plant’s GSL complement whilst limiting the amount of Se-GSLs produced.

S-glycosyltransferase UGT74B1 can glycosylate both S- and O-acceptors: mechanistic insights through substrate specificity

UGT74B1 from Arabidopsis thaliana is one of the few characterized glycosyltransferases able to generate a thioglycosidic linkage in vivo, using the sulfur atom of thiohydroximate as the nucleophile in the glycosylation reaction. This critical biosynthetic step in glucosinolate production has been documented. Yet little is known about the molecular mechanism that enables this rare and unusual glycosylation at the sulfur atom. To identify the role of this atom in the glycosylation reaction and unravel the mechanism of UGT74B1, we used a range of substrates containing either sulfur or oxygen. We first demonstrated that the enzyme could catalyze the glycosylation of thiohydroximates but also of O-containing hydroximates analogs. If KM values were shown to be close between analogs, the reaction catalytic rate kcat was 50–100 lower in the case of hydroximates. The glycosylation reaction is catalyzed through deprotonation of the acceptor, which was confirmed by the removal of catalytic carboxylic residues by site-directed mutagenesis. Moreover, using a range of simple phenols and thiophenols as UGT74B1 substrate acceptors for glycosylation, we concluded that the glycosylation reaction rate is correlated to the acceptor atom acidity, and not to the nature of this nucleophilic atom (oxygen or sulfur).

CIRCADIAN CLOCK-ASSOCIATED1 Controls Resistance to Aphids by Altering Indole Glucosinolate Production.

CIRCADIAN CLOCK-ASSOCIATED1 (CCA1), a well-known central circadian clock regulator, coordinates plant responses to environmental challenges. Its daily rhythmic expression in Arabidopsis (Arabidopsis thaliana) confers host resistance to the caterpillar Trichoplusia ni However, it is unclear whether CCA1 plays a role in defense against phloem sap-feeding aphids. In this study, we showed that green peach aphid (Myzus persicae) displayed an intrinsic circadian feeding rhythm. Under constant light, wild-type Columbia-0 (Col-0) Arabidopsis plants coentrained with aphids in the same light/dark cycles exhibited greater antixenotic activity than plants preentrained in the opposite cycle from the aphids. Consistently, circadian mutants cca1-1, cca1-11, lhy-21, ztl-1, ztl-4, and lux-2 suffered more severe damage than Col-0 plants when infested by aphids, suggesting that the Arabidopsis circadian clock plays a defensive role. However, the arrhythmic CCA1 overexpression line (CCA1-OX) displayed strong antixenotic and antibiotic activities despite its loss of circadian regulation. Aphids feeding on CCA1-OX plants exhibited lower reproduction and smaller body size and weight than those on Col-0. Apparently, CCA1 regulates both clock-dependent and -independent defense responses. Systematic investigation based on bioinformatics analyses indicated that resistance to aphids in CCA1-OX plants was due primarily to heightened basal indole glucosinolate levels. Interestingly, aphid feeding induced alternatively spliced intron-retaining CCA1a/b transcripts, which are normally expressed at low levels, whereas expression of the major fully spliced CCA1 transcript remained largely unchanged. We hypothesize that posttranscriptional modulation of CCA1 expression upon aphid infestation maximizes the potential of circadian-mediated defense and stress tolerance while ensuring normal plant development.

Production and application of 3-phosphoadenosine-5- phosphosulfate

Sulfated compounds are widely present in cytoplasm, on cell surface, and in extracellular matrix. These compounds play important roles in cell development, differentiation, immune response, detoxication, and cell signal transduction. 3-Phosphoadenosine-5-phosphosulfate (PAPS) is the universal sulfate group donor for the biosynthesis of sulfated compounds. Up to now, the synthesis of PAPS is still too expensive for industrial applications. This review focuses on the recent progress of PAPS production and summaries the application of PAPS, particularly in the production of glucosinolate, heparin, condroitin sulfate, and oxamniquine production.

Bioaccumulation of selected macro- and microelements and their impact on antioxidant properties and accumulation of glucosinolates and phenolic acids in in vitro cultures of Nasturtium officinale (watercress) microshoots

The study evaluated bioaccumulation capacity of macro- and microelements, their impact on the production of glucosinolates and phenolic acids and antioxidant properties in a microshoot culture model of Nasturtium officinale.Elements: calcium, chromium, copper, iron, lithium, magnesium, selenium and zinc were supplemented in different salt concentrations to culture media. Bioaccumulation of elements [mg/100 gDW] varied from 1.24 (Li,1 mg/l) to 498.62 (Cr,50 mg/l) and was dependent on the type of element and its concentration. The bioconcentration factor (BCF) ranged from 11.37 (Li,25 mg/l) to 4467.00 (Ca,1 mg/l). The total glucosinolate contents [mg/100gDW] varied from 108.11 (Cr,1 mg/l) to 172.90 (Ca,1 mg/l). The presence of four phenolic acids was confirmed in the microshoots. Their total contents [mg/100gDW] ranged from 19.35 (Mg,10 mg/l) to 139.21 (Fe,50 mg/l). The highest antioxidant activity [nM trolox/mgDW], as evaluated by CUPRAC and QUENCHER-CUPRAC methods, was equal to 55.50 (Cu,1 mg/l) and 161.10 (Li,5 mg/l), respectively. The results proved good correlations between all studied parameters.

Methyl jasmonate treated broccoli: Impact on the production of glucosinolates and consumer preferences

Applying methyl jasmonate can mimic the defense response to insect damage in broccoli and enhances the production of glucosinolates, especially inducible indolyl GS-neoglucobrassicin. Previous studies have suggested that glucosinolates and their hydrolysis products are anti-carcinogenic. Therefore, MeJA treatment may increase the nutritional quality of broccoli. However, there are few reports on the sensory evaluation and consumer acceptance of MeJA-treated broccoli. In this study, an untrained consumer panel could not detect any taste differences between steamed MeJA-treated and untreated broccoli, even though the steamed MeJA-treated broccoli contained 50% more glucosinolates than untreated broccoli. The partial least square-regression model suggested that neoglucobrassicin-derived hydrolysis compounds were the major metabolites that determined overall preference for raw MeJA-treated broccoli potentially due to their potential negative sensory qualities. The results imply that MeJA treatment can increase the nutritional quality of broccoli without sacrificing taste in precooked meals or frozen vegetables.

Overexpression of the MYB29 transcription factor affects aliphatic glucosinolate synthesis in Brassica oleracea

Key messageOverexpression of BoMYB29 gene up-regulates the aliphatic glucosinolate pathway in Brassica oleracea plants increasing the production of the anti-cancer metabolite glucoraphanin, and the toxic and pungent sinigrin.Isothiocyanates, the bio-active hydrolysis products of glucosinolates, naturally produced by several Brassicaceae species, play an important role in human health and agriculture. This study aims at correlating the content of aliphatic glucosinolates to the expression of genes involved in their synthesis in Brassica oleracea, and perform functional analysis of BoMYB29 gene. To this purpose, three genotypes were used: a sprouting broccoli, a cabbage, and a wild genotype (Winspit), a high glucosinolate containing accession. Winspit showed the highest transcript level of BoMYB28, BoMYB29 and BoAOP2 genes, and BoAOP2 expression was positively correlated with that of the two MYB genes. Further analyses of the aliphatic glucosinolates also showed a positive correlation between the expression of BoAOP2 and the production of sinigrin and gluconapin in Winspit. The Winspit BoMYB29 CDS was cloned and overexpressed in Winspit and in the DH AG1012 line. Overexpressing Winspit plants produced higher quantities of alkenyl glucosinolates, such as sinigrin. Conversely, the DH AG1012 transformants showed a higher production of methylsulphinylalkyl glucosinolates, including glucoraphanin, and, despite an up-regulation of the aliphatic glucosinolate genes, no increase in alkenyl glucosinolates. The latter may be explained by the absence of a functional AOP2 gene in DH AG1012. Nevertheless, an extract of DH AG1012 lines overexpressing BoMYB29 provided a chemoprotective effect on human colon cells. This work exemplifies how the genetic diversity of B. oleracea may be used by breeders to select for higher expression of transcription factors for glucosinolate biosynthesis to improve its natural, health-promoting properties.

Identification and Characterization of Genes Involved in Field Pennycress (Thlaspi arvense L.) Glucosinolate Production (nzz047.OR20-06-19)

ObjectivesField pennycress (Thlaspi arvense L.) is a winter annual oilseed undergoing development as a cash cover crop for the Upper Midwest. While it provides ecosystem benefits when planted between conventional crop rotations, there are few end-uses for wild pennycress. Pennycress is high in oil and protein, and current efforts aim to improve seed composition to generate environmentally sustainable food sources. Similar to Arabidopsis thaliana and oilseed rape, pennycress is rich in glucosinolates, which limits seed palatability and utility. This study aimed to identify mutations responsible for low glucosinolate pennycress lines in a mutant population. Methods39 low glucosinolate pennycress lines from a M3 mutant population were identified using brassica-calibrated NIRS and validated using column-exchange chromatography. Lines underwent whole genome sequencing to identify mutations in known brassica orthologs involved in glucosinolate production. We assessed the impact of three mutations on gene expression using RT-qPCR. ResultsWe identified mutations in three orthologs involved in glucosinolate production. Putative candidates were selected for co-expression validation based on amino acid changes in highly conserved regions of peptide sequences. Genes of interest included the transcription factor bHLH005, the amino acid chain elongation gene IMD1, and the core structure biosynthesis gene CYP83A1. The bHLH005 mutant showed a 48% reduction in sinigrin. Expression decreased in several glucosinolate-related genes including MAM1, AOP2-like, and CYP83A1 by 0.98, 0.82, and 0.78-fold respectively. The IMD1 mutant showed a 45% reduction in sinigrin. Expression decreased in IMD1, MAM1, AOP2-like, and CYP83A1 by 0.77, 0.81, 0.72, and 0.73-fold respectively. The CYP83A1 mutant showed a 19% reduction in sinigrin. Expression decreased in CYP83A1, MAM1, and AOP2-like by 0.14, 0.16, and 0.22-fold respectively. ConclusionsUnderstanding pennycress glucosinolate production can allow for the development of low-glucosinolate pennycress lines. We identified and validated three genes involved in pennycress glucosinolate production that can be used to develop low glucosinolate pennycress lines. Ultimately, this can improve the economic viability of the pennycress system by providing new end use opportunities. Funding SourcesPepsiCo R&D.

Comparative analysis of glucosinolate production in hairy roots of green and red kale (Brassica oleracea var. acephala)

Glucosinolates (GSLs) are sulfur- and nitrogen-containing secondary metabolites that function in plant defense and provide benefits to human health. In this study, using Agrobacterium rhizogenes R1000, green and red kale hairy roots were established. The expression levels of GSLs biosynthesis genes and their accumulation in both kale hairy roots were analyzed by quantitative real-time PCR and HPLC. The results showed that the expression of most indolic GSLs biosynthesis genes was higher in the hairy roots of green kale than in that of red kale. In contrast, the expression of BoCYP83A1 and BoSUR1 encoding key enzymes aromatic GSL biosynthesis was significantly higher in red kale hairy root. The HPLC analysis identified six GSLs. The levels of 4-methoxyglucobrassicin, glucobrassicin, and 4-hydroxyglucobrassicin were 6.21, 5.98, and 2 times higher, respectively, in green kale than in red kale, whereas the levels of neoglucobrassicin and gluconasturtiin were 16.2 and 3.48 times higher, respectively, in red kale than in green kale. Our study provides insights into the underlying mechanisms of GSLs biosynthesis in kale hairy roots and can be potentially used as “biological factories” for producing bioactive substances such as GSLs.

Influence of three different soil types on the interaction of two strains of Trichoderma harzianum with Brassica rapa subsp. sylvestris cv. esculenta, under soil mineral fertilization

The influence of Trichoderma fungi on growth, yield and nutrient status of food crops has been widely studied. However, a lack of knowledge still exists on the effect of agricultural soils with different geological nature, land use and management on Trichoderma-plant interaction. This study aims to evaluate the influence of three soil types, with different physicochemical properties and under fertilization by NPK-based nutrient solutions, on the efficacy of two T. harzianum (T22 and M10) strains on the growth, nutrient uptake and glucosinolate concentration in Brassica rapa subsp. sylvestris cv. esculenta. Overall data showed that the biofertilization by Trichoderma strains is very effective to enhance the fertility and productivity of the nutrient-poor soils even in the absence of fertilization by NPK-nutrient solutions, as well as to stimulate the production of glucosinolates by foliar biomass of B. rapa, although this effect was strongly dependent on the soil type.

Dissecting R gene and host genetic background effect on the Brassica napus defense response to Leptosphaeria maculans

While our understanding of the genetics underlying the Brassica-Leptosphaeria pathosystem has advanced greatly in the last decade, differences in molecular responses due to interaction between resistance genes and host genetic background has not been studied. We applied RNAseq technology to monitor the transcriptome profiles of Brassica napus (Bn) lines carrying one of four blackleg R genes (Rlm2, Rlm3, LepR1 & LepR2) in Topas or Westar background, during the early stages of infection by a Leptosphaeria maculans (Lm) isolate carrying the corresponding Avr genes. We observed upregulation of host genes involved in hormone signalling, cell wall thickening, response to chitin and glucosinolate production in all R gene lines at 3 day after inoculation (dai) albeit having higher level of expression in LepR1 and Rlm2 than in Rlm3 and LepR2 lines. Bn-SOBIR1 (Suppressor Of BIR1-1), a receptor like kinase (RLK) that forms complex receptor like proteins (RLPs) was highly expressed in LepR1 and Rlm2 at 3 dai. In contrast Bn-SOBIR1 induction was low in Rlm3 line, which could indicate that Rlm3 may function independent of SOBIR1. Expression of Salicylic acid (SA) related defense was enhanced in LepR1 and Rlm2 at 3 dai. In contrast to SA, expression of Bn genes with homology to PDF1.2, a jasmonic acid (JA) pathway marker, were increased in all Rlm and LepR lines at 6 and 9 dai. Effect of host genetic background on induction of defense, was determined by comparison of LepR1 and LepR2 in Topas vs Westar genotype (i.e. T-LepR1 vs W-LepR1 and T-LepR2 vs W-LepR2). In both cases (regardless of R gene) overall number of defense related genes at the earliest time point (3 dai) was higher in Tops compared to Westar. SA and JA markers genes such as PR1 and PDF1.2 were more induced in Topas compared to Westar introgression lines at this time point. Even in the absence of any R gene, effect of Topas genotype in enhanced defense, was also evident by the induction of PDF1.2 that started at a low level at 3 dai and peaked at 6 and 9 dai, while no induction in Westar genotype was observed at any of these time points. Overall, variation in time and intensity of expression of genes related to defense, was clearly dependent on both R gene and the host genotype.

Genome-Wide Analysis of Glycoside Hydrolase Family 1 β-glucosidase Genes in Brassica rapa and Their Potential Role in Pollen Development.

Glycoside hydrolase family 1 (GH1) β-glucosidases (BGLUs) are encoded by a large number of genes, and are involved in many developmental processes and stress responses in plants. Due to their importance in plant growth and development, genome-wide analyses have been conducted in model plants (Arabidopsis and rice) and maize, but not in Brassica species, which are important vegetable crops. In this study, we systematically analyzed B. rapa BGLUs (BrBGLUs), and demonstrated the involvement of several genes in pollen development. Sixty-four BrBGLUs were identified in Brassica databases, which were anchored onto 10 chromosomes, with 10 tandem duplications. Phylogenetic analysis revealed that 64 genes were classified into 10 subgroups, and each subgroup had relatively conserved intron/exon structures. Clustering with Arabidopsis BGLUs (AtBGLUs) facilitated the identification of several important subgroups for flavonoid metabolism, the production of glucosinolates, the regulation of abscisic acid (ABA) levels, and other defense-related compounds. At least six BrBGLUs might be involved in pollen development. The expression of BrBGLU10/AtBGLU20, the analysis of co-expressed genes, and the examination of knocked down Arabidopsis plants strongly suggests that BrBGLU10/AtBGLU20 has an indispensable function in pollen development. The results that are obtained from this study may provide valuable information for the further understanding of β-glucosidase function and Brassica breeding, for nutraceuticals-rich Brassica crops.

Stress response to CO2 deprivation by Arabidopsis thaliana in plant cultures.

After being the standard plant propagation protocol for decades, cultures of Arabidopsis thaliana sealed with Parafilm remain common today out of practicality, habit, or necessity (as in co-cultures with microorganisms). Regardless of concerns over the aeration of these cultures, no investigation has explored the CO2 transport inside these cultures and its effect on the plants. Thereby, it was impossible to assess whether Parafilm-seals used today or in thousands of older papers in the literature constitute a treatment, and whether this treatment could potentially affect the study of other treatments.For the first time we report the CO2 concentrations in Parafilm-sealed cultures of A. thaliana with a 1 minute temporal resolution, and the transcriptome comparison with aerated cultures. The data show significant CO2 deprivation to the plants, a drastic suppression of photosynthesis, respiration, starch accumulation, chlorophyll biosynthesis, and an increased accumulation of reactive oxygen species. Most importantly, CO2 deprivation occurs as soon as the cotyledons emerge. Gene expression analysis indicates a significant alteration of 35% of the pathways when compared to aerated cultures, especially in stress response and secondary metabolism processes. On the other hand, the observed increase in the production of glucosinolates and flavonoids suggests intriguing possibilities for CO2 deprivation as an organic biofortification treatment in high-value crops.

Increased Glucosinolate Production in Brassica oleracea var. italica Cell Cultures Due to Coronatine Activated Genes Involved in Glucosinolate Biosynthesis.

In this work, the effect of different elicitors and culture conditions on the production of glucosinolates in broccoli cell cultures was studied. The results showed that 0.5 μM coronatine was the best elicitor for increasing glucosinolate production (205-fold increase over untreated cells after 72 h of treatment). Furthermore, the expression levels of some genes related to the biosynthetic pathway of glucosinolates as well as three Myb transcription factors also have been studied. The highest glucosinolate levels found in coronatine-treated cells were closely correlated with the highest gene expression levels of Cyp79b2, Cyp83b1, St5a, Myb51, and Myb122 after 6 h of treatment. The data shown in this study provide new insight into the key metabolic steps involved in the biosynthesis of glucosinolates, which will be of use for future applications of metabolic engineering techniques in broccoli.