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The interspecies communication roles of γ-butyrolactones (GBLs) have been described for a long time but are still poorly understood. Herein, we analyzed more than 1000 Streptomyces strains and noticed a big quantitative gap between the strains with GBL biosynthetic genes and the strains with GBL receptor genes, which implies the wide-spread of GBLs as interspecies signals in Streptomyces and their great potential in the activation of silent natural product gene clusters. Streptomyces albidoflavus J1074, which has one GBL receptor gene but no GBL biosynthetic gene, was chosen as a target to study the possible interspecies communication roles of GBLs. At first, the GBL biosynthetic genes from Streptomyces coelicolor M145 were expressed in S. albidoflavus J1074, which enabled the S. albidoflavus strains to synthesize Streptomyces coelicolor butanolides (SCBs) and activated the production of paulomycins. Further studies showed that this activation process requires the participation of the GBL receptor gene XNR_4681. The results suggest that the expression of exogenous GBL biosynthetic genes can modulate the metabolisms of GBL non-producing strains, and this regulation role might be meaningful for silent gene cluster activation in Streptomyces. At final, we synthesized racemic-SCB2 and tried to simplify the activation process by adding SCB2 directly to S. albidoflavus J1074, which unfortunately failed to induce paulomycin production.

Current understanding of the role of plant hormones during cereal grain filling is confounded by contradictory reports on hormone production that is based on poor methodology. We report here on the accurate measurement of indole-3-acetic acid (IAA) and abscisic acid (ABA) by combined liquid chromatography-tandem mass spectrometry in multiple reaction-monitoring mode with heavy isotope labelled internal standards. ABA and IAA contents of superior versus inferior rice grains (ABA maxima 159 ng g–1 FW and 109 ng g–1 FW, IAA maxima 2 µg g–1 FW and 1.7 µg g–1 FW respectively) correlated with the expression of biosynthetic genes and with grain fill. Results confirm that grain ABA is produced primarily by OsNCED2(5), but suggest that ABA import and metabolism also play important roles in ABA regulation. The IAA content of grains is primarily influenced by OsYUC9 and OsYUC11. However, the distinct expression profile of OsYUC12 suggests a specific role for IAA produced by this enzyme. Co-expression of OsYUC12 with OsIAA29 indicates their involvement in a common signalling pathway. Co-expression and cis-element analysis identified several aleurone-specific transcriptional regulators as well as glutelin as strong candidates for detailed investigation for direct regulation by the auxin-signalling pathway.

In actinomycetes, many secondary metabolite biosynthetic genes are not expressed under typical laboratory culture conditions and various efforts have been made to activate these dormant genes. In this study, we focused on high-temperature culture. First, we examined the thermotolerance of 3160 actinomycete strains from our laboratory culture collection and selected 57 thermotolerant actinomycetes that grew well at 45 °C. These 57 thermotolerant actinomycetes were cultured for 5 days in liquid medium at both 30 °C and 45 °C. Culture broths were extracted with 1-butanol, and each extract was subjected to LC/MS analysis. The metabolic profiles of each strain were compared between the 30 °C and 45 °C cultures. We found that almost half of these thermotolerant actinomycetes produced secondary metabolites that were detected only in the 45 °C culture. This result suggests that high-temperature culture induces the production of dormant secondary metabolites. These compounds were named “heat shock metabolites (HSMs).” To examine HSM production in more detail, 18 strains were selected at random from the initial 57 strains and cultivated in six different media at 30 °C and 45 °C; as before, metabolic profiles of each strain in each medium were compared between the 30 °C and 45 °C cultures. From this analysis, we found a total of 131 HSMs. We identified several angucycline-related compounds as HSMs from two thermotolerant Streptomyces species. Furthermore, we discovered a new compound, murecholamide, as an HSM from thermotolerant Streptomyces sp. AY2. We propose that high-temperature culture of actinomycetes is a convenient method for activating dormant secondary metabolite biosynthetic genes.

Background: Pueraria candollei var. mirifica , a Thai medicinal plant used traditionally as a rejuvenating herb, is known as a rich source of phytoestrogens, including isoflavonoids and the highly estrogenic miroestrol and deoxymiroestrol. Although these active constituents in P. candollei var. mirifica have been known for some time, actual knowledge regarding their biosynthetic genes remains unknown. Results: A de novo transcriptome analysis was conducted using combined P. candollei var. mirifica tissues of young leaves, mature leaves, tuberous cortices, and cortex-excised tubers.A total of 166,923 contigs was assembled for functional annotation using protein databases and as a library for identification of genes that are potentially involved in the biosynthesis of isoflavonoids and miroestrol. Twenty-one differentially expressed genes from four separate libraries were identified as candidatesinvolved in these biosyntheticpathways, and their respective expressions were validated by quantitative real-time reverse transcription polymerase chain reaction. Notably, isoflavonoid profiling generated by LC-MS/MS was positively correlated with expression levels of isoflavonoid biosynthetic genes across the four types of tissues. Moreover, we identified R2R3 MYB transcription factors that may be involved in the regulation of isoflavonoid biosynthesis in P. candollei var. mirifica . To confirm the function of a key-isoflavone biosynthetic gene, P. candollei var. mirifica isoflavone synthase identified in our library was transiently co-expressed with an Arabidopsis MYB12 transcription factor ( At MYB12) in Nicotiana benthamiana leaves. Remarkably, the combined expression of these proteins ledto the production of the isoflavone genistein. Conclusions: Our results provide compelling evidence regarding the integration of transcriptome and metabolome as a powerful tool for unraveling biosynthetic pathways in plants.

Covering: 2005 up to 2019Natural products are of paramount importance in human medicine. Not only are most antibacterial and anticancer drugs derived directly from or inspired by natural products, many other branches of medicine, such as immunology, neurology, and cardiology, have similarly benefited from natural product-based drugs. Typically, the genetic material required to synthesize a microbial specialized product is arranged in a multigene biosynthetic gene cluster (BGC), which codes for proteins associated with molecule construction, regulation, and transport. The ability to connect natural product compounds to BGCs and vice versa, along with ever-increasing knowledge of biosynthetic machineries, has spawned the field of genomics-guided natural product genome mining for the rational discovery of new chemical entities. One significant challenge in the field of natural product genome mining is how to rapidly link orphan biosynthetic genes to their associated chemical products. This review highlights state-of-the-art genetic platforms to identify, interrogate, and engineer BGCs from diverse microbial sources, which can be broken into three stages: (1) cloning and isolation of genomic loci, (2) heterologous expression in a host organism, and (3) genetic manipulation of cloned pathways. In the future, we envision natural product genome mining will be rapidly accelerated by de novo DNA synthesis and refactoring of whole biosynthetic pathways in combination with systematic heterologous expression methodologies.

New technology for the derivatization of peptide natural products is required for drug development. Despite the recent advances in the genome sequencing technique enabling us to search for the biosynthetic genes for wide variety of natural products, the technical methods to get access to them are limited. A class of RiPPs, a recently emerged natural product family such as thioviridamide, is one of those possessing such unexplored chemical space. In this paper, we report a streamlined method to generate new thioviridamide derivatives and to assess their biological activities. Heterologous expression of 42 constructs in an engineered Streptomyces avermitilis host gave 35 designed thioviridamide derivatives, along with several unprecedented analogues. Moreover, cytotoxicity assay revealed that several derivatives showed more potent activities than those of prethioviridamide. These results indicate that this strategy can become one of the potential ways to produce supreme unnatural products.

Microalgae are a promising biofuel resource, but their high cost and low productivity hinder their commercial applications. In the present study, Monoraphidium sp. QLZ-3 was cultivated in walnut shell extracts (WSE) supplemented with carbon dioxide (CO2). Biomass was enhanced from 0.40 g L−1 to 1.18 g L−1, and lipid content reached 49.54% in WSE-12% CO2 media. Biomass and lipid productivity reached 196.88 and 97.52 mg L−1 d−1, which were 1.33- and 1.57-fold higher than those of the control, respectively. The amount of carbohydrates increased, but the protein contents decreased. Furthermore, the application of CO2 promoted nutrient and polyphenol absorption and upregulated the expression levels of lipid biosynthetic genes of this WSE-cultivated alga. These results indicated that coupling WSE and CO2 could be an efficient strategy to enhance biofuel production by microalgae.

Ochratoxin A (OTA) is a potent pentaketide nephrotoxin widely contaminating food and feed products (grains, legumes, coffee, dried fruits, meat derived products, beer and wine). OTA is the primary mycotoxin risk in wine and dried vine fruits. The contamination of grapes and wine is strongly related to climatic conditions during cultivation. There is little information on the impact that climate change environmental fluxes may have on fungal colonization and ochratoxin A contamination of grapes. It has been suggested that the Mediterranean region is a hot spot for the impact of climate change scenarios with temperatures expected to increase by +2-5°C and CO2 to double or triple (400 ppm vs 800/1200 ppm) in the medium term. The northern region of Apulia, in the south of Italy, is normally considered to be an area where the risk of OTA in vineyards is relatively low. This study has examined the effect of i) increased temperature in the alternating day/night cycle (15-28°C vs 18-34°C), ii) existing and predicted CO2 concentrations (400 vs 1000 ppm), iii) conditions where water is freely available or drought stress is imposed (0.99 vs 0.93 aw ) in A. carbonarius strains (ITEM 7444, ITEM 5010 and B7), grown on grape-based matrices, by analysis of growth rate, OTA production and expression of key toxin biosynthetic genes. The results showed that in both non-stressed aw and drought stressed conditions the effect of elevated CO2 (1000 ppm) caused a general positive stimulation of growth rates at both the temperature cycles. Furthermore, while differences in the alternating day/night temperatures at 400 ppm and 0.99 aw did not result in any variation in OTA production, elevated CO2 condition (1000 ppm) was observed to stimulate OTA production especially at the 15-28°C cycle. These results were confirmed by gene expression analysis showing higher expression levels of OTA related genes at higher CO2 concentration. This study showed for the first time that elevated CO2 concentrations under different fluxes in day/night temperatures in the Mediterranean region may result in an increasing of OTA risk contamination in the production chain, being grapes destined for wine production less resilient under climate change scenarios.

Little is known on the impact that climate change (CC) may have on Aspergillus carbonarius and Ochratoxin A (OTA) contamination of grapes, especially in the Mediterranean region - a hot spot for the impact of CC scenarios with temperature expected to increase by +2-5°C and CO2 to double or triple (400 vs 800/1200 ppm). This study examined the effect of (i) current and increased temperature in the alternating 11.5h dark/12.5h light cycle (15-28°C vs 18-34°C), representative of the North Apulia area, South Italy and (ii) existing and predicted CO2 concentrations (400 vs 1000 ppm), on growth, expression of biosynthetic genes (AcOTApks, AcOTAnrps, AcOTAhal, AcOTAp450, AcOTAbZIP) and regulatory genes of Velvet complex (laeA/veA/velB, "velvet complex") involved in OTA biosynthesis and OTA phenotypic production by three strains of A. carbonarius. The experiments made on a grape-based matrix showed that elevated CO2 resulted in a general stimulation of growth and OTA production. These results were supported by the up-regulation of both structural and regulatory genes involved in the OTA biosynthesis in elevated CO2 condition. Our work has shown for the first time that elevated CO2 concentration in the Mediterranean region may result in an increased risk of OTA contamination in the wine production chain.

Rediscovery of already known compounds is a major issue in microbial natural product drug discovery. In recent years, progress has been made in developing more efficient analytical approaches that quickly identify known compounds in a sample to minimise rediscovery. In parallel, whole genome sequencing of microorganisms has revealed their immense potential to produce secondary metabolites, yet the majority of biosynthetic genes remain silent under common laboratory culturing conditions. Therefore, increased research has focused on optimising culturing methods to activate the silent biosynthetic gene clusters. Co-cultivation of different microbial strains can activate biosynthetic gene clusters that remain silent under standard laboratory fermentations involving mono-cultures, hence, the technique has great potential for natural product drug discovery. However, innovative methods are still needed to evaluate the success of any co-cultured fermentation end-product. Here, the application of HSQC-TOCSY NMR spectra and subsequent PCoA to identify changes in the metabolite diversity induced through co-cultivation is described.

Carrot is considered one of the leading horticultural crops in the world in terms of its nutritional value, health benefits, and unique flavor based on its high content of carotenoids and volatile aroma compounds. Terpenes such as monoterpenes and sesquiterpenes represent some of the predominant volatile compounds that contribute to carrot aroma and flavor. Variation of terpene composition based on genotypic differences or environmental factors has significant effects on taste perception by consumers and, therefore, is a critical quality attribute for carrot breeders and growers. Surprisingly, little is known about the biosynthesis of volatile terpenes in carrots and the various enzymes involved in their formation. In this chapter, we provide an overview of volatile terpene sampling and terpene diversity in different D. carota genotypes. Facilitated by the recent elucidation of the carrot genome, we further describe and discuss latest findings of the function of genes and enzymes in the terpene synthase family involved in the biosynthesis of carrot terpene volatiles.

Covering: 1960s to end of August 2018 Plant glandular trichomes (GTs) are adaptive structures that are well known as "phytochemical factories" due to their impressive capacity to biosynthesize and store large quantities of specialized natural products. The natural products in GTs are chemically diverse and mostly function as defense chemicals, therefore GTs are frequently regarded as "the first defense line" of plants against biotic and abiotic stresses. More importantly, many GT natural products are commercially desirable, thanks to their significant biological activities, thus attracting extensive interest in their biosynthesis. Consequently, it is well known that plant GTs are not only important reservoirs of biologically active natural products but are also a valuable bank of novel biosynthetic genes and enzymes. The non-volatile or oxygenated natural products in plant GTs, which need longer biosynthetic pathways and more energy from the plants, are of particular interest due to their more extensive biological activities and high commercial value. This review mainly focuses on these non-volatile natural products in plant GTs, including their chemistry, biological activities and biosynthesis. The methods employed for investigating natural products and their biosynthesis in plant GTs are also comprehensively discussed.

Pasteurella multocida produces a capsule composed of different polysaccharides according to the capsular serotype (A, B, D, E, and F). Hyaluronic acid (HA) is a component of certain capsular types of this bacterium, especially capsular type A. Previously, 2 HA biosynthetic genes from a capsular type A strain were studied for the industrial-scale improvement of HA production. Molecular comparison of these genes across different capsular serotypes of P multocida has not been reported. This study aimed to compare 8 HA biosynthetic genes ( pgi, pgm, galU, hyaC, glmS, glmM, glmU, and hyaD) of 22 P multocida strains (A:B:D:F = 6:6:6:4) with those of other organisms using sequence and structural bioinformatics analyses. These 8 genes showed a high level of within-species similarity (98%-99%) compared with other organisms. Only the last gene of 4 strains with capsular type F (HN07, PM70, HNF01, and HNF02) significantly differed from those of other strains (82%). Analysis of amino acid patterns together with phylogenetic results showed that the HA biosynthetic genes of the type A were closely related within the group. The genes in the capsular type F strain were notably similar to those of the capsular type A strain. Protein structural analysis supported structural similarities of the encoded enzymes between the strains of capsular types A, B, D, and F, except for the Pgm, GlmS, GlmU, and HyaD proteins. Our bioinformatics analytic workflow proposed that variations observed within these genes could be useful for genetic engineering–based improvement of hyaluronic acid–producing enzymes.

Exocarp protects the fruit against external stresses by means of its special physical and biochemical properties. It is also a commercially important quality for fruits characterized with certain color and structure. The color of sand pear exocarp can be divided into three types of russet, semi-russet and green. The green of exocarp is formed by accumulation of chlorophyll in epidermal cells. Russeting is a disorder of the fruit skin that results from microscopic cracks caused by growth stresses and several additional factors and the subsequent formation of higher plasticity periderm membranes by the accumulation of suberin on the inner part of the cell wall of the outer epidermal cell layers. Genes and pathways that are specific to exocarp russet formation have been identified in sand pear and its genetically related apple. The cuticle biosynthetic genes were repressed while stress response genes and suberin deposition genes were enhanced underlying the exocarp russeting. One major ‘QTL’ associated with russet of exocarp was identified at the top of LG 8. However, despite these advances, important aspects of russet and semi-russet inheritance remain obscure. Central questions include whether russet and semi-russet are belong to one type of quantitative trait or not, and their genetic characters. These issues are reviewed. Greater emphasis on gene mapping and cloning, biochemical characterization of metabolic intermediates and putative enzymes identified to put together correct and detailed pathways will be required to solve these unknowns of the mechanisms underlying the exocarp russet and semi-russet formation.

Natural products are a group of bioactive structurally diverse chemicals produced by microorganisms and plants. These molecules and their derivatives have contributed to over a third of the therapeutic drugs produced in the last century. However, over the last few decades traditional drug discovery pipelines from natural products have become far less productive and far more expensive. One recent development with promise to combat this trend is the application of synthetic biology to therapeutic natural product biosynthesis. Synthetic biology is a young discipline with roots in systems biology, genetic engineering, and metabolic engineering. In this review, we discuss the use of synthetic biology to engineer improved yields of existing therapeutic natural products. We further describe the use of synthetic biology to combine and express natural product biosynthetic genes in unprecedented ways, and how this holds promise for opening up completely new avenues for drug discovery and production.

Terrestrial bacteria, especially actinomycetes, are known to be prolific producers of volatile compounds. We show here that bacteria from ocean sediments can also release complex bouquets of volatiles. The actinomycete Salinispora tropica produces cyclohexenyl compounds not previously known in nature, such as methyl cyclohex-2-ene-1-carboxylate (9), methyl 2-(cyclohex-2-en-1-yl)acetate (10), methyl (E/Z)-2-(cyclohex-2-en-1-ylidene)acetate (11/12), and related alcohols 8 and 13. These compounds were identified by GC/MS and confirmed by synthesis. In addition, rare spiroacetals, aromatic compounds, short-chain acids and esters, alcohols, and various cyclic compounds were produced by the bacteria. The biosynthesis of the cyclohexenyl compounds is closely coupled to that of cyclohexenylalanine (4), a building block of salinosporamide A, a proteasome inhibitor produced by S. tropica. Analysis of S. tropica strains that harbor knockouts of the salinosporamide biosynthetic genes salX and salD, coupled with feeding experiments, revealed that 3-(cyclohex-2-en-1-yl)-2-oxopropanoic acid (60) and 3-(cyclohex-2-en-1-ylidene)-2-oxopropanoic acid (isomers 61 and 62) are important intermediates in the biosynthesis of salinosporamide A, 4, and 8-13.

The role of brassinosteroids (BRs) in hyponastic growth induced by submergence was investigated in Arabidopsis thaliana. Under flooding conditions, exogenously applied BRs increased hyponastic growth of rosette leaves. This hyponastic growth was reduced in a BR insensitive mutant (bri1-5), while it was increased in a BR dominant mutant (bes1-D). Further, expression of hypoxia marker genes, HRE1 and HRE2, was elevated in submerged bes1-D. These results indicate that BRs exert a positive action on hyponastic growth of submerged Arabidopsis leaves. Expression of ethylene biosynthetic genes, such as ACS6, ACS8 and ACO1, which are up-regulated by submergence, was also activated by application of BRs and in bes1-D. The enhanced hyponastic growth in submerged bes1-D was significantly reduced by application of cobalt ion, suggesting that BRs control hyponastic growth via ethylene, which seems to be synthesized by ACO6 and ACO8 followed by ACO1 in submerged leaves. A double mutant, bes1-Dxaco1-1, showed hyponastic growth activity similar to that seen in aco1-1, demonstrating that the BR signaling for regulation of hyponastic growth seems to be an upstream event in ethylene-induced hyponastic growth under submergence in Arabidopsis.

Astragalosides (AGs) are triterpene saponins that uniquely occur in Astragalus species with multiple pharmacological activities. For the first time, we reported an elicitation strategy, using UV radiation (UV-A, UV-B, and UV-C), to enhance AG production in Astragalus membranaceus hairy root cultures (AMHRCs). The treatment of 32 day-old AMHRCs with 54 kJ/m 2 dose of UV-B radiation was capable of achieving the maximum production of AGs, i.e. 3.431 ± 0.092 mg/g dry weight (DW), which was 1.30-fold higher as compared to the level in non-treated control (2.645 ± 0.073 mg/g DW). Expressions of eleven genes involved in AG biosynthetic pathway were monitored by quantitative real-time PCR (qRT-PCR). Transcriptional results showed that only the HMGR gene was significantly up-regulated in response to UV-B elicitation. The present study demonstrated that UV-B elicitation is a feasible strategy to promote AG production in AMHRCs and that HMGR represents a potential key gene in AG biosynthetic pathway.

The influence of different illumination levels of blue light on the growth and intracellular pigment yields of Monascus strain M9 was investigated. Compared with darkness, constant exposure to blue light of 100 lux reduced the yields of six pigments, namely, rubropunctatamine (RUM), monascorubramine (MOM), rubropunctatin (RUN), monascorubrin (MON), monascin (MS), and ankaflavin (AK). However, exposure to varying levels of blue light had different effects on pigment production. Exposure to 100 lux of blue light once for 30 min/day and to 100 lux of blue light once and twice for 15 min/day could enhance RUM, MOM, MS, and AK production and reduce RUN and MON compared with non-exposure. Exposure to 100 lux twice for 30 min/day and to 200 lux once for 45 min/day decreased the RUM, MOM, MS, and AK yields and increased the RUN and MON. Meanwhile, the expression levels of pigment biosynthetic genes were analyzed by real-time quantitative PCR. Results indicated that gene MpPKS5, mppR1, mppA, mppB, mmpC, mppD, MpFasA, MpFasB, and mppF were positively correlated with the yields of RUN and MON, whereas mppE and mppR2 were associated with RUM, MOM, MS, and AK production.

Light quality and intensity have severe impact on Steroidal glycoalkaloid (SGA) biosynthesis, causing the quality degradation of potato by greening and accelerating SGA-induced toxicity. In this study, Atlantic and Haryoung cultivars responded differently when exposed to seven light qualities viz., purple, red, blue, green, yellow, UV and fluorescent lights. Haryoung tuber exhibited a 44 % lesser SGA accumulation in yellow light compared to fluorescent light. Further screening of selected light sources by investigating the expression pattern of key genes and SGA contents, showed lower transcript levels of key genes in yellow light which in-turn reflected in the least amount of SGA accumulation with a 22 % lesser SGA levels in yellow light compared to fluorescent light. This suppression in the SGA accumulation suggests that yellow light can be used as an alternative light source instead of fluorescent light which would help in minimizing the quality degradation and SGA- induced toxicity.

Covering: 2006 to 2016 The computational mining of genomes has become an important part in the discovery of novel natural products as drug leads. Thousands of bacterial genome sequences are publically available these days containing an even larger number and diversity of secondary metabolite gene clusters that await linkage to their encoded natural products. With the development of high-throughput sequencing methods and the wealth of DNA data available, a variety of genome mining methods and tools have been developed to guide discovery and characterisation of these compounds. This article reviews the development of these computational approaches during the last decade and shows how the revolution of next generation sequencing methods has led to an evolution of various genome mining approaches, techniques and tools. After a short introduction and brief overview of important milestones, this article will focus on the different approaches of mining genomes for secondary metabolites, from detecting biosynthetic genes to resistance based methods and “evo-mining” strategies including a short evaluation of the impact of the development of genome mining methods and tools on the field of natural products and microbial ecology.

Amaryllidaceae alkaloids are an example of the vast diversity of secondary metabolites with great therapeutic promise. The identification of novel compounds in this group with over 300 known structures continues to be an area of active study. The recent identification of norbelladine 4′-O-methyltransferase (N4OMT), an Amaryllidaceae alkaloid biosynthetic enzyme, and the assembly of transcriptomes for Narcissus sp. aff. pseudonarcissus and Lycoris aurea highlight the potential for discovery of Amaryllidaceae alkaloid biosynthetic genes with new technologies. Recent technical advances of interest include those in enzymology, next generation sequencing, genetic modification, nuclear magnetic resonance spectroscopy (NMR), and mass spectrometry (MS).

The involvement of ethylene gas in the ripening of peach fruit is a symbolic phenomenon that has a place in the history of research on plant hormones. It is well documented that fruit ripening is developmentally regulated and affected by environmental factors such as temperature and light. Blue light has been shown to induce pigment synthesis in postharvest fruit. However, information on its effect on fruit ripening is still limited. This study aimed to investigate the effect of light-emitting diode blue light treatment at 40 μmol m−2 s−1 on fruit ripening, ethylene biosynthesis and signalling in postharvest peaches during storage at 10 °C. Blue light exposure induced fruit ripening as expressed by the enhanced decline of fruit firmness and increases in skin colour parameters, which was associated with the increase in ethylene production because of the up-regulated expression of PpLOXs and ethylene biosynthetic genes such as PpACO1 and PpACS3. Meanwhile, six genes in relation to ethylene perception and signalling were also investigated using real-time PCR. Among them, blue light treatment significantly inhibited expression of PpERS1 and enhanced transcript levels of PpEIN2, PpEIL1, PpEIL2 and PpERF2, which led to an induction ethylene signalling pathway and resulted in turn into the promotion of ethylene production and fruit softening in postharvest peaches. Compared to control fruit stored at darkness, higher expression level of ethylene biosynthetic and signalling genes in blue light exposed peaches could be beneficial for its ethylene production and consequently fruit ripening.

Apples with red fruit flesh are desirable for breeding because they contain more health-beneficial compounds. That color trait is also considered more attractive to consumer. To obtain experimental evidence about the molecular mechanism for this pigmentation, we investigated how the practice of bagging fruit might affect levels of phenolic compounds as well as the expression of anthocyanin biosynthetic and regulatory genes. Two red-fleshed varieties from Xinjiang, P.R. China, were studied – ‘Xiahongrou’ and ‘No.1 Hongxun’. HPLC analysis showed that the contents of anthocyanins in both peel and flesh were decreased by bagging, but were increased again when fruits were re-exposed to sunlight. Except for F3H in ‘No.1 Hongxun’, transcription levels for most anthocyanin biosynthetic genes in the flesh were enhanced after bag removal. MYB10 is a key transcriptional factor in the anthocyanin pathway of these two apple varieties, particularly in the flesh. These results suggest that the development of red coloring when bags are removed could be a result of light-induced coordinated expression by MYB10 and anthocyanin biosynthetic genes in red-fleshed apples.

Gene elongation is a molecular mechanism consisting of an in-tandem duplication of a gene and divergence and fusion of the two copies, resulting in a gene constituted by two divergent paralogous modules. The aim of this work was to evaluate the importance of gene elongation in the evolution of histidine biosynthetic genes and to propose a possible evolutionary model for some of them. Concerning the genes hisA and hisF, which code for two homologous (&beta, /&alpha, )8-barrels, it has been proposed that the two extant genes could be the result of a cascade of gene elongation/domain shuffling events starting from an ancestor gene coding for just one (&beta, ) module. A gene elongation event has also been proposed for the evolution of hisB and hisD, structural analyses revealed the possibility of an early elongation event, resulting in the repetition of modules. Furthermore, it is quite possible that the gene elongations responsible for the evolution of the four proteins occurred before the earliest phylogenetic divergence. In conclusion, gene elongation events seem to have played a crucial role in the evolution of the histidine biosynthetic pathway, and they may have shaped the structures of many genes during the first steps of their evolution.

Anthocyanin biosynthesis exhibits a rhythmic oscillation pattern in some plants. To investigate the correlation between the oscillatory regulatory network and anthocyanin biosynthesis in pear, the anthocyanin accumulation and the expression patterns of anthocyanin late biosynthetic genes (ALBGs) were investigated in fruit skin of &lsquo, Red Zaosu&rsquo, (Pyrus bretschneideri Rehd.). The anthocyanin accumulated mainly during the night over three continuous days in the fruit skin, and the ALBGs&rsquo, expression patterns in &lsquo, fruit skin were oscillatory. However, the expression levels of typical anthocyanin-related transcription factors did not follow this pattern. Here, we found that the expression patterns of four PbREVEILLEs (PbRVEs), members of a class of atypical anthocyanin-regulated MYBs, were consistent with those of ALBGs in &lsquo, fruit skin over three continuous days. Additionally, transient expression assays indicated that the four PbRVEs promoted anthocyanin biosynthesis by regulating the expression of the anthocyanin biosynthetic genes encoding dihydroflavonol-4-reductase (DFR) and anthocyanidin synthase (ANS) in red pear fruit skin, which was verified using a dual-luciferase reporter assay. Moreover, a yeast one-hybrid assay indicated that PbRVE1a, 1b and 7 directly bound to PbDFR and PbANS promoters. Thus, PbRVEs promote anthocyanin accumulation at night by up-regulating the expression levels of PbDFR and PbANS in &lsquo, fruit skin.

Root-tubers of cassava (Manihot esculenta Crantz) accumulate high starchy carbohydrates, protein, and carotenoids. Carotenoids were associated with tuber colorus, i.e., yellow-fleshed tuber accumulates more carotenoids than white-fleshed root-tubers. Carotenoids serve nutrition for health, such as an antioxidant and provitamin A. This study aimed to analyze relative expressions (RE) of carotenoid genes in Indonesian yellow and white cassava (IYC and IWC, respectively). Nine-month-old fresh root-tubers of IYC (Adira-1, Mentega-2, and Ubi Kuning) and IWC (Adira-4 and Menti) were used for the analysis. RE of five genes: PSY (phytoene synthase), CRTISO (carotenoid isomerase), LYCa (α-lycopene cyclase), LYCb (β-lycopene cyclase), and BCH1 (beta carotene hydroxylase1), and a reference gene, i.e., EF1-A (elongation factor 1-alpha), were analyzed using a qRT-PCR. Results showed that RE of all genes were detected in IYC and IWC. In IWC, all genes were expressed high, suggesting biosynthetic fluxes through yellow α-carotenoids and xanthophylls. In YWC, variations on RE levels of all genes were observed. LYCa and LYCb were expressed high in Adira-1 and Mentega-2, indicating α- and β-carotenoids accumulate in root-tubers. However, BCH1 was expressed 2-3-folds lower in both accessions than all IWC, suggesting low zeaxanthin production and accumulation of β-carotene. In Ubi Kuning, α-carotenoids accumulation may not exist due to low LYCa RE. Also, higher BCH1 expression levels were observed in Ubi Kuning, suggesting a substantial zeaxanthin accumulation. Results are potential for improving carotenoid accumulation through breeding programs in cassava.

Sacred lotus (Nelumbo nucifera Gaertn.) is an ancient aquatic plant used throughout Asia for its nutritional and medicinal properties. Benzylisoquinoline alkaloids (BIAs), mostly within the aporphine and bisbenzylisoquinoline structural categories, are among the main bioactive constituents in the plant. The alkaloids of sacred lotus exhibit promising anti-cancer, anti-arrhythmic, anti-HIV, and anti-malarial properties. Despite their pharmacological significance, BIA metabolism in this non-model plant has not been extensively investigated. In this review, we examine the diversity of BIAs in sacred lotus, with an emphasis on the distinctive stereochemistry of alkaloids found in this species. Additionally, we discuss our current understanding of the biosynthetic genes and enzymes involved in the formation of 1-benzylisoquinoline, aporphine, and bisbenzylisoquinoline alkaloids in the plant. We conclude that a comprehensive functional characterization of alkaloid biosynthetic enzymes using both in vitro and in vivo methods is required to advance our limited knowledge of BIA metabolism in the sacred lotus.

The methylated flavone tricin has been associated with numerous health benefits, including reductions in intestinal and colon cancers in animal models. Tricin is found in a wide range of plant species and in many different tissues. However, whole cereal grains, such as rice, barley, oats, and wheat, are the only food sources of tricin, which is located in the bran portion of the grain. Variation in tricin levels was found in bran from rice genotypes with light brown, brown, red, and purple pericarp color, with the purple pericarp genotypes having the highest levels of tricin. Here, we analyzed tricin and tricin derivative levels in developing pericarp and embryo samples of a purple pericarp genotype, IAC600, that had high tricin and tricin derivative levels in the bran, and a light brown pericarp genotype, Cocodrie, that had no detectable tricin or tricin derivatives in the bran. Tricin and tricin derivatives were detected in both the pericarp and embryo of IAC600 but only in the embryo of Cocodrie. The purple pericarp rice had higher total levels of free tricin plus tricin derivatives than the light brown pericarp rice. When expressed on a per grain basis, most of the tricin component of IAC600 was in the pericarp. In contrast, Cocodrie had no detectable tricin in the pericarp samples but did have detectable chrysoeriol, a precursor of tricin, in the pericarp samples. We also used RNA-Seq analysis of developing pericarp and embryo samples of the two cultivars to compare the expression of genes involved in the flavonoid biosynthetic pathway. The results presented here suggest that understanding the basis of tricin accumulation in rice pericarp may lead to an approach to increasing tricin levels in whole grain rice. From analysis of gene expression levels in the pericarp samples it appears that regulation of the flavone specific genes is independent of regulation of the anthocyanin biosynthetic genes. It therefore may be feasible to develop brown pericarp rice cultivars that accumulate tricin in the pericarp.

The advances of genomic sequence analyses and genome mining tools have enabled the exploration of untapped microbial natural products. Through genome mining studies to discover cryptic natural products, we found biosynthetic genes encoding a new lasso peptide in the genome sequence of a soil bacterium, Streptomyces sp. KCB13F003 isolated from Ulleung Island (a small volcanic island), Korea. The production and purification of the encoded peptide, named ulleungdin, were achieved by optimizing the culture conditions followed by LC-MS-targeted isolation. Structure elucidation was performed by NMR spectroscopic and MS spectrometric analyses and chemical means (Marfey's and GITC derivatizations), proving ulleungdin to be a new 15-mer class II lasso peptide with a threaded structure. Biological evaluation with the cell invasion assay and time-lapse cell tracking analysis revealed that ulleungdin has significant inhibitory activities against cancer cell invasion and migration.

The Aspergillus niger aggregate contains 15 morphologically indistinguishable species which presence is related to ochratoxin A (OTA) and fumonisin B2 (FB2) contamination of foodstuffs. The taxonomy of this group was recently reevaluated and there is a need of new studies regarding the risk that these species might pose to food security. 258 isolates of A. niger aggregate obtained from a variety of products from Spain were classified by molecular methods being A. tubingensis the most frequently occurring (67.5%) followed by A. welwitschiae (19.4%) and A. niger (11.7%). Their potential ability to produce mycotoxins was evaluated by PCR protocols which allow a rapid detection of OTA and FB2 biosynthetic genes in their genomes. OTA production is not widespread in A. niger aggregate since only 17% of A. niger and 6% of A. welwitschiae isolates presented the complete biosynthetic cluster whereas the lack of the cluster was confirmed in all A. tubingensis isolates. On the other hand, A. niger and A. welwitschiae seem to be important FB2 producers with 97% and 29% of the isolates, respectively, presenting the complete cluster. The genes involved in OTA and FB2 were overexpressed in producing isolates and their expression was related to mycotoxin synthesis.

The accumulation of beneficial biochemical compounds in different parts of pomegranate ( Punica granatum L.) fruit determines fruit quality and highly depends on environmental conditions. We investigated the effects of agro-climatic conditions on major biochemical compounds and on the expression of major anthocyanin biosynthetic genes in the peels and arils of a yellow-peeled and pink-ariled pomegranate cultivar in three agro-climatologically different locations in Sri Lanka. Drier and warmer climates promoted the accumulation of the measured biochemical compounds, i.e. total phenolic content (TPC), antioxidant capacity (AOX), and α, β, and total punicalagin, in both peels and arils compared to wetter and cooler climates. Pomegranate DFR, F3H, and ANS transcripts in both peels and arils showed higher relative expression in hotter and drier regions, compared to those grown in cooler and wetter conditions. Therefore, growing pomegranates in drier and warmer environments maximizes the production of beneficial biochemical compounds and associated gene expression in pomegranate fruit.

Covering: up to 2018 Ovothiols are sulfur-containing natural products biosynthesized by marine invertebrates, microalgae, and bacteria. These compounds are characterized by unique chemical properties suggestive of numerous cellular functions. For example, ovothiols may be cytoprotectants against oxidative stress, serve as building blocks of more complex structures and may act as molecular messengers for inter- and intracellular signaling. Detailed understanding of ovothiol physiological role in marine organisms may unearth novel concepts in cellular redox biochemistry and highlight the therapeutic potential of this antioxidant. The recent discovery of ovothiol biosynthetic genes has paved the way for a systematic investigation of ovothiol-modulated cellular processes. In this highlight we review the early research on ovothiol and we discuss key questions that may now be addressed using genome-based approaches. This highlight article provides an overview of recent progress towards elucidating the biosynthesis, function and potential application of ovothiols.

Pakchoi (Brassica rapa subsp. chinensis) is cultivated for its nutritional value, particularly with regard to vitamins, minerals and dietary fibers. However, limited metabolic information is available on the phyto-nutritional traits of pakchoi. Our GC-TOF MS analysis showed that green pakchoi has higher contents of carbon metabolism-associated metabolites such as sugars, sugar derivatives and inositol, while purple pakchoi has higher levels of nitrogen metabolism-associated metabolites such as amino acids and amino acid derivatives. To compare the content and composition of secondary metabolites in green and purple pakchoi, we analyzed phenylpropanoid-derived compounds and anthocyanins in mature leaves using an HPLC-UV system. This analysis identified 9 phenylpropanoid-derived compounds and 12 anthocyanins in the mature leaves of green and purple pakchoi. The level of rutin was significantly higher in purple pakchoi compared with green pakchoi, consistent with the expression of phenylpropanoid biosynthetic genes in the two pakchoi cultivars. The data obtained from this comprehensive metabolic profiling would be helpful to improve our understanding of the nutritional values of pakchoi cultivars as food sources.

The emerging and reemerging forms of fungal infections encountered in the course of allogeneic bone marrow transplantations, cancer therapy, and organ transplants have necessitated the discovery of antifungal compounds with enhanced efficacy and better compatibility. A very limited number of antifungal compounds are in practice against the various forms of topical and systemic fungal infections. The trends of new antifungals being introduced into the market have remained insignificant while resistance towards the introduced drug has apparently increased, specifically in patients undergoing long-term treatment. Considering the immense potential of natural microbial products for the isolation and screening of novel antibiotics for different pharmaceutical applications as an alternative source has remained largely unexplored. Endophytes are one such microbial community that resides inside all plants without showing any symptoms with the promise of producing diverse bioactive molecules and novel metabolites which have application in medicine, agriculture, and industrial set ups. This review substantially covers the antifungal compounds, including volatile organic compounds, isolated from fungal endophytes of medicinal plants during 2013–2018. Some of the methods for the activation of silent biosynthetic genes are also covered. As such, the compounds described here possess diverse configurations which can be a step towards the development of new antifungal agents directly or precursor molecules after the required modification.

Tebuconazole, which consists of a pair of enantiomers with different fungicidal activities, is one of the most common fungicides used in the control of Fusarium graminearum. In this study, the fungicidal activity of rac-tebuconazole and its enantiomers against F. graminearum was determined at 0.997, 0.975, and 0.950 aw and at 20, 25, and 30 °C on wheat-based media. Then, F. graminearum was treated with rac-tebuconazole and its enantiomers at the EC10, EC50, and EC90 levels under different culture conditions, and DON production was measured. Finally, expression of the DON biosynthetic genes (TRI5 and TRI6) was quantified by real-time RT-PCR after incubation with EC50 doses of rac-tebuconazole and its enantiomers for 4, 8, and 14 days at 30 °C and aw 0.997. The results showed that the fungicidal activity of tebuconazole was strongly influenced by temperature, aw, and the combined factors. (−)-Tebuconazole is higher in fungicidal activity than (+)-tebuconazole and rac-tebuconazole with 24–99-fold and 1.8–6.7...

Seed germination plays important roles in the establishment of seedlings and their subsequent growth; however, seed germination is inhibited by salinity, and the inhibitory mechanism remains elusive. Our results indicate that NaCl treatment inhibits rice seed germination by decreasing the contents of bioactive gibberellins (GAs), such as GA1 and GA4, and that this inhibition can be rescued by exogenous bioactive GA application. To explore the mechanism of bioactive GA deficiency, the effect of NaCl on GA metabolic gene expression was investigated, revealing that expression of both GA biosynthetic genes and GA-inactivated genes was up-regulated by NaCl treatment. These results suggest that NaCl-induced bioactive GA deficiency is caused by up-regulated expression of GA-inactivated genes, and the up-regulated expression of GA biosynthetic genes might be a consequence of negative feedback regulation of the bioactive GA deficiency. Moreover, we provide evidence that NaCl-induced bioactive GA deficiency inhibits rice seed germination by decreasing α-amylase activity via down-regulation of α-amylase gene expression. Additionally, exogenous bioactive GA rescues NaCl-inhibited seed germination by enhancing α-amylase activity. Thus, NaCl treatment reduces bioactive GA content through promotion of bioactive GA inactivation, which in turn inhibits rice seed germination by decreasing α-amylase activity via down-regulation of α-amylase gene expression.

Anthocyanins are the resultant end-point metabolites of phenylapropanoid/flavonoid (F/P) pathway which is regulated at transcriptional level via a series of structural genes. Identifying the key genes and their potential interactions can provide us with the clue for novel points of intervention for improvement of the trait in strawberry. We profiled the expressions of putative regulatory and biosynthetic genes of cultivated strawberry in three developmental and characteristically colored stages of fruits of contrastingly anthocyanin rich cultivars: Tokun, Maehyang and Soelhyang. Besides FaMYB10, a well-characterized positive regulator, FaMYB5, FabHLH3 and FabHLH3-delta might also act as potential positive regulators, while FaMYB11, FaMYB9, FabHLH33 and FaWD44-1 as potential negative regulators of anthocyanin biosynthesis in these high-anthocyanin cultivars. Among the early BGs, Fa4CL7, FaF3H, FaCHI1, FaCHI3, and FaCHS, and among the late BGs, FaDFR4-3, FaLDOX, and FaUFGT2 showed significantly higher expression in ripe fruits of high anthocyanin cultivars Maehyang and Soelhyang. Multivariate analysis revealed the association of these genes with total anthocyanins. Increasingly higher expressions of the key genes along the pathway indicates the progressive intensification of pathway flux leading to final higher accumulation of anthocyanins. Identification of these key genetic determinants of anthocyanin regulation and biosynthesis in Korean cultivars will be helpful in designing crop improvement programs.

Plants synthesize volatile compounds to attract pollinators. The volatiles emitted by flowers are often complex mixtures of organic compounds; pollinators are capable of distinctly recognizing different volatile compounds. Plants also produce volatile compounds to protect themselves against herbivores and pathogens. Some of the volatile compounds produced in floral and vegetative tissues are toxic to insects and microbes. To adapt changes in the environment, plants have evolved the ability to synthesize a unique set of volatiles. Intensive studies have identified and characterized the enzymes responsible for the formation of plant volatiles. In particular, many biosynthetic genes have been isolated and their enzymatic functions have been proposed. This review describes how plants have evolved the biosynthetic pathways leading to the formation of green leaf volatiles and phenylpropene volatiles.

We develop a novel synthetic biology platform for rapid, scalable expression of fungal biosynthetic genes and encoded metabolites., For decades, fungi have been a source of U.S. Food and Drug Administration–approved natural products such as penicillin, cyclosporine, and the statins. Recent breakthroughs in DNA sequencing suggest that millions of fungal species exist on Earth, with each genome encoding pathways capable of generating as many as dozens of natural products. However, the majority of encoded molecules are difficult or impossible to access because the organisms are uncultivable or the genes are transcriptionally silent. To overcome this bottleneck in natural product discovery, we developed the HEx (Heterologous EXpression) synthetic biology platform for rapid, scalable expression of fungal biosynthetic genes and their encoded metabolites in Saccharomyces cerevisiae. We applied this platform to 41 fungal biosynthetic gene clusters from diverse fungal species from around the world, 22 of which produced detectable compounds. These included novel compounds with unexpected biosynthetic origins, particularly from poorly studied species. This result establishes the HEx platform for rapid discovery of natural products from any fungal species, even those that are uncultivable, and opens the door to discovery of the next generation of natural products.

Actinobacteria are important sources for discovery of secondary metabolites for application in medicine and agriculture. Genome mining methods employ combinations of molecular biology techniques, including next-generation sequencing, and bioinformatics-based methods reinvigorating interest in the biosynthetic potential of gene clusters in actinobacteria, opening new avenues for the discovery of natural bioactive products. Notwithstanding the recent advancements in genomics, the field of natural products research is currently undergoing a paradigm shift. However, recent insights gained from genomics revealed a massive existence of previously unrecognized biosynthetic potential in actinobacteria. Interestingly genomics-guided natural product discovery is considered a feasible approach for the discovery of novel and bioactive secondary metabolites from the actinobacteria. In this review an attempt is made to highlight novel perspectives in natural product genomics, and analyze the strengths and limitations of the genomics technology, where elucidation of certain aspects could lead to noteworthy biotechnological applications.

We evaluated the impact of increased temperature and sunlight exposure, features of global warming, on the skin coloration of ‘Kyoho’ table grape berries. Skin coloration, anthocyanin accumulation, abscisic acid (ABA) content, and transcript levels of anthocyanin biosynthetic genes in the skins were examined during the ripening season under continuous 24 °C, 27 °C and 30 °C temperatures together with a sun or shade treatment. Skin coloration and anthocyanin accumulation were sufficient in the berries of plants ripened at 24 °C with a sun-exposure treatment. They also had higher ABA contents and anthocyanin biosynthetic gene transcript levels than grapes under higher temperatures. The skins of grapes ripened at 27 °C or 30 °C showed insufficient coloration and low levels of anthocyanin accumulation, ABA content, and anthocyanin biosynthetic gene transcripts levels, irrespective of the light conditions. These results suggest that temperatures exceeding 27 °C during the ripening season lead to insufficient berry coloration as a result of low levels of ABA and anthocyanin biosynthetic gene expression levels.