Production Of Specific Metabolites Research Articles

Application of 1H HR-MAS NMR-Based Metabolite Fingerprinting of Marine Microalgae

Natural products from the marine environment as well as microalgae, have been known for the complexity of the metabolites they produce due to their adaptability to different environmental conditions, which has been an inexhaustible source of several bioactive properties, such as antioxidant, anti-tumor, and antimicrobial. This study aims to characterize the main metabolites of three species of microalgae (Nannochloropsis oceanica, Chaetoceros muelleri, and Conticribra weissflogii), which have important applications in the biofuel and nutrition industries, by 1H High-resolution magic angle spinning nuclear magnetic resonance (1H HR-MAS NMR), a method which is non-destructive, is highly reproducible, and requires minimal sample preparation. Even though the three species were found in the same ecosystem and a superior production of lipid compounds was observed, important differences were identified in relation to the production of specialized metabolites. These distinct properties favor the use of these compounds as leaders in the development of new bioactive compounds, especially against environmental, human, and animal pathogens (One Health), and demonstrate their potential in the development of alternatives for aquaculture.

Improvement of phenolic compounds production in callus cultures of Cereus hildmannianus (K.) Schum. by elicitation

Cereus hildmannianus (Cactaceae) is a medicinal plant with various medicinal and industrial applications. Plant biotechnology is an attractive approach for specialized metabolite production under controlled conditions, with the elicitation process being considered a highly effective strategy for increasing the production of bioactive compounds. In this regard, the present study investigated the effects of elicitors, sucrose (S, 1–8 g L−1), UV-C light (UV-C, 10–80 min), salicylic acid (SA, 50–200 µmol), and jasmonic acid (JA, 50–200 µmol), on the profile of esterase isozymes, total phenolic and total flavonoid contents, antioxidant activity (FRAP, DPPH, and ABTS), and the dereplication method by Ultra-High-Performance Liquid Chromatography coupled with Mass Spectrometry (UHPLC-ESI-Q-TOF–MS/MS). The isozyme profile was not significantly changed. The highest accumulations of total phenolics were observed in callus tissues induced with 100 µmol JA (390.8 µg GAE mg−1 DW), 200 µmol SA (355.5 µg GAE mg−1 DW), 20 min UV-C (182.9 µg GAE mg−1 DW), and 6 g L−1 S (122.1 µg GAE mg−1 DW); the highest concentrations of total flavonoids were observed following culture with 200 µmol SA (89.8 µg QE mg−1 DW) and 2 g L−1 S (28.4 µg QE mg−1 DW). Furthermore, the dereplication study allowed putative identification of thirty-six compounds (thirty phenolic acids and organic acids, and six flavonoids). The highest antioxidant activities were obtained with callus tissues induced with 100 µmol JA, 200 µmol SA, 80 min UV-C, and 1 g L−1 S. The elicitors were able to alter the production of phenolic compounds in callus cultures of C. hildmannianus. These results are promising for the clean and sustainable bioproduction of bioactive molecules for pharmaceutical and cosmeceutical purposes, prioritizing the conservation of the species. Elicitation of callus tissues of Cereus hildmannianus with sucrose, UV-C light, salicylic acid, and jasmonic acid enhances the phenolic compound content with antioxidant activity, which is known to be associated with promising antiaging activity.

Lunaemycins, New Cyclic Hexapeptide Antibiotics from the Cave Moonmilk-Dweller Streptomyces lunaelactis MM109T

Streptomyces lunaelactis strains have been isolated from moonmilk deposits, which are calcium carbonate speleothems used for centuries in traditional medicine for their antimicrobial properties. Genome mining revealed that these strains are a remarkable example of a Streptomyces species with huge heterogeneity regarding their content in biosynthetic gene clusters (BGCs) for specialized metabolite production. BGC 28a is one of the cryptic BGCs that is only carried by a subgroup of S. lunaelactis strains for which in silico analysis predicted the production of nonribosomal peptide antibiotics containing the non-proteogenic amino acid piperazic acid (Piz). Comparative metabolomics of culture extracts of S. lunaelactis strains either holding or not holding BGC 28a combined with MS/MS-guided peptidogenomics and 1H/13C NMR allowed us to identify the cyclic hexapeptide with the amino acid sequence (D-Phe)-(L-HO-Ile)-(D-Piz)-(L-Piz)-(D-Piz)-(L-Piz), called lunaemycin A, as the main compound synthesized by BGC 28a. Molecular networking further identified 18 additional lunaemycins, with 14 of them having their structure elucidated by HRMS/MS. Antimicrobial assays demonstrated a significant bactericidal activity of lunaemycins against Gram-positive bacteria, including multi-drug resistant clinical isolates. Our work demonstrates how an accurate in silico analysis of a cryptic BGC can highly facilitate the identification, the structural elucidation, and the bioactivity of its associated specialized metabolites.

Evaluation of Biosynthetic Pathway and Engineered Biosynthesis of Morphine with CRISPR

As a secondary metabolite produced by opium poppy (Papaver somniferum), morphine is a valuable subject for research and application purposes within the medical field. Thanks to the research conducted throughout the past few decades, the main pathway for the biosynthesis of morphine has been thoroughly elucidated. In addition, whole genome sequencing and evolutionary genomics of opium poppy have provided us with crucial information regarding specific details of such a pathway on the molecular level, which includes coding regions and functions of key enzymes that play vital roles in the production of specific metabolites. These advantages can be combined with the application of current genome editing techniques, such as CRISPR, to allow the regulated and optimized production of desirable metabolites through manipulating particular genes. This research focuses on the functional evaluation of key enzymes in the biosynthetic pathway of morphine in opium poppy, further exploring possibilities of regulated production of morphine with CRISPR.

Metabolomics of bacterial-fungal pairwise interactions reveal conserved molecular mechanisms.

Bacterial-fungal interactions (BFIs) can shape the structure of microbial communities, but the small molecules mediating these BFIs are often understudied. We explored various optimization steps for our microbial culture and chemical extraction protocols for bacterial-fungal co-cultures, and liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed that metabolomic profiles are mainly comprised of fungi derived features, indicating that fungi are the key contributors to small molecules in BFIs. LC-inductively coupled plasma MS (LC-ICP-MS) and MS/MS based dereplication using database searching revealed the presence of several known fungal specialized metabolites and structurally related analogues in these extracts, including siderophores such as desferrichrome, desferricoprogen, and palmitoylcoprogen. Among these analogues, a novel putative coprogen analogue possessing a terminal carboxylic acid motif was identified from Scopulariopsis sp. JB370, a common cheese rind fungus, and its structure was elucidated via MS/MS fragmentation. Based on these findings, filamentous fungal species appear to be capable of producing multiple siderophores with potentially different biological roles (i.e. various affinities for different forms of iron). These findings highlight that fungal species are important contributors to microbiomes via their production of abundant specialized metabolites and that elucidating their role in complex communities should continue to be a priority.

Computation-aided studies related to the induction of specialized metabolite biosynthesis in microbial co-cultures: An introductory overview

Co-cultivation is an effective method of inducing the production of specialized metabolites (SMs) in microbial strains. By mimicking the ecological interactions that take place in natural environment, this approach enables to trigger the biosynthesis of molecules which are not formed under monoculture conditions. Importantly, microbial co-cultivation may lead to the discovery of novel chemical entities of pharmaceutical interest. The experimental efforts aimed at the induction of SMs are greatly facilitated by computational techniques. The aim of this overview is to highlight the relevance of computational methods for the investigation of SM induction via microbial co-cultivation. The concepts related to the induction of SMs in microbial co-cultures are briefly introduced by addressing four areas associated with the SM induction workflows, namely the detection of SMs formed exclusively under co-culture conditions, the annotation of induced SMs, the identification of SM producer strains, and the optimization of fermentation conditions. The computational infrastructure associated with these areas, including the tools of multivariate data analysis, molecular networking, genome mining and mathematical optimization, is discussed in relation to the experimental results described in recent literature. The perspective on the future developments in the field, mainly in relation to the microbiome-related research, is also provided.

Hairy Root Induction of Taxus baccata L. by Natural Transformation with Rhizobium rhizogenes

Paclitaxel (Taxol®) is a potent anticancer agent, but the widespread pharmaceutical use of paclitaxel is hampered by its limited availability due to low accumulation levels in the native yew (Taxus spp.) plants. Currently, hairy root culture is an emerging biotechnological tool that presents several advantages such as reduced costs and higher specialized metabolite production, therefore, its application to paclitaxel production can be of commercial and medicinal interest. The objective of present study was to induce hairy root in Taxus baccata L. by transformation with the wild type Rhizobium rhizogenes A4 strain. Thus, T. baccata was inoculated by three different inoculation methods: (a) ex vitro seedlings inoculation by direct injection of a liquid bacterial culture; (b) ex vitro needles inoculation by liquid co-culturing with bacteria; (c) ex vitro shoots inoculation by dipping liquid bacterial culture. Hairy roots were formed only from ex vitro seedlings inoculated by the direct inoculation method, with transformation efficiency of 14.3%. Formation of hairy roots was observed two months after inoculation. This project forms the basis for the establishment of hairy root cultures from T. baccata for the production of paclitaxel.

Cyclitol metabolism is a central feature of Burkholderia leaf symbionts.

The symbioses between plants of the Rubiaceae and Primulaceae families with Burkholderia bacteria represent unique and intimate plant-bacterial relationships. Many of these interactions have been identified through PCR-dependent typing methods, but there is little information available about their functional and ecological roles. We assembled 17 new endophyte genomes representing endophytes from 13 plant species, including those of two previously unknown associations. Genomes of leaf endophytes belonging to Burkholderia s.l. show extensive signs of genome reduction, albeit to varying degrees. Except for one endophyte, none of the bacterial symbionts could be isolated on standard microbiological media. Despite their taxonomic diversity, all endophyte genomes contained gene clusters linked to the production of specialized metabolites, including genes linked to cyclitol sugar analog metabolism and in one instance non-ribosomal peptide synthesis. These genes and gene clusters are unique within Burkholderia s.l. and are likely horizontally acquired. We propose that the acquisition of secondary metabolite gene clusters through horizontal gene transfer is a prerequisite for the evolution of a stable association between these endophytes and their hosts.

MALDI Mass Spectrometry Imaging Reveals the Existence of an N-Acyl-homoserine Lactone Quorum Sensing System in Pseudomonas putida Biofilms.

Quorum sensing (QS) is generally used to describe the process involving the release and recognition of signaling molecules, such as N-acyl-homoserine lactones, by bacteria to coordinate their response to population density and biofilm development. However, detailed information on the heterogeneity of QS metabolites in biofilms remains largely unknown. Here, we describe the utilization of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) to follow the production of specific metabolites, including QS metabolites, during Pseudomonas putida biofilm development. To do so, a method to grow an agar-based biofilm was first established, and MALDI-MSI was used to detect and visualize the distribution of QS metabolites in biofilms at different cultivation times. This study demonstrated that N-acyl-homoserine lactones are homogeneously produced in the early stages of P. putida biofilm formation. In contrast, the spatial distribution of quinolones and pyochelin correlated with the swarming motility of P. putida in mature biofilms. These two metabolites are involved in the production of extracellular polymeric substances and iron chelators. Our study thus contributes to establishing the specific temporal regulation and spatial distribution of N-acyl-homoserine lactone-related metabolites and quinolone and pyochelin in P. putida biofilms.

Genomic potential of Streptomyces roseochromogenes NRRL 3504 for the production of specialized metabolites: analysis in silico

Streptomyces roseochromogenes NRRL 3504 is the only known producer of aminocoumarin antibiotic clorobiocin – an inhibitor of bacterial DNA-gyrase and topoisomerase IV. Sequencing of NRRL 3504 genome revealed a plethora of specialized metabolite biosynthetic gene clusters (BGCs) within the latter, attesting to the significant potential of this species for the production of various as-yet-unknown bioactive compounds. Here we report bioinformatic analysis of NRRL 3504 genome aimed to better understand what kind of small molecules this strain could produce and genetic mechanisms that may limit their production. In addition to the most commonly applied bioinformatic service for BGCs detection antiSMASH, we turned to alternative tools for secondary metabolome in silico analysis such as PRISM, DeepBGC, ARTS, SEMPI and GECCO. While different genome mining applications pointed to a common core set of BGCs within the NRRL 3504, each tool having its own algorithm of BGCs detection also discovered certain number of non-overlapping clusters. This was especially true for machine learning tool DeepBGC that unearthed the biggest number of BGCs. To summarize the obtained results we used BGCViz tool, which visualizes and integrates BGC annotations from various sources by using genomic coordinates. We discuss the genetic and structural diversity of the BGCs and outline the most interesting, in opinion, targets for further investigations. Most of the described BGCs are most likely silent due to very low or zero transcription. Therefore, it might be needed to find the ways to activate the transcription of the BGCs of interest. To this end, we mined NRRL 3504 genome for the orthologs of global regulatory genes known to be involved in regulation of specialized metabolism of S. coelicolor A3(2). We were able to identify almost all plausible global regulators of interest in NRRL 3504, implying that overall scheme of regulation of specialized metabolism in A3(2) and NRRL 3504 might be similar. Results of our work set the stage for a more detailed experimental scrutiny of silent specialized metabolome of NRRL 3504.

Specialized metabolism by trichome-enriched Rubisco and fatty acid synthase components.

Acylsugars, specialized metabolites with defense activities, are secreted by trichomes of many solanaceous plants. Several acylsugar metabolic genes (AMGs) remain unknown. We previously reported multiple candidate AMGs. Here, using multiple approaches, we characterized additional AMGs. First, we identified differentially expressed genes between high- and low-acylsugar-producing F2 plants derived from a cross between cultivated tomato (Solanum lycopersicum) and a wild relative (Solanum pennellii), which produce acylsugars that are ∼1% and ∼20% of leaf dry weight, respectively. Expression levels of many known and candidate AMGs positively correlated with acylsugar amounts in F2 individuals. Next, we identified lycopersicum-pennellii putative orthologs with higher nonsynonymous to synonymous substitutions. These analyses identified four candidate genes, three of which showed enriched expression in stem trichomes compared to underlying tissues (shaved stems). Virus-induced gene silencing confirmed two candidates, Sopen05g009610 [beta-ketoacyl-(acyl-carrier-protein) reductase; fatty acid synthase component] and Sopen07g006810 (Rubisco small subunit), as AMGs. Phylogenetic analysis indicated that Sopen05g009610 is distinct from specialized metabolic cytosolic reductases but closely related to two capsaicinoid biosynthetic reductases, suggesting evolutionary relationship between acylsugar and capsaicinoid biosynthesis. Analysis of publicly available datasets revealed enriched expression of Sopen05g009610 orthologs in trichomes of several acylsugar-producing species. Similarly, orthologs of Sopen07g006810 were identified as solanaceous trichome-enriched members, which form a phylogenetic clade distinct from those of mesophyll-expressed "regular" Rubisco small subunits. Furthermore, δ13C analyses indicated recycling of metabolic CO2 into acylsugars by Sopen07g006810 and showed how trichomes support high levels of specialized metabolite production. These findings have implications for genetic manipulation of trichome-specialized metabolism in solanaceous crops.

Genome-wide association mapping reveals genes underlying population-level metabolome diversity in a fungal crop pathogen

BackgroundFungi produce a wide range of specialized metabolites (SMs) involved in biotic interactions. Pathways for the production of SMs are often encoded in clusters of tightly arranged genes identified as biosynthetic gene clusters. Such gene clusters can undergo horizontal gene transfers between species and rapid evolutionary change within species. The acquisition, rearrangement, and deletion of gene clusters can generate significant metabolome diversity. However, the genetic basis underlying variation in SM production remains poorly understood.ResultsHere, we analyzed the metabolite production of a large population of the fungal pathogen of wheat, Zymoseptoria tritici. The pathogen causes major yield losses and shows variation in gene clusters. We performed untargeted ultra-high performance liquid chromatography-high resolution mass spectrometry to profile the metabolite diversity among 102 isolates of the same species. We found substantial variation in the abundance of the detected metabolites among isolates. Integrating whole-genome sequencing data, we performed metabolite genome-wide association mapping to identify loci underlying variation in metabolite production (i.e., metabolite-GWAS). We found that significantly associated SNPs reside mostly in coding and gene regulatory regions. Associated genes encode mainly transport and catalytic activities. The metabolite-GWAS identified also a polymorphism in the 3′UTR region of a virulence gene related to metabolite production and showing expression variation.ConclusionsTaken together, our study provides a significant resource to unravel polymorphism underlying metabolome diversity within a species. Integrating metabolome screens should be feasible for a range of different plant pathogens and help prioritize molecular studies.

Balancing Trade-Offs Imposed by Growth Media and Mass Spectrometry for Bacterial Exometabolomics.

The bacterial exometabolome consists of a vast array of specialized metabolites, many of which are only produced in response to specific environmental stimuli. For this reason, it is desirable to control the extracellular environment with a defined growth medium composed of pure ingredients. However, complex (undefined) media are expected to support the robust growth of a greater variety of microorganisms than defined media. Here, we investigate the trade-offs inherent to a range of complex and defined solid media for the growth of soil microorganisms, production of specialized metabolites, and detection of these compounds using direct infusion mass spectrometry. We find that complex media support growth of more soil microorganisms, as well as allowing for the detection of more previously discovered natural products as a fraction of total m/z features detected in each sample. However, the use of complex media often caused mass spectrometer injection failures and poor-quality mass spectra, which in some cases resulted in over a quarter of samples being removed from analysis. Defined media, while more limiting in growth, generated higher quality spectra and yielded more m/z features after background subtraction. These results inform future exometabolomic experiments requiring a medium that supports the robust growth of many soil microorganisms. IMPORTANCE Bacteria are capable of producing and secreting a rich diversity of specialized metabolites. Yet, much of their exometabolome remains hidden due to challenges associated with eliciting specialized metabolite production, labor-intensive sample preparation, and time-consuming analysis techniques. Using our versatile three-dimensional (3D)-printed culturing platform, SubTap, we demonstrate that rapid exometabolomic data collection from a diverse set of environmental bacteria is feasible. We optimized our platform by surveying Streptomyces isolated from soil on a variety of media types to assess viability, degree of specialized metabolite production, and compatibility with downstream LESA-DIMS analysis. Ultimately, this will enable data-rich experimentation, allowing for a better understanding of bacterial exometabolomes.

Response of Plant Immunity Markers to Early and Late Application of Extracellular DNA from Different Sources in Tomato (Solanum lycopersicum)

As a recently explored agricultural practice, the controlled elicitation of plants offers high potential in multiple crop needs as growth promotion, activation of defenses and the production of specific metabolites. Extracellular DNA has been identified as a plant immune system elicitor but some aspects of the plant response have not been explored. In the present work, five DNA treatments were applied in tomato plants and the response of catalase, superoxide dismutase, phenylalanine ammonia lyase activities, hydrogen peroxide, total phenolics and flavonoid contents in leaves were spectrophotometrically measured. Treatments differed by the source and concentration of DNA and plant phenological stage of application. Furthermore, mathematical modeling and principal component analysis were performed to explore the behavior of each variable and their interaction. The most effective treatment was the self-DNA application in young plants based on the intensity and duration of immune system activation. The information given by the measured variables correlated positively with the phenylpropanoid pathway markers and negatively with catalase and superoxide dismutase activities. Results reported here propose an easy way to evaluate plant immune response activation by DNA and any other elicitor and provide useful information for future development of controlled elicitation strategies in crop production.

Allelopathic Potential of Mangroves from the Red River Estuary against the Rice Weed Echinochloa crus-galli and Variation in Their Leaf Metabolome.

Mangroves are the only forests located at the sea–land interface in tropical and subtropical regions. They are key elements of tropical coastal ecosystems, providing numerous ecosystem services. Among them is the production of specialized metabolites by mangroves and their potential use in agriculture to limit weed growth in cultures. We explored the in vitro allelopathic potential of eight mangrove species’ aqueous leaf extracts (Avicennia marina, Kandelia obovata, Bruguiera gymnorhiza, Sonneratia apetala, Sonneratia caseolaris, Aegiceras corniculatum, Lumnitzera racemosa and Rhizophora stylosa) on the germination and growth of Echinochloa crus-galli, a weed species associated with rice, Oryza sativa. Leaf methanolic extracts of mangrove species were also studied via UHPLC-ESI/qToF to compare their metabolite fingerprints. Our results highlight that A. corniculatum and S. apetala negatively affected E. crus-galli development with a stimulating effect or no effect on O. sativa. Phytochemical investigations of A. corniculatum allowed us to putatively annotate three flavonoids and two saponins. For S. apetala, three flavonoids, a tannin and two unusual sulfated ellagic acid derivatives were found. Some of these compounds are described for the first time in these species. Overall, A. corniculatum and S. apetala leaves are proposed as promising natural alternatives against E. crus-galli and should be further assessed under field conditions.

The sage genome provides insight into the evolutionary dynamics of diterpene biosynthesis gene cluster in plants.

The widely cultivated medicinal and ornamental plant sage (Salvia officinalis L.) is an evergreen shrub of the Lamiaceae family, native to the Mediterranean. We assembled a high-quality sage genome of 480 Mb on seven chromosomes, and identified a biosynthetic gene cluster (BGC) encoding two pairs of diterpene synthases (diTPSs) that, together with the cytochromes P450 (CYPs) genes located inside and outside thecluster, form two expression cascades responsible for the shoot and root diterpenoids, respectively, thus extending BGC functionality from co-regulation to orchestrating metabolite production in different organs. Phylogenomic analysis indicates that the Salvia clades diverged in the early Miocene. In East Asia, most Salvia species are herbaceous and accumulate diterpenoids in storage roots. Notably, in Chinese sage S.miltiorrhiza, the diterpene BGC has contracted and the shoot cascade has been lost. Our data provide genomic insights of micro-evolution of growth type-associated patterning of specialized metabolite production in plants.

Effects of temperature, irradiance, and pH on the growth and biochemical composition of Haslea ostrearia batch-cultured in an airlift plan-photobioreactor.

Haslea ostrearia is a pennate diatom that produces marennine, a water-soluble blue pigment responsible for the greening phenomenon and the increase of organoleptic quality of oysters. Apart from the oyster industry, there is a growing interest in the mass cultivation of this diatom due to the biological activities of marennine. To gain knowledge about the feasibility to upscale production of this diatom, in particular, in the context of global warming, the effects of different temperatures (20, 25, and 30°C), irradiances (100, 200, and 300μmol photons m-2s-1), and pH (7.0, 8.0, and 9.0) on growth and biochemical composition were studied in H. ostrearia cultured in an airlift plan-photobioreactor. The maximum growth rate of H. ostrearia (0.9 ± 0.0day-1) was obtained at 20°C, 200μmol photons m-2s-1, and pH 7.0, referred to as control conditions. The highest concentration in Chla (2.5 ± 0.1µg 10-6 cells) and total fatty acids (71.6 ± 1.4mgg-1 of dry weight, DW) was observed at 20°C, 300μmol photons m-2s-1, and pH 7.0. The highest concentration of carotenoids (1.4 ± 0.1µg 10-6 cells), Chlc (1.3 ± 0.1µg 10-6 cells), and extracellular marennine (33.1 ± 0.2µg 10-6 cells) was observed at 30°C, 200μmol photons m-2s-1, and pH 7.0, and a higher protein content (309.7 ± 24.5mgg-1 of DW) at 25°C, 200μmol photons m-2s-1, and pH 7.0. The biomass of H. ostrearia was enriched with C14:0 fatty acid at 30°C, 200μmol photons m-2s-1, and pH 7.0, and with C16:0 and C16:1n - 7 fatty acids at control conditions. However, DHA C22:6n - 3 (ω-3), C22:0, and C20:0 were only observed at 300µmol photons m-2s-1, 20°C, and pH 7.0. A high abundance of essential polyunsaturated fatty acids C22:1n - 9 (ω-9) + C20:5n - 3 (EPA) was observed at 100µmol photons m-2s-1, 20°C, and pH 7.0. It is thus possible to anticipate and tune the production of specific metabolites through the control of growth conditions of the benthic diatom H. ostrearia. KEY POINTS: • Validation of H. ostrearia culture in a new photobioreactor in batch mode • Biochemical composition variation of H. ostrearia in function of growth conditions • Growth inhibition and unbalanced metabolites induced by the treatment conditions.

Zebularine, a DNA Methylation Inhibitor, Activates Anthocyanin Accumulation in Grapevine Cells

Through its role in the regulation of gene expression, DNA methylation can participate in the control of specialized metabolite production. We have investigated the link between DNA methylation and anthocyanin accumulation in grapevine using the hypomethylating drug, zebularine and Gamay Teinturier cell suspensions. In this model, zebularine increased anthocyanin accumulation in the light, and induced its production in the dark. To unravel the underlying mechanisms, cell transcriptome, metabolic content, and DNA methylation were analyzed. The up-regulation of stress-related genes, as well as a decrease in cell viability, revealed that zebularine affected cell integrity. Concomitantly, the global DNA methylation level was only slightly decreased in the light and not modified in the dark. However, locus-specific analyses demonstrated a decrease in DNA methylation at a few selected loci, including a CACTA DNA transposon and a small region upstream from the UFGT gene, coding for the UDP glucose:flavonoid-3-O-glucosyltransferase, known to be critical for anthocyanin biosynthesis. Moreover, this decrease was correlated with an increase in UFGT expression and in anthocyanin content. In conclusion, our data suggest that UFGT expression could be regulated through DNA methylation in Gamay Teinturier, although the functional link between changes in DNA methylation and UFGT transcription still needs to be demonstrated.

How Metabolomics Provides Novel Insights on Celiac Disease and Gluten-Free Diet: A Narrative Review.

Celiac disease (CD) is an inflammatory autoimmune disorder triggered by the ingestion of gluten from wheat and other cereals. Nowadays, its positive diagnosis is based on invasive approaches such as the histological examination of intestinal biopsies and positive serology screening of antibodies. After proven diagnosis, the only admissible treatment for CD individuals is strict life-long adherence to gluten-free diet (GFD), although it is not a conclusive therapy. Acting by different mechanisms and with different etiologies, both CD and GFD have a great impact on gut microbiota that result in a different taxa composition. Altered production of specific metabolites reflects these microbiota changes. In this light, the currently available literature reports some suggestions about the possible use of specific metabolites, detected by meta-omics analyses, as potential biomarkers for a CD non-invasive diagnosis. To highlight insights about metabolomics application in CD study, we conducted a narrative dissertation of selected original articles published in the last decade. By applying a systematic search, it clearly emerged how the metabolomic signature appears to be contradictory, as well as poorly investigated.

Production and Functionalities of Specialized Metabolites from Different Organic Sources.

Medicinal plants are rich sources of specialized metabolites that are of great importance to plants, animals, and humans. The usefulness of active biological compounds cuts across different fields, such as agriculture, forestry, food processing and packaging, biofuels, biocatalysts, and environmental remediation. In recent years, research has shifted toward the use of microbes, especially endophytes (bacteria, fungi, and viruses), and the combination of these organisms with other alternatives to optimize the production and regulation of these compounds. This review reinforces the production of specialized metabolites, especially by plants and microorganisms, and the effectiveness of microorganisms in increasing the production/concentration of these compounds in plants. The study also highlights the functions of these compounds in plants and their applications in various fields. New research areas that should be explored to produce and regulate these compounds, especially in plants and microbes, have been identified. Methods involving molecular studies are yet to be fully explored, and next-generation sequencing possesses an interesting and reliable approach.