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Over the decades, extensive research efforts have been undertaken to understand how secondary plant metabolites are affected by genetic, environmental, and agronomic factors. Understanding the genetic basis of stress-response metabolite biosynthesis is crucial for sustainable agriculture production amidst frequent occurrence of climatic anomalies. Although it is known that environmental factors influence phytochemical profiles and their content, studies of plant compounds in relation to stress mitigation are only emerging and largely hindered by phytochemical diversities and technical shortcomings in measurement techniques. Despite these challenges, considerable success has been achieved in profiling of secondary metabolites such as glucosinolates, flavonoids, carotenoids, phenolic acids and alkaloids. In this study, we aimed to understand the roles of glucosinolates, flavonoids, carotenoids, phenolic acids and alkaloids in relation to their abiotic stress response, with a focus on the developing of stress-resilient crops. The focal genus is the Brassica since it (i) possesses variety of specialized phytochemicals that are important for its plant defense against major abiotic stresses, and (ii) hosts many economically important crops that are sensitive to adverse growth conditions. We summarize that augmented levels of specialized metabolites in Brassica primarily function as stress mitigators against oxidative stress, which is a secondary stressor in many abiotic stresses. Furthermore, it is clear that functional characterization of stress-response metabolites or their genetic pathways describing biosynthesis is essential for developing stress-resilient Brassica crops.

This study aimed to determine whether leaf extracts from seven Eruca vesicaria subsp. sativa cultivars and their biochemically active compounds (glucosinolates and downstream-derived products) inhibit mycelia growth of three well-known pathogenic oomycetes, Phytopythium chamaehyphon, Phytopythium vexans and Phytophthora citrophthora; being the most significant in the development of Kiwifruit Vine Decline Syndrome (KVDS). Leaf extract quantity of 10, 20 and 30 mg were inoculated in Petri dish (90 mm Ø, each 22 mL of liquid medium – Potato Dextrose Agar), for in vitro bioassays. A pathogen plug was placed in the centre of each plate and the Oomycota colony perimeter was marked 5 days after inoculation. Radial colony growth was measured from 4 marks per plate 5, 10, and 15 days after inoculation, further elaborated with Image J software image analysis. Growth rates for all strains were inhibited by around 67% after 15 days. This was most pronounced when applying the highest concentration of leaf extract. By using Liquid Chromatography Mass Spectrometry (LC-MS) and Gas Chromatography Mass Spectrometry (GC-MS), fifteen glucosinolate compounds, of which glucosativin was found in the highest quantity, were identified. Concentrations of hydrolysis products produced by leaves (erucin and sativin) were also investigated, and were significantly associated with colony radial growth, especially towards Pp. chamaehyphon and Pp. vexans. Three downstream products of glucosinolates (two pure isothiocyanates, AITC and PEITC; and one indole I3C; all commonly present in Brassicaceae) were also tested, and a statistically significant inhibition of growth was observed at the highest concentration (0.6 µL).

This study aimed to determine whether leaf extracts from seven Eruca vesicaria subsp. sativa cultivars and their biochemically active compounds (glucosinolates and downstream-derived products) inhibit mycelia growth of three well-known pathogenic oomycetes, Phytopythium chamaehyphon, Phytopythium vexans and Phytophthora citrophthora; being the most significant in the development of Kiwifruit Vine Decline Syndrome (KVDS). Leaf extract quantity of 10, 20 and 30 mgwereinoculated in Petridish (90mmØ, each 22mLof liquidmedium-Potato Dextrose Agar), for in vitro bioassays. A pathogen plug was placed in the centre of each plate and the Oomycota colony perimeter was marked 5 days after inoculation. Radial colony growth was measured from 4 marks per plate 5, 10, and 15 days after inoculation, further elaborated with Image J software image analysis. Growthrates forall strainswereinhibitedbyaround67%after15days. This wasmostpronouncedwhenapplying thehighest concentrationof leaf extract. By using Liquid Chromatography Mass Spectrometry (LC-MS) and Gas Chromatography Mass Spectrometry (GC-MS), fifteen glucosinolate compounds, of which glucosativin was found in the highest quantity, were identified. Concentrations of hydrolysis products produced by leaves (erucin and sativin) were also investigated, and were significantly associated with colony radial growth, especially towards Pp. chamaehyphon and Pp. vexans. Three downstream products of glucosinolates (two pure isothiocyanates, AITC and PEITC; and one indole I3C; all commonly present in Brassicaceae) were also tested, and a statistically significant inhibition of growth was observed at the highest concentration (0.6 µL). [ABSTRACT FROM AUTHOR]

Rocket (Eruca vesicaria subsp. sativa) is a source of sulfur-containing glucosinolates (GSLs). GSLs and their breakdown hydrolysis products (GHPs) are responsible for health-related benefits, such as anti-cancer and anti-neurodegenerative properties. Understanding how phytochemical composition changes between cultivation environments is key to developing cultivars with improved nutritional quality. Two consecutive harvests (first and second regrowth) of crops, grown in both Italy and the UK, were used to determine the phytochemical and transcriptomic differences between six lines of Eruca. Samples were taken upon delivery from field sites (D0) and after five days of cold storage (D5) for each location. Leaves were analysed for sulfur content, volatile organic compounds (VOCs), GSLs, GHPs, and sugars. Transcriptome data were associated with metabolite profiles to identify differentially expressed genes between plants grown in the two environments. VOC compounds (carbon disulfide, methyl thiocyanate) were associated with growth environment and with differences in sulfur metabolism gene expression (APR2, LSU2, LSU3, SDI1, SiR), GSL biosynthesis (MYB28, FMOGS-OX2) and GHP formation (ESM1, TGG1, TGG2). The concentrations of sugars were an order of magnitude greater in UK grown samples (up to 29.9 mg g-1 dry weight; dw). Sulfur content was significantly higher in the Italy plant samples (11.4 – 20.1 mg g-1 dw), which was in turn associated with higher concentrations of GSLs (pentyl GSL, up to 15.8 μmol g-1 dw; sinigrin, up to 0.005 μmol g-1 dw; glucoraphanin, up to 5.1 μmol g-1 dw; glucorucolamine, up to 23.6 μmol g-1 dw; neoglucobrassicin, up to 5.3 μmol g-1 dw) and hydrolysis products (sativin, up to 13.5 μmol g-1 dw; erucin, up to 1 μmol g-1 dw; sulforaphane, up to 34.7 μmol g-1 dw). VOC profiles of plants cultivated in the UK were distinct from Italy grown plants, with higher relative abundances of alkanes and esters in second cut and shelf-life (D5) samples. The data indicate a significant interaction of cultivar response with environment, highlighting the difficulty of producing Eruca crops with consistent phytochemical and postharvest traits. Genes with differential expression between plants grown in Italy and the UK could be used as markers of phytochemical quality and composition.

Growing microgreens on trays without substrate in a vertical multilayered growing unit offers several advantages over traditional agriculture methods. This study investigated the yield performance and nutritional quality of five selections of radish microgreens grown in sprouting trays, without a substrate using only water, in an indoor multilayer cultivation system using artificial light. Various parameters were measured, including fresh weight, dry matter, chlorophyll, minerals, amino acids, phenolics, flavonoids, anthocyanins, vitamin C, glucosinolates, and antioxidant activity with four different in vitro assays. After ten days, the biomass had increased by 6-10 times, and the dry matter varied from 4.75-7.65%. The highest yield was obtained from ‘Asia red’, while the lowest was from ‘Koregon red’. However, ‘Koregon red’ and ‘Asia red’ had the highest dry matter. ‘Asia red’ was found to have the highest levels of both Chls and vitamin C compared to the other cultivars, while ‘Koregon red’ exhibited the highest levels of total phenolics and flavonoids. Although variations in the levels of individual glucosinolates were observed, there were no significant differences in the total content of glucosinolates among the five cultivars. ‘Asia purple’ had the highest anthocyanin content, while ‘Asia green 2’ had the lowest. The K, Mg, and Na concentrations were significantly highest in ‘Asia green 2’, and the highest Ca was recorded in ‘Asia purple’. Overall, ‘Asia purple’ and ‘Koregon red’ were the best cultivars in terms of nutritional quality among the tested radish microgreens. These cultivars exhibited high levels of dry weight, total phenolics, flavonoids, anthocyanins, essential and total amino acids, and antioxidant activities. Moreover, the implementation of this vertical cultivation method for microgreens, which relies solely on water and seeds known for their tall shoots during the sprouting could hold promise as a sustainable approach. This method can effectively be utilized for cultivar screening and fulfilling the nutritional and functional needs of the population while minimizing the environmental impacts associated with traditional agriculture practices.

Rocket (Eruca vesicaria subsp. sativa) is a source of sulfur-containing glucosinolates (GSLs). GSLs and their breakdown hydrolysis products (GHPs) are responsible for health-related benefits, such as anti-cancer and anti-neurodegenerative properties. Understanding how phytochemical composition changes between cultivation environments is key to developing cultivars with improved nutritional quality. Two consecutive harvests (first and second regrowth) of crops, grown in both Italy and the UK, were used to determine the phytochemical and transcriptomic differences between six lines of Eruca. Samples were taken upon delivery from field sites (D0) and after five days of cold storage (D5) for each location. Leaves were analysed for sulfur content, volatile organic compounds (VOCs), GSLs, GHPs, and sugars. Transcriptome data were associated with metabolite profiles to identify differentially expressed genes between plants grown in the two environments. VOC compounds (carbon disulfide, methyl thiocyanate) were associated with growth environment and with differences in sulfur metabolism gene expression (APR2, LSU2, LSU3, SDI1, SiR), GSL biosynthesis (MYB28, FMOGS-OX2) and GHP formation (ESM1, TGG1, TGG2). The concentrations of sugars were an order of magnitude greater in UK grown samples (up to 29.9 mg g-1 dry weight; dw). Sulfur content was significantly higher in the Italy plant samples (11.4 -- 20.1 mg g-1 dw), which was in turn associated with higher concentrations of GSLs (pentyl GSL, up to 15.8 mmol g-1 dw; sinigrin, up to 0.005 mmol g-1 dw; glucoraphanin, up to 5.1 mmol g-1 dw; glucorucolamine, up to 23.6 mmol g-1 dw; neoglucobrassicin, up to 5.3 mmol g-1 dw) and hydrolysis products (sativin, up to 13.5 mmol g-1 dw; erucin, up to 1 mmol g-1 dw; sulforaphane, up to 34.7 mmol g-1 dw). VOC profiles of plants cultivated in the UK were distinct from Italy grown plants, with higher relative abundances of alkanes and esters in second cut and shelf-life (D5) samples. The data indicate a significant interaction of cultivar response with environment, highlighting the difficulty of producing Eruca crops with consistent phytochemical and postharvest traits. Genes with differential expression between plants grown in Italy and the UK could be used as markers of phytochemical quality and composition. [ABSTRACT FROM AUTHOR]

Glucosinolates (GSLs) and their degradation products in radish confer plant defense, promote human health, and generate pungent flavor. However, the intact GSLs in radish have not been investigated comprehensively yet. Here, an accurate qualitative and quantitative analyses of 15 intact GSLs from radish, including four major GSLs of glucoraphasatin (GRH), glucoerucin (GER), glucoraphenin (GRE), and 4-methoxyglucobrassicin (4MGBS), were conducted using UHPLC-HRMS/MS in combination with UHPLC-QqQ-MS/MS. Simultaneously, three isomers of hexyl GSL, 3-methylpentyl GSL, and 4-methylpentyl GSL were identified in radish. The highest content of GSLs was up to 232.46 μmol/g DW at the 42 DAG stage in the ‘SQY’ taproot, with an approximately 184.49-fold increase compared to the lowest content in another sample. That the GSLs content in the taproots of two radishes fluctuated in a similar pattern throughout the five vegetative growth stages according to the metabolic profiling, whereas the GSLs content in the ‘55’ leaf steadily decreased over the same period. Additionally, the proposed biosynthetic pathways of radish-specific GSLs were elucidated in this study. Our findings will provide an abundance of qualitative and quantitative data on intact GSLs, as well as a method for detecting GSLs, thus providing direction for the scientific progress and practical utilization of GSLs in radish.

Glucosinolates are key defense compounds of plants in Brassicales order, and their accumulation in seeds is essential for the protection of the next generation. Recently, members of the Usually Multiple Amino acids Move In and Out Transporter (UMAMIT) family were shown to be essential for facilitating transport of seed-bound glucosinolates from site of synthesis within the reproductive organ to seeds. Here, we set out to identify amino acid residues responsible for glucosinolate transport activity of the main seed glucosinolate exporter UMAMIT29 in Arabidopsis thaliana. Based on a predicted model of UMAMIT29, we propose that the substrate transporting cavity consists of 51 residues, of which four are highly conserved residues across all the analyzed homologs of UMAMIT29. A comparison of the putative substrate binding site of homologs within the brassicaceous-specific, glucosinolate-transporting clade with the non-brassicaceous-specific, non-glucosinolate-transporting UMAMIT32 clade identified 11 differentially conserved sites. When each of the 11 residues of UMAMIT29 was individually mutated into the corresponding residue in UMAMIT32, five mutant variants (UMAMIT29#V27F, UMAMIT29#M86V, UMAMIT29#L109V, UMAMIT29#Q263S, and UMAMIT29#T267Y) reduced glucosinolate transport activity over 75% compared to wild-type UMAMIT29. This suggests that these residues are key for UMAMIT29-mediated glucosinolate transport activity and thus potential targets for blocking the transport of glucosinolates to the seeds. [ABSTRACT FROM AUTHOR]

Polycomb group proteins (PcG) play a crucial role in developmental programs in eukaryotic organisms, including plants. PcG-mediated gene repression is achieved by epigenetic histone modification on target chromatins. Loss of PcG components leads to severe developmental defects. CURLY LEAF (CLF), a PcG component in Arabidopsis, catalyzes the trimethylation of histone H3 on lysine 27 (H3K27me3), a repressive histone mark in numerous genes in Arabidopsis. In this study, we isolated a single homolog of Arabidopsis CLF, namely, BrCLF, in Brassica rapa ssp. trilocularis. Transcriptomic analysis revealed that BrCLF participated in B. rapa developmental processes, such as seed dormancy, leaf and flower organ development, and floral transition. BrCLF was also involved in stress signaling and stress-responsive metabolism, such as aliphatic and indolic glucosinolate metabolism in B. rapa. Epigenome analysis showed that H3K27me3 was substantially enriched in genes related to these developmental and stress-responsive processes. Thus, this study provided a basis for elucidating the molecular mechanism of the PcG-mediated regulation of development and stress responses in B. rapa.

Watercress (Nasturtium officinale) is a rich source of secondary metabolites with disease-preventing and/or health-promoting properties. Herein, we have utilized extraction procedures to isolate fractions of polyphenols, glucosinolates and isothiocyanates to determine their identification, and quantification. In doing so, we have utilized reproducible analytical methodologies based on liquid chromatography with tandem mass spectrometry by either positive or negative ion mode. Due to the instability and volatility of isothiocyanates, we followed an ammonia derivatization protocol which converts them into respective ionizable thiourea derivatives. The analytes’ content distribution map was created on watercress flowers, leaves and stems. We have demonstrated that watercress contains significantly higher levels of gluconasturtiin, phenethyl isothiocyanate, quercetin-3-O-rutinoside and isorhamnetin, among others, with their content decreasing from flowers (82.11 ± 0.63, 273.89 ± 0.88, 1459.30 ± 12.95 and 289.40 ± 1.37 ng/g of dry extract respectively) to leaves (32.25 ± 0.74, 125.02 ± 0.52, 1197.86 ± 4.24 and 196.47 ± 3.65 ng/g of det extract respectively) to stems (9.20 ± 0.11, 64.7 ± 0.9, 41.02 ± 0.18, 65.67 ± 0.84 ng/g of dry extract respectivbely). Pearson’s correlation analysis has shown that the content of isothiocyanates doesn’t depend only on the bioconversion of individual glucosinolates but also on other glucosinolates of the same group. Overall, we have provided comprehensive analytical data of the major watercress metabolites thereby providing an opportunity to exploit different parts of watercress for potential therapeutic applications.

The synthesis of indole-3-acetonitrile (IAN) from the indolic glucosinolate (iGSL) glucobrassicin (GB) is a unique trait of members of the Brassicales. To assess the contribution of this pathway to indole-3-acetic acid (IAA) synthesis under stress conditions, drought stress (DS) experiments with Arabidopsis thaliana were performed in vitro. Analysis of GSLs in DS plants revealed higher contents of GB in shoots and roots compared to control plants. Deuterium incorporation experiments showed the highest turnover of GB compared to all other GSLs during drought conditions. Evidence suggests the involvement of the thioglucosidase BGLU18 in the degradation of GB. The nitrile specifier proteins NSP1 and NSP5 are known to direct the GSL hydrolysis towards formation of IAN. Nitrilases like NIT2 are able to subsequently synthesize IAA from IAN. Expression of BGLU18, NSP1, NSP5 and NIT2 and contents of GB, IAN and IAA were significantly elevated in DS plants compared to control plants suggesting the increased use of GB as IAA source. Significantly higher contents of reactive oxygen species in DS bglu18 and epithionitrile specifier protein (esp) mutants compared to Col-0 indicate higher stress levels in these mutants highlighting the need for both proteins in DS plants. Furthermore, GB accumulation in leaves was higher in both mutants during DS when compared to Col-0 indicating enhanced synthesis of GB due to a lack of breakdown products. This work provides the first evidence for the breakdown of iGSLs to IAN which seems to be used for synthesis of IAA in DS A. thaliana plants.

Wasabi (Eutrema japonicum), also known as Japanese horseradish, is a perennial herb widely used in Japanese cuisine for its special flavour. The health-promoting phytochemicals and antioxidant capacity of four organs (leaf, petiole, rhizome, and root) of two cultivars (Chuankui–1 and Chuankui–2) of wasabi from two producing areas, Leibo and Guangyuan in Sichuan Province, China, were investigated in this study. The results showed that leaves were rich in pigments, soluble protein, ascorbic acid, and total phenolics and had the highest antioxidant capacity. Soluble sugars were highest in the petioles and were 1.1- to 5-fold higher than those in the other three organs. Glucosinolates and glucosinolate breakdown products (GBPs) were the most abundant in rhizomes, and their maximum values were 271.61 mmol kg-1 DW and 249.78 mmol kg-1 DW, respectively. The rhizomes of Chuankui–1 in Leibo and the leaves of Chuankui–1 in Guangyuan were superior in terms of glucosinolates and GBPs. These findings provide new insights that will aid the use of wasabi cultivars; they also have implications for the environmental characteristics needed to obtain better quality wasabi products. In the future, metabolome and transcriptome can be used to analyze the potential mechanism of differences among typical varieties, origins and parts.

Brassicaceae family includes an important group of plants of great scientific interest, e.g., the model plant Arabidopsis thaliana, and of economic interest, such as crops of the genus Brassica (Brassica oleracea, Brassica napus, Brassica rapa, etc.). This group of plants is characterized by the synthesis and accumulation in their tissues of secondary metabolites called glucosinolates (GSLs), sulfur-containing compounds mainly involved in plant defense against pathogens and pests. Brassicaceae plants are among the 30% of plant species that cannot establish optimal associations with mycorrhizal hosts (together with other plant families such as Proteaceae, Chenopodiaceae, and Caryophyllaceae), and GSLs could be involved in this evolutionary process of non-interaction. However, this group of plants can establish beneficial interactions with endophytic fungi, which requires a reduction of defensive responses by the host plant and/or an evasion, tolerance, or suppression of plant defenses by the fungus. Although much remains to be known about the mechanisms involved in the Brassicaceae-endophyte fungal interaction, several cases have been described, in which the fungi need to interfere with the GSL synthesis and hydrolysis in the host plant, or even directly degrade GSLs before they are hydrolyzed to antifungal isothiocyanates. Once the Brassicaceae-endophyte fungus symbiosis is formed, the host plant can obtain important benefits from an agricultural point of view, such as plant growth promotion and increase in yield and quality, increased tolerance to abiotic stresses, and direct and indirect control of plant pests and diseases. This review compiles the studies on the interaction between endophytic fungi and Brassicaceae plants, discussing the mechanisms involved in the success of the symbiosis, together with the benefits obtained by these plants. Due to their unique characteristics, the family Brassicaceae can be seen as a fruitful source of novel beneficial endophytes with applications to crops, as well as to generate new models of study that allow us to better understand the interactions of these amazing fungi with plants.

Biofortification is the process that aims to enrich crops in micronutrients and valuable compounds. Selenium (Se) biofortification has particularly attracted increasing interest in recent times due to the growing number of individuals suffering from Se deficiency. Selenate and selenite are the Se forms most frequently administered to crops. In this study, Se was applied foliarly as selenate at 2.5, 5, or 10 mg per plant to two rocket species, Diplotaxis tenuifolia and Eruca sativa, grown in soil and the effects in terms of Se enrichment and content of primary and secondary metabolites were comparatively analyzed. We also compared our results with those obtained previously when selenate was supplied to the same species in hydroponics by addition to the nutrient solution. In most cases, the results were the opposite. In E. sativa, foliar Se treatment was more effective in promoting Se accumulation, sulfur (S), cysteine, and glucosinolates. No significant effect of Se was evident on total phenolic content, but there were individual phenols. Among amino acids, the content of proline was increased by Se, perhaps to counteract osmotic stress due to high Se accumulation. In D. tenuifolia, the content of S and cysteine decreased under Se treatment, but the amount of glutathione was steady, suggesting a preferred assimilation of cysteine toward the synthesis of this antioxidant. Consistent, the content of methionine and glucosinolates was reduced. The content of total phenolics was enhanced only by the low Se dosage. In both species, selenocysteine (SeCys) was identified, the content of which was higher compared to plants grown hydroponically. Concluding, most metabolic differences between rocket species were observed at high Se supplementation. Low Se foliar fertilization was effective in an enriching rocket in Se without affecting other phytochemicals. However, the Se dosages sufficient for biofortification could be even lower, as the Se concentration in rocket treated with 2.5 mg Se per plant was still very high and the edible part should not be eaten undiluted. Also, a single method of Se supplementation does not appear to be optimal for all plant species or the same species, as the metabolic responses could be very different.

IQ Domain 1 (IQD1) is a novel Arabidopsis thaliana calmodulin-binding protein, which was found to be a positive regulator of glucosinolate (GS) accumulation and plant defense responses against insects. We demonstrate here that the IQD1 overexpressing line (IQD1OXP) was also more resistant also to the necrotrophic fungus Botrytis cinerea, whereas an IQD1 knockout line (iqd1-1) was much more sensitive. Furthermore, we showed that IQD1 is up-regulated by jasmonic acid (JA) and downregulated by salicylic acid (SA). A comparison of whole transcriptome expression between iqd1-1 and wild type plants revealed a substantial downregulation of genes involved in plant defense and hormone regulation. Further examination revealed a marked reduction of SA and increases in the levels of ethylene, JA and abscisic acid response genes in the iqd1-1 line. Moreover, quantification of SA, JA, and abscisic acids in IQD1OXP and iqd1-1 lines relative to the wild type, showed a significant reduction in endogenous JA levels in the knockout line, simultaneously with increased SA levels. Relations between IQD1OXP and mutants defective in plant-hormone response indicated that IQD1 cannot rescue the absence of NPR1 or impaired SA accumulation in the NahG line. IQD1 cannot rescue ein2 or eto1 mutations connected to the ethylene pathway involved in both defense responses against B. cinerea and in regulating GS accumulation. Furthermore, IQD1cannot rescue the aos, coi1 or jar1mutations, all involved in the defense response against B. cinerea and it depends on JAR1 to control indole glucosinolate accumulation. We also found that in the B. cinerea, which infected the iqd1-1 mutant, the most abundant upregulated group of proteins is involved in the degradation of complex carbohydrates, as correlated with the sensitivity of this mutant. In summary, our results suggest that IQD1 is an important A. thaliana defensive protein against B. cinerea that is integrated into several important pathways, such as those involved in plant defense and hormone responses.

Initiation of plant immune signaling requires recognition of conserved molecular patterns from microbes and herbivores by plasma membrane-localized pattern recognition receptors. Additionally, plants produce and secrete numerous small peptide hormones, termed phytocytokines, which act as secondary danger signals to modulate immunity. In Arabidopsis, the Brassicae-specific SERINE RICH ENDOGENOUS PEPTIDE (SCOOP) family consists of 14 members that are perceived by the leucine-rich repeat receptor kinase MALE DISCOVERER 1-INTERACTING RECEPTOR LIKE KINASE 2 (MIK2). Recognition of SCOOP peptides elicits generic early signaling responses but knowledge on how and if SCOOPs modulate specific downstream immune defenses is limited. We report here that depletion of MIK2 or the single PROSCOOP12 precursor results in decreased Arabidopsis resistance against the generalist herbivore Spodoptera littoralis but not the specialist Pieris brassicae. Increased performance of S. littoralis on mik2-1 and proscoop12 is accompanied by a diminished accumulation of jasmonic acid, jasmonate-isoleucine and indolic glucosinolates. Additionally, we show transcriptional activation of the PROSCOOP gene family in response to insect herbivory. Our data therefore indicate that perception of endogenous SCOOP peptides by MIK2 modulates the jasmonate pathway and thereby contributes to enhanced defense against a generalist herbivore.

Glucosinolates (GSLs) are naturally occurring secondary metabolites found in the Brassicaceae family, which mainly synthesize in the siliques with a wide range of functions. In this study, we investigated the effects of lights on metabolites in siliques of rapeseed through ultra high-performance liquid chromatography (UPLC)—heated electrospray ionization (HESI)–tandem mass spectrometry (MS/MS). A total of 249 metabolites, including 29 phenolic acids, 38 flavonoids, 22 GSLs, 93 uncalculated and 67 unknown compounds, were identified in siliques of rapeseed. Meanwhile, 62 metabolites showed significant differences after shading treatment, which were mainly GSLs and unknown compounds. Interestingly, the amounts of 10 GSLs had high accumulation levels in siliques, while the expression levels of their corresponding biosynthetic genes (AOP, GSL-OH, IGMT, and ST5a) were obviously reduced after shading treatment. Further evidence showed that the amounts of GSLs were significantly reduced in seeds, in accordance with the expression profiles of transporter genes (BnaGTRs). Our findings indicated that lights could affect the accumulation and transportation of GSLs from siliques to seeds in rapeseed. Therefore, this study facilitates a better understanding of metabolic characteristics of siliques and provides insight into the importance of light for GSLs accumulation and transportation in siliques and seeds of rapeseed.

Watercress (Nasturtium officinale) is a rich source of secondarymetabolites with disease-preventing and/or health-promoting properties. Herein, we have utilized extraction procedures to isolate fractions of polyphenols, glucosinolates and isothiocyanates to determine their identification, and quantification. In doing so, we have utilized reproducible analytical methodologies based on liquid chromatography with tandem mass spectrometry by either positive or negative ion mode. Due to the instability and volatility of isothiocyanates, we followed an ammonia derivatization protocol which converts them into respective ionizable thiourea derivatives. The analytes' content distribution map was created on watercress flowers, leaves and stems. We have demonstrated that watercress contains significantly higher levels of gluconasturtiin, phenethyl isothiocyanate, quercetin-3-O-rutinoside and isorhamnetin, among others, with their content decreasing from flowers (82.11 ± 0.63, 273.89 ± 0.88, 1459.30 ± 12.95 and 289.40 ± 1.37 ng/g of dry extract respectively) to leaves (32.25 ± 0.74, 125.02 ± 0.52, 1197.86 ± 4.24 and 196.47 ± 3.65 ng/g of det extract respectively) to stems (9.20 ± 0.11, 64.7 ± 0.9, 41.02 ± 0.18, 65.67 ± 0.84 ng/g of dry extract respectivbely). Pearson's correlation analysis has shown that the content of isothiocyanates doesn't depend only on the bioconversion of individual glucosinolates but also on other glucosinolates of the same group. Overall, we have provided comprehensive analytical data of the major watercress metabolites thereby providing an opportunity to exploit different parts of watercress for potential therapeutic applications. [ABSTRACT FROM AUTHOR]

Current research was conducted to explore the effects of liquiritoside on the growth and physiochemical features of Chinese flowering cabbage (Brassica rapa subsp. parachinensis) under lead (Pb) stress. Lead stressed B. rapa plants exhibited decreased growth parameters, chlorophyll, and carotenoid contents. Moreover, Pb toxicity escalated the synthesis of malondialdehyde (MDA), hydrogen peroxide (H2O2), flavonoids, phenolics, and proline in treated plants. Nevertheless, foliar application of liquiritoside mitigated Pb toxicity by decreasing oxidative stress by reducing cysteine, H2O2, and MDA contents in applied plants. Liquiritoside significantly increased plant height, shoot fresh weight and dry weight, number of leaves, and marketable value of Chinese flowering cabbage plants exposed to Pb toxicity. This biotic elicitor also enhanced the proline, glutathione, total phenolics, and flavonoid contents in Chinese flowering cabbage plants exposed to Pb stress compared with the control. Additionally, total glucosinolate content, phytochelatins (PCs), and non-protein thiols were effectively increased in plants grown under Pb regimes compared with the control plants. Overall, foliar application of liquiritoside can markedly alleviate Pb stress by restricting Pb translocation in Chinese flowering cabbage.

Biofortification is the process that aims to enrich crops in micronutrients and valuable compounds. Selenium (Se) biofortification has particularly attracted increasing interest in recent times due to the growing number of individuals suffering from Se deficiency. Selenate and selenite are the Se forms most frequently administered to crops. In this study, Se was applied foliarly as selenate at 2.5, 5, or 10 mg per plant to two rocket species, Diplotaxis tenuifolia and Eruca sativa, grown in soil and the effects in terms of Se enrichment and content of primary and secondary metabolites were comparatively analyzed. We also compared our results with those obtained previously when selenate was supplied to the same species in hydroponics by addition to the nutrient solution. In most cases, the results were the opposite. In E. sativa, foliar Se treatment was more effective in promoting Se accumulation, sulfur (S), cysteine, and glucosinolates. No significant effect of Se was evident on total phenolic content, but there were individual phenols. Among amino acids, the content of proline was increased by Se, perhaps to counteract osmotic stress due to high Se accumulation. In D. tenuifolia, the content of S and cysteine decreased under Se treatment, but the amount of glutathione was steady, suggesting a preferred assimilation of cysteine toward the synthesis of this antioxidant. Consistent, the content of methionine and glucosinolates was reduced. The content of total phenolics was enhanced only by the low Se dosage. In both species, selenocysteine (SeCys) was identified, the content of which was higher compared to plants grown hydroponically. Concluding, most metabolic differences between rocket species were observed at high Se supplementation. Low Se foliar fertilization was effective in an enriching rocket in Se without affecting other phytochemicals. However, the Se dosages sufficient for biofortification could be even lower, as the Se concentration in rocket treated with 2.5 mg Se per plant was still very high and the edible part should not be eaten undiluted. Also, a single method of Se supplementation does not appear to be optimal for all plant species or the same species, as the metabolic responses could be very different. [ABSTRACT FROM AUTHOR]

Hybrids have been successfully used to improve crop productivity, including Brassicas. Nucleo-cytoplasmic interactions have been reported to influence the expression of resistance to insect pests in several crops. We studied the effects of Cytoplasmic Male Sterility (CMS) in Brassica juncea carrying alien cytoplasms and their respective maintainer (B) lines on the antibiosis mechanism of resistance, involving development, survival, reproduction potential and population build-up of mustard aphid, Lipaphis erysimi, and the levels of defense phyto-chemicals. Present findings revealed that the numbers of aphids/plant, aphid multiplication rate and aphid resistance index were lower on ber CMS under natural, mori CMS under artificial infestation conditions, and juncea under both the test conditions indicating nucleocytoplasmic interactions for aphid reaction. Across cytoplasms, nymphal, reproductive and total developmental periods were significantly longer on SEJ 8, NPJ 161, LES 39, and NPJ 93, while the reproductive potential and survival were lower on PM 30, Pusa Tarak and SEJ 8 as compared to other nuclear backgrounds. Across nuclear backgrounds, nymphal, reproductive and total developmental periods were significantly longer on ber CMS, while reproductive potential and survival were lower on ber and mori CMS as compared to other cytoplasms. Total glucosinolates were significantly greater and myrosinase lower in Pusa Agrani, SEJ 8, LES 39, PM 30, NPJ 112, and Pusa Tarak as compared to the other nuclear backgrounds. Furthermore, total glucosinolates were significantly greater and myrosinase lower in ber CMS and juncea as compared to other cytoplasms. The studies suggest that CMS as well as cytoplasmic and nuclear gene interactions regulate the expression of defense compounds such as glucosinolates and determine the expression of resistance/susceptibility to L. erysimi. These findings shall help in identification of suitable L. erysimi tolerant nucleo-cytoplasmic combinations for their deployment in B. juncea hybrid breeding program. [ABSTRACT FROM AUTHOR]

Brassicaceae family includes an important group of plants of great scientific interest, e.g., the model plant Arabidopsis thaliana, and of economic interest, such as crops of the genus Brassica (Brassica oleracea, Brassica napus, Brassica rapa, etc.). This group of plants is characterized by the synthesis and accumulation in their tissues of secondary metabolites called glucosinolates (GSLs), sulfur-containing compounds mainly involved in plant defense against pathogens and pests. Brassicaceae plants are among the 30% of plant species that cannot establish optimal associations with mycorrhizal hosts (together with other plant families such as Proteaceae, Chenopodiaceae, and Caryophyllaceae), and GSLs could be involved in this evolutionary process of non-interaction. However, this group of plants can establish beneficial interactions with endophytic fungi, which requires a reduction of defensive responses by the host plant and/or an evasion, tolerance, or suppression of plant defenses by the fungus. Although much remains to be known about the mechanisms involved in the Brassicaceae-endophyte fungal interaction, several cases have been described, in which the fungi need to interfere with the GSL synthesis and hydrolysis in the host plant, or even directly degrade GSLs before they are hydrolyzed to antifungal isothiocyanates. Once the Brassicaceae-endophyte fungus symbiosis is formed, the host plant can obtain important benefits from an agricultural point of view, such as plant growth promotion and increase in yield and quality, increased tolerance to abiotic stresses, and direct and indirect control of plant pests and diseases. This review compiles the studies on the interaction between endophytic fungi and Brassicaceae plants, discussing the mechanisms involved in the success of the symbiosis, together with the benefits obtained by these plants. Due to their unique characteristics, the family Brassicaceae can be seen as a fruitful source of novel beneficial endophytes with applications to crops, as well as to generate new models of study that allow us to better understand the interactions of these amazing fungi with plants. [ABSTRACT FROM AUTHOR]

Terrestrial orchids can form tubers, organs modified to store energy reserves. Tubers are an attractive source of nutrients, and salep, a flour made from dried orchid tubers, is the source of traditional beverages. Tubers also contain valuable secondary metabolites and are used in traditional medicine. The extensive harvest of wild orchids is endangering their populations in nature; however, orchids can be cultivated and tubers mass-produced. This work illustrates the importance of plant-fungus interaction in shaping the content of orchid tubers in vitro. Orchid plants of Dactylorhiza sp. grown in asymbiotic culture were inoculated with a fungal isolate from Tulasnella calospora group and, after 3 months of co-cultivation, tubers were analyzed. The fungus adopted the saprotrophic mode of life, but no visible differences in the morphology and biomass of the tubers were detected compared to the mock-treated plants. To elucidate the mechanisms protecting the tubers against fungal infestation, proteome, metabolome, and lipidome of tubers were analyzed. In total, 1,526, 174, and 108 proteins, metabolites, and lipids were quantified, respectively, providing a detailed snapshot of the molecular process underlying plant-microbe interaction. The observed changes at the molecular level showed that the tubers of inoculated plants accumulated significantly higher amounts of antifungal compounds, including phenolics, alkaloid Calystegine B2, and dihydrophenanthrenes. The promoted antimicrobial effects were validated by observing transient inhibition of Phytophthora cactorum growth. The integration of omics data highlighted the promotion of flavonoid biosynthesis, the increase in the formation of lipid droplets and associated production of oxylipins, and the accumulation of auxin in response to T. calospora. Taken together, these results provide the first insights into the molecular mechanisms of defense priming in orchid tubers and highlight the possible use of fungal interactors in biotechnology for the production of orchid secondary metabolites. [ABSTRACT FROM AUTHOR]

Western hemlock (Tsuga heterophylla) is highly susceptible to Annosus root and butt rot disease, caused by Heterobasidion occidentale across its native range in western North America. Understanding molecular mechanisms of tree defense and dissecting genetic components underlying disease resistance will facilitate forest breeding and disease control management. The aim of this study was to profile host transcriptome reprogramming in response to pathogen infection using RNA-seq analysis. Inoculated seedlings were clearly grouped into three types: quantitative resistant (QR), susceptible (Sus), and un-infected (Uif), based on profiles of H. occidentale genes expressed in host tissues. Following de novo assembly of a western hemlock reference transcriptome with more than 33,000 expressed genes, the defensive transcriptome reprogramming was characterized and a set of differentially expressed genes (DEGs) were identified with gene ontology (GO) annotation. The QR seedlings showed controlled and coordinated molecular defenses against biotic stressors with enhanced biosynthesis of terpenoids, cinnamic acids, and other secondary metabolites. The Sus seedlings showed defense responses to abiotic stimuli with a few biological processes enhanced (such as DNA replication and cell wall organization), while others were suppressed (such as killing of cells of other organism). Furthermore, non-synonymous single nucleotide polymorphisms (ns-SNPs) of the defense- and resistance-related genes were characterized with high genetic variability. Both phylogenetic analysis and principal coordinate analysis (PCoA) revealed distinct evolutionary distances among the samples. The QR and Sus seedlings were well separated and grouped into different phylogenetic clades. This study provides initial insight into molecular defense and genetic components of western hemlock resistance against the Annosus root and butt rot disease. Identification of a large number of genes and their DNA variations with annotated functions in plant resistance and defense promotes the development of genomics-based breeding strategies for improved western hemlock resistance to H. occidentale. [ABSTRACT FROM AUTHOR]

The plant microbiome is an increasingly intensive research area, with significance in agriculture, general plant health, and production of bioactive natural products. Correlations between the fungal endophytic communities and plant chemistry can provide insight into these interactions, and suggest key contributors on both the chemical and fungal side. In this study, roots of various horseradish (Armoracia rusticana) accessions grown under the same conditions were sampled in two consecutive years and chemically characterized using a quality controlled, untargeted metabolomics approach by LC-ESI-MS/MS. Sinigrin, gluconasturtiin, glucoiberin, and glucobrassicin were also quantified. Thereafter, a subset of roots from eight accessions (n = 64) with considerable chemical variability was assessed for their endophytic fungal community, using an ITS2 amplicon-based metagenomic approach using a custom primer with high coverage on fungi, but no amplification of host internal transcribed spacer (ITS). A set of 335 chemical features, including putatively identified flavonoids, phospholipids, peptides, amino acid derivatives, indolic phytoalexins, a glucosinolate, and a glucosinolate downstream product was detected. Major taxa in horseradish roots belonged to Cantharellales, Glomerellales, Hypocreales, Pleosporales, Saccharomycetales, and Sordariales. Most abundant genera included typical endophytes such as Plectosphaerella, Thanatephorus, Podospora, Monosporascus, Exophiala , and Setophoma. A surprising dominance of single taxa was observed for many samples. In summary, 35.23% of reads of the plant endophytic fungal microbiome correlated with changes in the plant metabolome. While the concentration of flavonoid kaempferol glycosides positively correlated with the abundance of many fungal strains, many compounds showed negative correlations with fungi including indolic phytoalexins, a putative glucosinolate but not major glucosinolates and a glutathione isothiocyanate adduct. The latter is likely an in vivo glucosinolate decomposition product important in fungal arrest. Our results show the potency of the untargeted metabolomics approach in deciphering plant–microbe interactions and depicts a complex array of various metabolite classes in shaping the endophytic fungal community. [ABSTRACT FROM AUTHOR]

Cruciferous plants in the order Brassicales defend themselves from herbivory using glucosinolates: sulfur-containing pro-toxic metabolites that are activated by hydrolysis to form compounds, such as isothiocyanates, which are toxic to insects and other organisms. Some herbivores are known to circumvent glucosinolate activation with glucosinolate sulfatases (GSSs), enzymes that convert glucosinolates into inactive desulfoglucosinolates. This strategy is a major glucosinolate detoxification pathway in a phloem-feeding insect, the silverleaf whitefly Bemisia tabaci, a serious agricultural pest of cruciferous vegetables. In this study, we identified and characterized an enzyme responsible for glucosinolate desulfation in the globally distributed B. tabaci species MEAM1. In in vitro assays, this sulfatase showed a clear preference for indolic glucosinolates compared with aliphatic glucosinolates, consistent with the greater representation of desulfated indolic glucosinolates in honeydew. B. tabaci might use this detoxification strategy specifically against indolic glucosinolates since plants may preferentially deploy indolic glucosinolates against phloem-feeding insects. In vivo silencing of the expression of the B. tabaci GSS gene via RNA interference led to lower levels of desulfoglucosinolates in honeydew. Our findings expand the knowledge on the biochemistry of glucosinolate detoxification in phloem-feeding insects and suggest how detoxification pathways might facilitate plant colonization in a generalist herbivore.

Quantitative trait loci (QTL) mapping studies enable identification of loci that are part of regulatory networks controlling various phenotypes. Detailed investigations of genes within these loci are required to ultimately understand the function of individual genes and how they interact with other players in the network. In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds. Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation. Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation.

The evolution of plant metabolic diversity is largely driven by gene duplication and ensuing sub-functionalization and/or neo-functionalization to generate new enzymatic activities. However, it is not clear whether the transcription factors (TFs) regulating these new enzyme encoding genes were required to co-evolve with these genes in a similar fashion or if these new genes can be captured by existing conserved TFs to provide the appropriate expression pattern. In this study, we found two conserved TFs, MYB115, and MYB118, co-expressed with the key enzyme encoding genes in the newly evolved benzoyloxy glucosinolate (GLS) pathway. These TFs interacted with the promoters of the GLS biosynthetic genes and negatively influenced their expression. Similarly, the GLS profiles of these two TFs knockouts showed that they influenced the aliphatic GLS accumulation within seed, leaf and flower, while they mainly expressed in seeds. Further studies indicated that they are functionally redundant and epistatically interact to control the transcription of GLS genes. Complementation study confirmed their roles in regulating the aliphatic GLS biosynthesis. These results suggest that the newly evolved enzyme encoding genes for novel metabolites can be regulated by conserved TFs, which helps to improve our model for newly evolved genes regulation.

Naturally variable regulatory networks control different biological processes including reproduction and defense. This variation within regulatory networks enables plants to optimize defense and reproduction in different environments. In this study we investigate the ability of two enzyme-encoding genes in the glucosinolate pathway, AOP2 and AOP3, to affect glucosinolate accumulation and flowering time. We have introduced the two highly similar enzymes into two different AOP (null) accessions, Col-0 and Cph-0, and found that the genes differ in their ability to affect glucosinolate levels and flowering time across the accessions. This indicated that the different glucosinolates produced by AOP2 and AOP3 serve specific regulatory roles in controlling these phenotypes. While the changes in glucosinolate levels were similar in both accessions, the effect on flowering time was dependent on the genetic background pointing to natural variation in cross-talk between defense chemistry and onset of flowering. This variation likely reflects an adaptation to survival in different environments.

Effect of packaging baby mustard into bags of different color under light exposure on its visual quality and the content of chlorophyll, carotenoids, and glucosinolates at 20°C was investigated. Packaging with seven color bags under light exposure prolonged the shelf life, especially green (GB), blue (BB), and transparent (TB) bags with holes, and their shelf life was 1.7, 1.6, and 1.6 times that of the control, respectively. The GB and BB treatments delayed the deterioration of the sensory quality in baby mustard during storage. The BB and TB treatments not only increased chlorophyll and carotenoids content in baby mustard during storage but also enhanced the accumulation of glucosinolates by inhibiting their degradation, especially the BB treatment. Overall, the results demonstrate that the BB treatment is a promising technique for maintaining the postharvest quality of baby mustard. [ABSTRACT FROM AUTHOR]

IQ Domain 1 (IQD1) is a novel Arabidopsis thaliana calmodulin-binding protein, which was found to be a positive regulator of glucosinolate (GS) accumulation and plant defense responses against insects. We demonstrate here that the IQD1 overexpressing line (IQD1 OXP) was also more resistant also to the necrotrophic fungus Botrytis cinerea , whereas an IQD1 knockout line (iqd1-1) was much more sensitive. Furthermore, we showed that IQD1 is up-regulated by jasmonic acid (JA) and downregulated by salicylic acid (SA). A comparison of whole transcriptome expression between iqd1-1 and wild type plants revealed a substantial downregulation of genes involved in plant defense and hormone regulation. Further examination revealed a marked reduction of SA and increases in the levels of ethylene, JA and abscisic acid response genes in the iqd1-1 line. Moreover, quantification of SA, JA, and abscisic acids in IQD1 OXP and iqd1-1 lines relative to the wild type, showed a significant reduction in endogenous JA levels in the knockout line, simultaneously with increased SA levels. Relations between IQD1 OXP and mutants defective in plant-hormone response indicated that IQD1 cannot rescue the absence of NPR1 or impaired SA accumulation in the NahG line. IQD1 cannot rescue ein2 or eto1 mutations connected to the ethylene pathway involved in both defense responses against B. cinerea and in regulating GS accumulation. Furthermore, IQD1cannot rescue the aos , coi1 or jar1 mutations, all involved in the defense response against B. cinerea and it depends on JAR1 to control indole glucosinolate accumulation. We also found that in the B. cinerea , which infected the iqd1-1 mutant, the most abundant upregulated group of proteins is involved in the degradation of complex carbohydrates, as correlated with the sensitivity of this mutant. In summary, our results suggest that IQD1 is an important A. thaliana defensive protein against B. cinerea that is integrated into several important pathways, such as those involved in plant defense and hormone responses. [ABSTRACT FROM AUTHOR]

Broccoli cultivars that have enhanced accumulation of methionine-derived glucosinolates have been developed through the introgression of a novel allele of the MYB28 transcription factor from the wild species Brassica villosa. Through a novel k-mer approach, we characterised the extent of the introgression of unique B. villosa genome sequences into high glucosinolate broccoli genotypes. RNAseq analyses indicated that the introgression of the B. villosa MYB28 C2 allele resulted in the enhanced expression of the MYB28 transcription factor, and modified expression of genes associated with sulphate absorption and reduction, and methionine and glucosinolate biosynthesis when compared to standard broccoli. A adenine-thymine (AT) short tandem repeat (STR) was identified within the 5′ untranslated region (UTR) B. villosa MYB28 allele that was absent from two divergent cultivated forms of Brassica oleracea , which may underpin the enhanced expression of B. villosa MYB28. [ABSTRACT FROM AUTHOR]

Initiation of plant immune signaling requires recognition of conserved molecular patterns from microbes and herbivores by plasma membrane-localized pattern recognition receptors. Additionally, plants produce and secrete numerous small peptide hormones, termed phytocytokines, which act as secondary danger signals to modulate immunity. In Arabidopsis, the Brassicae -specific SERINE RICH ENDOGENOUS PEPTIDE (SCOOP) family consists of 14 members that are perceived by the leucine-rich repeat receptor kinase MALE DISCOVERER 1-INTERACTING RECEPTOR LIKE KINASE 2 (MIK2). Recognition of SCOOP peptides elicits generic early signaling responses but knowledge on how and if SCOOPs modulate specific downstream immune defenses is limited. We report here that depletion of MIK2 or the single PROSCOOP12 precursor results in decreased Arabidopsis resistance against the generalist herbivore Spodoptera littoralis but not the specialist Pieris brassicae. Increased performance of S. littoralis on mik2-1 and proscoop12 is accompanied by a diminished accumulation of jasmonic acid, jasmonate-isoleucine and indolic glucosinolates. Additionally, we show transcriptional activation of the PROSCOOP gene family in response to insect herbivory. Our data therefore indicate that perception of endogenous SCOOP peptides by MIK2 modulates the jasmonate pathway and thereby contributes to enhanced defense against a generalist herbivore. [ABSTRACT FROM AUTHOR]

Glucosinolates (GSLs) are naturally occurring secondary metabolites found in the Brassicaceae family, which mainly synthesize in the siliques with a wide range of functions. In this study, we investigated the effects of lights on metabolites in siliques of rapeseed through ultra high-performance liquid chromatography (UPLC)—heated electrospray ionization (HESI)–tandem mass spectrometry (MS/MS). A total of 249 metabolites, including 29 phenolic acids, 38 flavonoids, 22 GSLs, 93 uncalculated and 67 unknown compounds, were identified in siliques of rapeseed. Meanwhile, 62 metabolites showed significant differences after shading treatment, which were mainly GSLs and unknown compounds. Interestingly, the amounts of 10 GSLs had high accumulation levels in siliques, while the expression levels of their corresponding biosynthetic genes (AOP , GSL-OH , IGMT , and ST5a) were obviously reduced after shading treatment. Further evidence showed that the amounts of GSLs were significantly reduced in seeds, in accordance with the expression profiles of transporter genes (BnaGTRs). Our findings indicated that lights could affect the accumulation and transportation of GSLs from siliques to seeds in rapeseed. Therefore, this study facilitates a better understanding of metabolic characteristics of siliques and provides insight into the importance of light for GSLs accumulation and transportation in siliques and seeds of rapeseed. [ABSTRACT FROM AUTHOR]

ABA-induced H SB 2 sb O SB 2 sb accumulation can protect plant parts (such as pumpkin-grafted cucumber leaves) against Ca(NO SB 3 sb ) SB 2 sb via ABA/H SB 2 sb O SB 2 sb signaling-led induction of ROS-scavenging machinery (Shu et al.). I salsugineum i chloroplasts showed a constitutive increase and the cell's steady-state regulation of H SB 2 sb O SB 2 sb level which prepared this plant for ROS-control mainly due to an efficient ROS-scavenging machinery including glucosinolates content and well-coordinated tuning of hormonal signaling (Pilarska et al.). H SB 2 sb O SB 2 sb directly mediates metal-induced oxidative signaling, where the production of H SB 2 sb O SB 2 sb may involve H SB 2 sb O SB 2 sb receptors, redox-sensitive transcription factors and inhibition of phosphatases (Miller et al., [8]). Keywords: abiotic stresses; hydrogen peroxide; H2O2-metabolism; priming; plant stress tolerance EN abiotic stresses hydrogen peroxide H2O2-metabolism priming plant stress tolerance 1 4 4 01/31/22 20220128 NES 220128 Reactive oxygen species (ROS) of varied types can be yielded in plants at several primary sites (such as the chloroplast, mitochondria, and peroxisomes) under normal aerobic metabolism I via i processes including photosynthetic and respiratory electron transport chains. [Extracted from the article]

Idioblasts are defined by abnormal shapes, sizes, and contents that are different from neighboring cells. Myrosin cells are Brassicales-specific idioblasts and accumulate a large amount of thioglucoside glucohydrolases (TGGs, also known as myrosinases) in their vacuoles. Myrosinases convert their substrates, glucosinolates, into toxic compounds when herbivories and pests attack plants. In this review, we highlight the similarities and differences between myrosin cells and vascular cells/guard cells (GCs) because myrosin cells are distributed along vascular cells, especially the phloem parenchyma, and myrosin cells share the master transcription factor FAMA with GCs for their cell differentiation. In addition, we analyzed the overlap of cell type-specific genes between myrosin cells and GCs by using published single-cell transcriptomics (scRNA-seq) data, suggesting significant similarities in the gene expression patterns of these two specialized cells. [ABSTRACT FROM AUTHOR]

The fungal pathogen, Leptosphaeria maculans causes a severe and economically important disease to Brassica crops globally, well-known as blackleg. Besides, the anti-oxidative defense response of glucosinolates to fungal pathogens is widely established. Despite notable importance of glucosinolates in blackleg disease resistance the association of glucosinolate pathway genes in glucosinolate mediated defense response after L. maculans infection remains incompletely understood. The current study was designed to identify glucosinolate-biosynthesis specific genes among the eight selected candidates induced by L. maculans and associated alterations in glucosinolate profiles to explore their roles in blackleg resistance at the seedling stage of cabbage plants. The defense responses of four cabbage inbred lines, two resistant and two susceptible, were investigated using two L. maculans isolates, 03-02s and 00-100s. Pathogen-induced glucosinolate accumulation dynamically changed from two days after inoculation to four days after inoculation. In general, glucosinolate biosynthetic genes were induced at 24 h after inoculation and glucosinolate accumulation enhanced at two days after inoculation. An increase in either aliphatic (GIB, GRA) or indolic (GBS and MGBS) glucosinolates was associated with seedling resistance of cabbage. Pearson correlation showed the enhanced accumulation of MGBS, GBS, GIB, GIV and GRA after the inoculation of fungal isolates was associated with expression of specific genes. Principal component analysis separated two resistant cabbage lines—BN4098 and BN4303 from two susceptible cabbage lines—BN4059 and BN4072 for variable coefficients of disease scores, glucosinolate accumulation and expression levels of genes. Enhanced MGBS content against both fungal isolates, contributing to seedling resistance in two interactions—BN4098 × 03-02s and BN4303 × 00-100s and enhanced GBS content only in BN4098 × 03-02s interaction. Aliphatic GRA took part in resistance of BN4098 × 00-100s interaction whereas aliphatic GIB took part is resistance of BN4098 × 03-02s interaction. Aliphatic GIV accumulated upon BN4098 × 03-02s interaction but GSL-OH-Bol033373 and CYP81F2-Bol026044 showed enhanced expression in BN4303 × 03-02s interaction. The association between the selected candidate genes, corresponding glucosinolates, and seedling resistance broaden the horizon of glucosinolate conciliated defense against L. maculans in cabbage seedlings.

Glucosinolates are a diverse group of plant metabolites that characterize the order Brassicales. The MAM locus is one of the most significant QTLs for glucosinolate diversity. However, most of what we understand about evolution at the locus is focused on only a few species and not within a phylogenetic context. In this study, we utilize a micro-synteny network and phylogenetic inference to investigate the origin and diversification of the MAM/IPMS gene family. We uncover unique MAM-like genes found at the orthologous locus in the Cleomaceae that shed light on the transition from IPMS to MAM. In the Brassicaceae, we identify six distinct MAM clades across Lineages I, II, and III. We characterize the evolutionary impact and consequences of local duplications, transpositions, whole genome duplications, and gene fusion events, generating several new hypothesizes on the function and diversity of the MAM locus.

Growing microgreens on trays without substrate in a vertical multilayered growing unit offers several advantages over traditional agriculture methods. This study investigated the yield performance and nutritional quality of five selections of radish microgreens grown in sprouting trays, without a substrate using only water, in an indoor multilayer cultivation system using artificial light. Various parameters were measured, including fresh weight, dry matter, chlorophyll, minerals, amino acids, phenolics, flavonoids, anthocyanins, vitamin C, glucosinolates, and antioxidant activity with four different in vitro assays. After ten days, the biomass had increased by 6-10 times, and the dry matter varied from 4.75-7.65%. The highest yield was obtained from 'Asia red', while the lowest was from 'Koregon red'. However, 'Koregon red' and 'Asia red' had the highest dry matter. 'Asia red' was found to have the highest levels of both Chls and vitamin C compared to the other cultivars, while 'Koregon red' exhibited the highest levels of total phenolics and flavonoids. Although variations in the levels of individual glucosinolates were observed, there were no significant differences in the total content of glucosinolates among the five cultivars. 'Asia purple' had the highest anthocyanin content, while 'Asia green 2' had the lowest. The K, Mg, and Na concentrations were significantly highest in 'Asia green 2', and the highest Ca was recorded in 'Asia purple'. Overall, 'Asia purple' and 'Koregon red' were the best cultivars in terms of nutritional quality among the tested radish microgreens. These cultivars exhibited high levels of dry weight, total phenolics, flavonoids, anthocyanins, essential and total amino acids, and antioxidant activities. Moreover, the implementation of this vertical cultivation method for microgreens, which relies solely on water and seeds known for their tall shoots during the sprouting could hold promise as a sustainable approach. This method can effectively be utilized for cultivar screening and fulfilling the nutritional and functional needs of the population while minimizing the environmental impacts associated with traditional agriculture practices., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Tilahun, Baek, An, Choi, Lee, Hong and Jeong.)

Rocket cultivation is increasing to supply the expanding ready-to-eat market because of its unique taste, but crops are often over fertilized to avoid nitrogen deficiencies. This leads to nitrate accumulation in leaves, and the products of their degradation (nitrites and nitrosamines) have been related to several health problems. Nitrate concentrations in rocket and other leafy vegetables are subject to limits by the EU legislation, yet rocket holds a great nutritional value. Degradation products of glucosinolates (isothiocyanates) have been consistently linked with benefits to human health. We investigated the influence of nitrogen application (1 and 8 mM), species [Eruca sativa (L.) Cav. and Diplotaxis tenuifolia (L.) DC.] and light spectrum (full spectrum, red, blue and red + blue) on the nitrate concentration, nitrate reductase activity and glucosinolate content of rocket grown in a soil-less system. Red light decreased the nitrate concentration with respect to the blue spectrum (4,270 vs. 7,100 mg⋅kg–1 of fresh weight, respectively), but such reduction was influenced by the species and the nitrogen level (significantly higher in D. tenuifolia and with the higher concentration of N). The nitrate reductase activity increased under red light in D. tenuifolia, with the lower N concentration. Rocket is known to contain several health-promoting compounds mainly antioxidants and glucosinolates, as secondary metabolites that act as part of plant defense mechanisms. The total content of glucosinolates was mainly affected by the species (D. tenuifolia showed the highest concentrations). Our results will help growers to tailor light spectra with the aim of reducing nitrate concentration and to remain within EU legislative limits, without any detrimental influence on other qualitative parameters in rocket.

Glucosinolates are defense-related secondary metabolites found in Brassicaceae. When Brassicaceae come under attack, glucosinolates are hydrolyzed into different forms of glucosinolate hydrolysis products (GHPs). Among the GHPs, isothiocyanates are the most comprehensively characterized defensive compounds, whereas the functional study of nitriles, another group of GHP, is still limited. Therefore, this study investigates whether 3-butenenitrile (3BN), a nitrile, can trigger the signaling pathways involved in the regulation of defense responses in Arabidopsis thaliana against biotic stresses. Briefly, the methodology is divided into three stages, (i) evaluate the physiological and biochemical effects of exogenous 3BN treatment on Arabidopsis, (ii) determine the metabolites involved in 3BN-mediated defense responses in Arabidopsis, and (iii) assess whether a 3BN treatment can enhance the disease tolerance of Arabidopsis against necrotrophic pathogens. As a result, a 2.5 mM 3BN treatment caused lesion formation in Arabidopsis Columbia (Col-0) plants, a process found to be modulated by nitric oxide (NO). Metabolite profiling revealed an increased production of soluble sugars, Krebs cycle associated carboxylic acids and amino acids in Arabidopsis upon a 2.5 mM 3BN treatment, presumably via NO action. Primary metabolites such as sugars and amino acids are known to be crucial components in modulating plant defense responses. Furthermore, exposure to 2.0 mM 3BN treatment began to increase the production of salicylic acid (SA) and jasmonic acid (JA) phytohormones in Arabidopsis Col-0 plants in the absence of lesion formation. The production of SA and JA in nitrate reductase loss-of function mutant (nia1nia2) plants was also induced by the 3BN treatments, with a greater induction for JA. The SA concentration in nia1nia2 plants was lower than in Col-0 plants, confirming the previously reported role of NO in controlling SA production in Arabidopsis. A 2.0 mM 3BN treatment prior to pathogen assays effectively alleviated the leaf lesion symptom of Arabidopsis Col-0 plants caused by Pectobacterium carotovorum ssp. carotovorum and Botrytis cinerea and reduced the pathogen growth on leaves. The findings of this study demonstrate that 3BN can elicit defense response pathways in Arabidopsis, which potentially involves a coordinated crosstalk between NO and phytohormone signaling.

Glucosinolates (GLSs) are amino acid-derived defense compounds characteristic of the Brassicales order. Cytochromes P450s of the CYP79 family are the entry point into the biosynthetic pathway of the GLS core structure and catalyze the conversion of amino acids to oximes. In Arabidopsis thaliana, CYP79A2, CYP79B2, CYP79B3, CYP79F1, and CYP79F2 have been functionally characterized and are responsible for the biosynthesis of phenylalanine-, tryptophan-, and methionine-derived GLSs, respectively. However, the substrate(s) for CYP79C1 and CYP79C2 were unknown. Here, we investigated the function of CYP79C1 and CYP79C2 by transiently co-expressing the genes together with three sets of remaining genes required for GLS biosynthesis in Nicotiana benthamiana. Co-expression of CYP79C2 with either the aliphatic or aromatic core structure pathways resulted in the production of primarily leucine-derived 2-methylpropyl GLS and phenylalanine-derived benzyl GLS, along with minor amounts of GLSs from isoleucine, tryptophan, and tyrosine. Co-expression of CYP79C1 displayed minor amounts of GLSs from valine, leucine, isoleucine, and phenylalanine with the aliphatic core structure pathway, and similar GLS profile (except the GLS from valine) with the aromatic core structure pathway. Additionally, we co-expressed CYP79C1 and CYP79C2 with the chain elongation and aliphatic core structure pathways. With the chain elongation pathway, CYP79C2 still mainly produced 2-methylpropyl GLS derived from leucine, accompanied by GLSs derived from isoleucine and from chain-elongated mono- and dihomoleucine, but not from phenylalanine. However, co-expression of CYP79C1 only resulted in GLSs derived from chain-elongated amino acid substrates, dihomoleucine and dihomomethionine, when the chain elongation pathway was present. This shows that CYP79 activity depends on the specific pathways co-expressed and availability of amino acid precursors, and that description of GLS core structure pathways as “aliphatic” and “aromatic” pathways is not suitable, especially in an engineering context. This is the first characterization of members of the CYP79C family. Co-expression of CYP79 enzymes with engineered GLS pathways in N. benthamiana is a valuable tool for simultaneous testing of substrate specificity against multiple amino acids.

While the defensive function of glucosinolates is well established, their possible role as a nutrient reservoir is poorly understood and glucosinolate turnover pathways have not been elucidated. Previous research showed that glucosinolate content in germinating seeds of Arabidopsis thaliana Columbia-0 (Col-0) increases within the first two to four days on culture medium and then decreases below the level at day 0. In this study we used previously characterized T-DNA mutants to investigate if enzymes known to be involved in glucosinolate breakdown upon tissue damage affect the time course of glucosinolate content in germinating seeds. Besides dormant seeds, we analyzed seeds subjected to stratification in water for up to 72 h or germination on plates for up to ten days. Although seeds of tgg1 tgg2 (deficient in above-ground classical myrosinases) had higher glucosinolate levels than Col-0, the changes during germination were not different to those in seeds of Col-0. This demonstrates that TGG1/TGG2 are not responsible for the decline in glucosinolate content upon germination and suggests the involvement of other enzymes. Expression data extracted from publically available databases show a number of β-glucosidases of the BGLU18–BGLU33 clade to be expressed at specific time points of seed maturation and germination identifying them as good candidates for a role in glucosinolate turnover. Although nitrile-specifier proteins (NSPs) act downstream of myrosinases upon glucosinolate breakdown in tissue homogenates, mutants deficient in either seed-expressed NSP2 or seedling-expressed NSP1 were affected in glucosinolate content in seeds and during stratification or germination when compared to Col-0 indicating a direct role in turnover. The mutant lines nsp1-1, nsp2-1 and nsp2-2 had significantly higher glucosinolate levels in dry seeds than Col-0. After 24 h of stratification in water, nsp2-2 seeds contained 2.3 fold higher levels of glucosinolate than Col-0 seeds. This might indicate downregulation of hydrolytic enzymes when nitrile formation following glucosinolate hydrolysis is impaired. The time course of total glucosinolate content during ten days of germination depended on functional NSP1. Based on the present data, we propose a number of experiments that might aid in establishing the pathway(s) of glucosinolate turnover in germinating A. thaliana seeds.

Current research was conducted to explore the effects of liquiritoside on the growth and physiochemical features of Chinese flowering cabbage (Brassica rapa subsp. parachinensis) under lead (Pb) stress. Lead stressed B. rapa plants exhibited decreased growth parameters, chlorophyll, and carotenoid contents. Moreover, Pb toxicity escalated the synthesis of malondialdehyde (MDA), hydrogen peroxide (H2O2), flavonoids, phenolics, and proline in treated plants. Nevertheless, foliar application of liquiritoside mitigated Pb toxicity by decreasing oxidative stress by reducing cysteine, H2O2, and MDA contents in applied plants. Liquiritoside significantly increased plant height, shoot fresh weight and dry weight, number of leaves, and marketable value of Chinese flowering cabbage plants exposed to Pb toxicity. This biotic elicitor also enhanced the proline, glutathione, total phenolics, and flavonoid contents in Chinese flowering cabbage plants exposed to Pb stress compared with the control. Additionally, total glucosinolate content, phytochelatins (PCs), and non-protein thiols were effectively increased in plants grown under Pb regimes compared with the control plants. Overall, foliar application of liquiritoside can markedly alleviate Pb stress by restricting Pb translocation in Chinese flowering cabbage. [ABSTRACT FROM AUTHOR]

Halogeton glomeratus is a succulent annual herbaceous halophyte belonging to the Chenopodiaceae family, has attracted wide attention as a promising candidate for phytoremediation and as an oilseed crop and noodle-improver. More importantly, H. glomeratus has important medicinal value in traditional Chinese medicine. However, there are few comprehensive studies on the nutrients, particularly secondary metabolites. Here, we adopted untargeted metabolomics to compare the differences in metabolites of different tissues (root, stem, leaf, and seed) and identify the compounds related to pharmacological effects and response to abiotic stress in H. glomeratus. A total of 2,152 metabolites were identified, and the metabolic profiles of root, stem, leaf, and seed samples were clearly separated. More than 50% of the metabolites showed significant differences among root, stem, leaf, and seed. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of differential metabolites suggested an extensive alteration in the metabolome among the different organs. Furthermore, the identified metabolites related to pharmacological effects and response to abiotic stress included flavones, flavonols, flavandiols, glucosinolates, isoquinolines, pyridines, indoles, amino acids, lipids, carbohydrates, and ATP-binding cassette transporters. These metabolites have application in treating human cardiovascular diseases, cancers, diabetes, and heart disease, induce sleeping and have nutritive value. In plants, they are related to osmotic adjustment, alleviating cell damage, adjusting membrane lipid action and avoiding toxins. To the best of our knowledge, this is the first metabolomics-based report to overview the metabolite compounds in H. glomeratus and provide a reference for future development and utilization of H. glomeratus. [ABSTRACT FROM AUTHOR]

The biosynthesis of defensive secondary metabolites, such as glucosinolates (GSLs), is a costly process, which requires nutrients, ATP, and reduction equivalents, and, therefore, needs well-orchestrated machinery while coordinating defense and growth. We discovered that the key repressor of light signaling, the CONSTITUTIVE PHOTOMORPHOGENIC 1/SUPPRESSOR OF PHYTOCHROME A-105 (COP1/SPA) complex, is a crucial component of GSL biosynthesis regulation. Various mutants in this COP1/SPA complex exhibited a strongly reduced level of GSL and a low expression of jasmonate (JA)-dependent genes. Furthermore, cop1 , which is known to accumulate DELLA proteins in the dark, shows reduced gibberellin (GA) and JA signaling, thereby phenocopying other DELLA-accumulating mutants. This phenotype can be complemented by a dominant gain-of-function allele of MYC3 and by crossing with a mutant having low DELLA protein levels. Hence, SPA1 interacts with DELLA proteins in a yeast two-hybrid screen, whereas high levels of DELLA inhibit MYC function and suppress JA signaling. DELLA accumulation leads to reduced synthesis of GSL and inhibited growth. Thus, the COP1/SPA-mediated degradation of DELLA not only affects growth but also regulates the biosynthesis of GSLs. [ABSTRACT FROM AUTHOR]

Cruciferous plants in the order Brassicales defend themselves from herbivory using glucosinolates: sulfur-containing pro-toxic metabolites that are activated by hydrolysis to form compounds, such as isothiocyanates, which are toxic to insects and other organisms. Some herbivores are known to circumvent glucosinolate activation with glucosinolate sulfatases (GSSs), enzymes that convert glucosinolates into inactive desulfoglucosinolates. This strategy is a major glucosinolate detoxification pathway in a phloem-feeding insect, the silverleaf whitefly Bemisia tabaci , a serious agricultural pest of cruciferous vegetables. In this study, we identified and characterized an enzyme responsible for glucosinolate desulfation in the globally distributed B. tabaci species MEAM1. In in vitro assays, this sulfatase showed a clear preference for indolic glucosinolates compared with aliphatic glucosinolates, consistent with the greater representation of desulfated indolic glucosinolates in honeydew. B. tabaci might use this detoxification strategy specifically against indolic glucosinolates since plants may preferentially deploy indolic glucosinolates against phloem-feeding insects. In vivo silencing of the expression of the B. tabaci GSS gene via RNA interference led to lower levels of desulfoglucosinolates in honeydew. Our findings expand the knowledge on the biochemistry of glucosinolate detoxification in phloem-feeding insects and suggest how detoxification pathways might facilitate plant colonization in a generalist herbivore. [ABSTRACT FROM AUTHOR]

Myrosinase enzymes play a key role in the chemical defense of plants of the order Brassicales. Upon herbivory, myrosinases hydrolyze the β-S-linked glucose moiety of glucosinolates, the characteristic secondary metabolites of brassicaceous plants, which leads to the formation of different toxic hydrolysis products. The specialist flea beetle, Phyllotreta armoraciae , is capable of accumulating high levels of glucosinolates in the body and can thus at least partially avoid plant myrosinase activity. In feeding experiments with the myrosinase-deficient Arabidopsis thaliana tgg1 × tgg2 (tgg) mutant and the corresponding Arabidopsis Col-0 wild type, we investigated the influence of plant myrosinase activity on the metabolic fate of ingested glucosinolates in adult P. armoraciae beetles. Arabidopsis myrosinases hydrolyzed a fraction of ingested glucosinolates and thereby reduced the glucosinolate sequestration rate by up to 50% in adult beetles. These results show that P. armoraciae cannot fully prevent glucosinolate hydrolysis; however, the exposure of adult beetles to glucosinolate hydrolysis products had no impact on the beetle's energy budget under our experimental conditions. To understand how P. armoraciae can partially prevent glucosinolate hydrolysis, we analyzed the short-term fate of ingested glucosinolates and found them to be rapidly absorbed from the gut. In addition, we determined the fate of ingested Arabidopsis myrosinase enzymes in P. armoraciae. Although we detected Arabidopsis myrosinase protein in the feces, we found only traces of myrosinase activity, suggesting that P. armoraciae can inactivate plant myrosinases in the gut. Based on our findings, we propose that the ability to tolerate plant myrosinase activity and a fast glucosinolate uptake mechanism represent key adaptations of P. armoraciae to their brassicaceous host plants. [ABSTRACT FROM AUTHOR]

Alteration and induction of plant secondary metabolites after herbivore attack have been shown in almost all the studied plant species. Induction can be at the local site of damage, or systemic, such as from roots to shoots. In addition to immediate induction, previous herbivore bouts have been shown to “prime” the plants for a stronger and faster response only after a subsequent attack happens. Whereas several studies revealed a link between root herbivory and increased resistance against aboveground (AG) herbivory, the evidence of root defense priming against subsequent AG herbivory is currently lacking. To address this gap, we induced Cardamine hirsuta roots by applying jasmonic acid (JA), and, after a time lag, we subjected both control and JA-treated plants to AG herbivory by the generalist herbivore Spodoptera littoralis. We addressed the effect of root JA addition on AG herbivore resistance by measuring larval weight gain and tested the effect of root induction on abundance and composition of glucosinolates (GSLs) in shoots, prior, and after subsequent herbivory. We observed a strong positive effect of root induction on the resistance against AG herbivory. The overall abundance and identity of GSLs was globally affected by JA induction and by herbivore feeding, independently, and we found a significant correlation between larval growth and the shoot GSL profiles only after AG herbivory, 11 days after induction in roots. Contrary to expectations of priming, we observed that JA induction in roots altered the GSLs profile in the leaves that was maintained through time. This initial modification was sufficient to maintain a lower caterpillar weight gain, even 11 days post-root induction. Altogether, we show that prior root defense induction increases AG insect resistance by modifying and maintaining variation in GSL profiles during insect feeding.

Plants within the Brassicales order generate glucosinolate hydrolysis products that can exert different biological effects on several organisms. Here, we evaluated the physiological effects of one of these compounds, benzyl cyanide (phenylacetonitrile), when exogenously applied on Arabidopsis thaliana. Treatment with benzyl cyanide led to a dose-dependent reduction of primary root length and total biomass. Further morphological changes like elongated hypocotyls, epinastic cotyledons, and increased formation of adventitious roots resembled a severe auxin-overproducer phenotype. The elevated auxin response was confirmed by histochemical staining and gene expression analysis of auxin-responsive genes. Nitriles are converted by specific enzymes, nitrilases (NIT1-3), to their corresponding carboxylic acids. The nitrilase mutants nit1 and nit2 tolerated benzyl cyanide treatments better than the wild type, with nit2 being less resistant than nit1. A NIT2RNAi line suppressing several nitrilases was resistant to all tested benzyl cyanide concentrations. When exposed to phenylacetic acid (PAA) – the corresponding carboxylic acid of benzyl cyanide – wild type and mutant seedlings were, however, equally susceptible and showed a more severe auxin phenotype than upon cyanide treatment. Here, we demonstrate that the auxin-like effects triggered by benzyl cyanide on Arabidopsis are due to its nitrilase-mediated conversion to the natural auxin PAA.