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Alternaria sp. MG1, an endophytic fungus previously isolated from Merlot grape, produces resveratrol from glucose, showing similar metabolic flux to the phenylpropanoid biosynthesis pathway, currently found solely in plants. In order to identify the resveratrol biosynthesis pathway in this strain at the gene level, de novo transcriptome sequencing...
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... the resveratrol biosynthesis pathway is only found in plants (Figure 1; Tan et al., 2012;Tavares et al., 2013). ...
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... (ko00010) contributes to the early stage of resveratrol biosynthesis by converting different substrates (e.g., α-D-glucose and β-D-glucose) to phosphoenolpyruvic acid (PEP; Figure 5A-1). In total, 48 unigenes, coding 20 enzymes such as HK, PGI, pfkA, ALDO, tpiA, GADPH, PGK, PGAM, and ENO were identified in this pathway. ...
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... total, 48 unigenes, coding 20 enzymes such as HK, PGI, pfkA, ALDO, tpiA, GADPH, PGK, PGAM, and ENO were identified in this pathway. The unigene expression analysis showed the annotated unigenes encoding these enzymes exhibited a wide range of abundance variation (Figure 5B-1). For example, c10831 and c3258 showed an abundance of 2697.6 and 1093.4 ...
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Citations
... Stilbenes are synthesized via the Shikimate pathway, where stilbene synthase (STS) catalyzes the formation of simple monomeric stilbene (i.e., resveratrol, pinosylvin or piceatannol). Based on encouraging therapeutic evidence, stilbene research has fueled the best deal of interest in characterizing stilbene compounds and their natural sources for their pharmacological properties (Che et al. 2016a(Che et al. , 2016bBraga et al. 2018). ...
In modern times, medicine is predominantly based on evidence-based practices, whereas in ancient times, indigenous people relied on plant-based medicines with factual evidence documented in ancient books or folklore that demonstrated their effectiveness against specific infections. Plants and microbes account for 70% of drugs approved by the USFDA (U.S. Food and Drug Administration). Stilbenes, polyphenolic compounds synthesized by plants under stress conditions, have garnered significant attention for their therapeutic potential, bridging ancient wisdom with modern healthcare. Resveratrol, the most studied stilbene, initially discovered in grapes, red wine, peanuts, and blueberries, exhibits diverse pharmacological properties, including cardiovascular protection, antioxidant effects, anticancer activity, and neuroprotection. Traditional remedies, documented in ancient texts like the Ayurvedic Charak Samhita, foreshadowed the medicinal properties of stilbenes long before their modern scientific validation. Today, stilbenes are integral to the booming wellness and health supplement market, with resveratrol alone projected to reach a market value of 90 million US$ by 2025. However, challenges in stilbene production persist due to limited natural sources and costly extraction methods. Bioprospecting efforts reveal promising candidates for stilbene production, particularly endophytic fungi, which demonstrate high-yield capabilities and genetic modifiability. However, the identification of optimal strains and fermentation processes remains a critical consideration. The current review emphasizes the knowledge of the medicinal properties of Stilbenes (i.e., cardiovascular, antioxidant, anticancer, anti-inflammatory, etc.) isolated from plant and microbial sources, while also discussing strategies for their commercial production and future research directions. This also includes examples of novel stilbenes compounds reported from plant and endophytic fungi.
... Various types of N-glycan biosynthesis ( Figure S2) are an important pathway for polysaccharide biosynthesis and belong to the biosynthesis and metabolism of sugar in the KEGG metabolic pathway. This pathway is related to the biosynthesis of resveratrol (Che, Shi, Gao, & Zhang, 2016) and the pharmacological effects of Cordyceps sinensis (Xin et al., 2019). Therefore, It's an important focus of functional food processing and biomedicine research. ...
The development of binary protein systems featuring superior nutritional properties and applied range is an interesting and challenging task in the food industry. In this study, the tilapia-soybean protein co-precipitates (TSPCs) with different mass ratios of tilapia meat and soybean meal were constructed. Results of physicochemical properties showed that the highest solubility and thermal stability values of TSPCs were 81.90 % and 90.30 °C, respectively. TSPCs have the full complement of amino acids and enhanced nutritional quality compared to tilapia protein isolate (TPI) and soybean protein isolate (SPI). TSPC2:1 and TSPC1:1 contained the highest levels of tryptophan, aspartic acid, glycine, histidine, and arginine relative to TPI and SPI. The in vitro protein digestibility and protein digestibility corrected amino acid scores of TSPCs were also higher than that of SPI. SDS-PAGE revealed that TSPCs contained protein subunits from TPI and SPI. Moreover, the lysine-to-arginine ratio and β subunit were greatly correlated with protein digestibility with correlation coefficients of −0.962 (P < 0.01) and −0.971 (P < 0.01), respectively. Compared to SPI, TSPCs displayed a lower lysine-to-arginine ratio and β-conglycinin content, which improved its digestibility. Proteomic analysis indicated that TSPC1:1 had 989 unique proteins, which gives TSPCs enhanced biological properties compared to TPI and SPI, allowing them to participate in a broad range of biochemical metabolic and signal transduction pathways. The study would advance the utilization of mixed proteins toward exceptional food industry applications.
... Deciphering the resveratrol biosynthesis pathway and its relevant key genes in endophytic fungi is substantial for delicate control of the production condition and overcoming the problems of low and unstable yield (Che et al., 2016b). The complete biosynthetic pathway of resveratrol is only identified in plants as the phenylpropanoid pathway (Figure 2). ...
... MG1 was conducted during resveratrol production and a total of 84 genes were annotated encompassing four major resveratrol metabolic pathways that represent 38 coding enzymes (20 for glycolysis, 10 for phenylalanine biosynthesis, 4 for phenylpropanoid biosynthesis, and 4 for stilbenoid biosynthesis). The expression of PAL, 4CL, and CYP73A-related genes was observed (Che et al., 2016b). Additionally, Lu et al. (2019) used the WGS and obtained the same results and found that 4CL and CHS were up-regulated, resulting in a twofold increase in resveratrol production. ...
... Additionally, Lu et al. (2019) used the WGS and obtained the same results and found that 4CL and CHS were up-regulated, resulting in a twofold increase in resveratrol production. Resveratrol synthase (ST) was not annotated in the previous studies (Che et al., 2016b;Lu et al., 2019), however, CHS was successfully annotated instead (Jeandet et al., 2010). CHS is an isoenzyme of ST that is responsible for the biosynthesis of naringenin and possesses catalytic activity to form resveratrol, whereas naringenin and resveratrol are produced from the same substrate, p-coumaric acid . ...
Resveratrol biosynthesis, optimization, induction, bio-transformation and bio-degradation in mycoendophytes. Resveratrol (3,4,5-trihydroxystilbene) is a naturally occurring polyphenolic stilbene compound produced by certain plant species in response to biotic and abiotic factors. Resveratrol has sparked a lot of interest due to its unique structure and approved therapeutic properties for the prevention and treatment of many diseases such as neurological disease, cardiovascular disease, diabetes, inflammation, cancer, and Alzheimer's disease. Over the last few decades, many studies have focused on the production of resveratrol from various natural sources and the optimization of large-scale production. Endophytic fungi isolated from various types of grapevines and Polygonum cuspidatum, the primary plant sources of resveratrol, demonstrated intriguing resveratrol-producing ability. Due to the increasing demand for resveratrol, one active area of research is the use of endophytic fungi and metabolic engineering techniques for resveratrol's large-scale production. The current review addresses an overview of endophytic fungi as a source for production, as well as biosynthesis pathways and relevant genes incorporated in resveratrol biosynthesis. Various approaches for optimizing resveratrol production from endophytic fungi, as well as their bio-transformation and bio-degradation, are explained in detail.
... Shotgun metagenome analysis of bacterial endophytes at different plant ages revealed that siderophore and auxin-related plant growth promoting traits are highly represented in 3-year-old plants of Panax ginseng (Hong et al., 2019). Besides genome sequencing, transcriptome analysis, is also useful in the discovery of new genes and to understand their regulation (Che et al., 2016). The transcriptome of an endophyte Cladosporium cladosporioides indicated 40 unigenes which were related to paclitaxel biosynthesis (taxadiene synthase, taxadiene 5-alphahydroxylase, 2-alpha-hydroxytaxane 2-O-benzoyltransferase) and were homologous to the genes of host plant, Taxus media (Miao et al., 2018). ...
Endophytes as a ubiquitous associate of the plant are considered as a promising candidate for sustainable agriculture. These organisms play a pivotal role in the regulation of the primary and secondary metabolism of their host plant. The direct and long-lasting interaction of endophytes with the host enables them to escape from harsh environmental conditions. Especially, their endophytic nature makes them better candidates over epiphytes and rhizospheric microbes in interaction with plants. Current research findings revealed that the endophytes help plants in making nutrient acquisition from the soil, nitrogen fixation, phosphate availability, phytohormone and antimicrobial production. There is a huge potential for developing novel products like endophytes-based microbial formulations and elicitors to improve plant health, ameliorating stress tolerance in plants and source of therapeutically important secondary metabolites. The present review specifically dealt with attributes such as host-tissue specificity of endophytes, the importance of seed-associated endophytes, endophyte-parasite plant-host plant interaction as well as their applications in plant in-vitro systems and as microbial consortium. In addition, the conserved endophytic microbial communities in different plants are also looked upon possibly to understand the plant-endophytic microbiome on similar lines of the animal-gut microbiome. Primarily, the purpose of this review is to implicate the endophytic flora as probiotics influencing overall plant health and their survival under extreme environmental conditions.
... Another study identified 18,570 unigenes with an N50 length of 2153 bp and a mean length of 1122.38 bp in Alternaria sp. MG1 isolated from Vitis vinifera (Che et al. 2016). In contrast, the predicted number of unigenes in A. alternata PN1, A. alternata PN2, A. alternata ATCC34957, A. brassicae J3, and A. solani altNL03003 were 11,495, 11,387, 12,500, 11,593, and 11,804 respectively (Rajarammohan et al. 2019). ...
Background: Alternaria panax is the causative agent of black spot disease in Panax notoginseng, which causes significant yield loss. However, the molecular mechanisms underlying the pathogenicity remain mostly unknown.
Objective: We sequenced the transcriptome of A. panax during infecting P. notoginseng leaves using next-generation RNA-seq to understand the molecular aspects of black spot disease.
Methods: In this study, we sequenced the A. panax transcriptome during infecting P. notoginseng leaves through next-generation sequencing to explore the pathogenesis genes that may be responsible for black spot disease on P. notoginseng.
Result: The de novo transcriptome assembly of A. panax produced 23,036 unigenes, of which 18,096 genes were functionally annotated by at least one protein database. GO enrichment analysis and enriched KEGG pathways of differentially up-regulated genes suggest that most of the genes are associated with metabolic processes, catalytic activity, starch, and sucrose metabolism during infection. Many pathogenesis-associated genes, including genes encoding secreted proteins, candidate secreted effectors, cell wall degrading enzymes, transcription factors, and transporters, were upregulated in A. panax during infection. In addition, the secondary metabolite biosynthesis genes, including cytochrome P450, and nonribosomal peptide synthetases, were also identified in this study.
Conclusion: Differential gene expression analysis has confirmed that A. panax infection was mainly present in the middle and final stages. The findings show that these pathogenesis-associated genes in A. panax may be critical factors for the P. notoginseng black spots disease.
... High quantities of this compound are required for applications in functional food [88]. At present resveratrol is mainly extracted from plant by-products, with limited availability [67,89]. ...
... De novo transcriptome sequencing of Alternaria sp. MG1, an endophytic strain isolated from grape which is able to synthesize resveratrol from glucose, identified 20 genes for glycolysis, 10 for phenylalanine biosynthesis, and 4 genes for phenylpropanoid and stilbenoid biosynthesis [67]. Phenylalanine ammonia-lyase (PAL), cytochrome P-450 CYP73A, 4-coumarate coenzyme A ligase (4CL), and chalcone synthase (CHS) were annotated for converting L-phenylalanine into resveratrol. ...
Recent investigations carried out all over the world have disclosed the capacity of a multitude of endophytic fungal species/strains to produce bioactive compounds which are the same or structurally related to those synthesized by their host plants. This intriguing phenomenon has implemented huge research activity aimed at ascertaining the nature of the biological processes underlying this convergence, as well as to characterize the genetic bases of the biosynthetic schemes. Insights on these basic issues may support the biotechnological exploitation of wild strains, and their eventual improvement through genome editing. Aspects concerning the use of next generation sequencing technologies for the comparative elucidation of the biosynthetic pathways operating in endophytic fungi and their host plants are reviewed in this paper in view of applicative perspectives. Our literature search yielded 21 references dealing with at least 26 strains which have been the subject of experimental activities involving massively parallel genome and transcriptome sequencing methods in the last eight years.
... As an example, RNAseq of Alternaria sp. MG1 was done to identify putative genes involved in the proposed biosynthetic pathway of 11 (Che et al., 2016). An RNAseq library was constructed, sequenced by Illumina technology, assembled to unigenes, annotated and analyzed for gene and pathway expression. ...
The bioprospecting of secondary metabolites from endophytic fungi received great attention in the 1990s and 2000s, when the controversy around taxol production from Taxus spp. endophytes was at its height. Since then, hundreds of reports have described the isolation and characterization of putative secondary metabolites from endophytic fungi. However, only very few studies also report the genetic basis for these phenotypic observations. With low sequencing cost and fast sample turnaround, genetics- and genomics-based approaches have risen to become comprehensive approaches to study natural products from a wide-range of organisms, especially to elucidate underlying biosynthetic pathways. However, in the field of fungal endophyte biology, elucidation of biosynthetic pathways is still a major challenge. As a relatively poorly investigated group of microorganisms, even in the light of recent efforts to sequence more fungal genomes, such as the 1000 Fungal Genomes Project at the Joint Genome Institute (JGI), the basis for bioprospecting of enzymes and pathways from endophytic fungi is still rather slim. In this review we want to discuss the current approaches and tools used to associate phenotype and genotype to elucidate biosynthetic pathways of secondary metabolites in endophytic fungi through the lens of bioprospecting. This review will point out the reported successes and shortcomings, and discuss future directions in sampling, and genetics and genomics of endophytic fungi. Identifying responsible biosynthetic genes for the numerous secondary metabolites isolated from endophytic fungi opens the opportunity to explore the genetic potential of producer strains to discover novel secondary metabolites and enhance secondary metabolite production by metabolic engineering resulting in novel and more affordable medicines and food additives.
... However, genome sequencing indicated that the genes in the host plant genus were significantly different from those found in endophytic fungi. They seemed to evolve independently (Yang et al. 2014;Che et al. 2016b;Lu et al. 2016Lu et al. , 2019. Investigating the mechanisms of the synthesis of plant metabolites by endophytes plays an important role for the commercial exploitation of endophytic fungi as new producers of bioactive molecules, especially those plants produced in low levels (Che et al. 2016a). ...
Due to increasingly limited water resources, diminishing farmland acreage, and potentially negative effects of climate change, an urgent need exists to improve agricultural productivity to feed the ever-growing population. Plants interact with microorganisms at all trophic levels, adapting growth, developmental, and defense responses within a complicated network of community members. Endophytic fungi have been widely reported for their ability to aid in the defense of their host plants. Currently, many reports focus on the application of endophytic fungi with the capability to produce valuable bioactive molecules, while others focus on endophytic fungi as biocontrol agents. Plant responses upon endophytic fungi colonization are also good for the immune system of the plant. In this paper, the possible mechanisms between endophytic fungi and their hosts were reviewed. During long-term evolution, plants have acquired numerous beneficial strategies in response to endophytic fungi colonization. The interaction of endophytic fungi with plants modulates the relationship between plants and both biotic and abiotic stresses. It has previously been reported that this endophytic relationship confers additional defensive mechanisms on the modulation of the plant immune system, as the result of the manipulation of direct antimicrobial metabolites such as alkaloids to indirect phytohormones, jasmonic acid, or salicylic acid. Furthermore, plants have evolved to cope with combinations of stresses and experiments are required to address specific questions related to these multiple stresses. This review summarizes our current understanding of the intrinsic mechanism to better utilize these benefits for plant growth and disease resistance. It contributes new ideas to increase plant fitness and crop productivity.
... The draft model was obtained via homologous alignment with reference strains using previously described procedures, followed by manual curation. Firstly, 97 reactions in the biosynthesis pathways of phenylpropanoid, stilbene synthesis, flavonoid synthesis, and lignin biosynthesis were added to the model by referring to the KEGG database, BLASTp, and literature reports [24,37]. A total of 220 transport reactions annotated from the TCDB database and 133 exchange reactions were also added into the model iYL1539. ...
... Nucleic acids (DNA and RNA), proteins, lipids, carbohydrates, and ash were considered in the biomass equation. Total DNA and RNA were extracted [37] and their specific composition was calculated based on the genome sequence with a GC content of 50.96%. Amino acid composition was analyzed using an amino acid analyzer (L-8900, HITACHI, Japan). ...
... Primers of target genes (4CL and CHS) were designed according to the transcriptome data of Alternaria sp. MG1 [37] and the sequences are shown in Table 4. For each gene, the expression value was normalized with respect to the reference gene EF1 [76]. ...
Background
Alternaria sp. MG1, an endophytic fungus isolated from grape, is a native producer of resveratrol, which has important application potential. However, the metabolic characteristics and physiological behavior of MG1 still remains mostly unraveled. In addition, the resveratrol production of the strain is low. Thus, the whole-genome sequencing is highly required for elucidating the resveratrol biosynthesis pathway. Furthermore, the metabolic network model of MG1 was constructed to provide a computational guided approach for improving the yield of resveratrol.
Results
Firstly, a draft genomic sequence of MG1 was generated with a size of 34.7 Mbp and a GC content of 50.96%. Genome annotation indicated that MG1 possessed complete biosynthesis pathways for stilbenoids, flavonoids, and lignins. Eight secondary metabolites involved in these pathways were detected by GC–MS analysis, confirming the metabolic diversity of MG1. Furthermore, the first genome-scale metabolic network of Alternaria sp. MG1 (named iYL1539) was reconstructed, accounting for 1539 genes, 2231 metabolites, and 2255 reactions. The model was validated qualitatively and quantitatively by comparing the in silico simulation with experimental data, and the results showed a high consistency. In iYL1539, 56 genes were identified as growth essential in rich medium. According to constraint-based analysis, the importance of cofactors for the resveratrol biosynthesis was successfully demonstrated. Ethanol addition was predicted in silico to be an effective method to improve resveratrol production by strengthening acetyl-CoA synthesis and pentose phosphate pathway, and was verified experimentally with a 26.31% increase of resveratrol. Finally, 6 genes were identified as potential targets for resveratrol over-production by the recently developed methodology. The target-genes were validated using salicylic acid as elicitor, leading to an increase of resveratrol yield by 33.32% and the expression of gene 4CL and CHS by 1.8- and 1.6-fold, respectively.
Conclusions
This study details the diverse capability and key genes of Alternaria sp. MG1 to produce multiple secondary metabolites. The first model of the species Alternaria was constructed, providing an overall understanding of the physiological behavior and metabolic characteristics of MG1. The model is a highly useful tool for enhancing productivity by rational design of the metabolic pathway for resveratrol and other secondary metabolites.
Electronic supplementary material
The online version of this article (10.1186/s12934-019-1063-7) contains supplementary material, which is available to authorized users.
... The N50 length of the unigenes was 2,451 bp, which was longer than the N50 obtained from the transcriptome sequencing of Alternaria sp. MG1 (N50 = 2,153 bp) using the Illumina HiSeq 2500 platform in a previous study (Che et al., 2016). Collectively, the results indicate that the quality and integrity of the obtained sequences are high. ...
A total of 32,284 unigenes were obtained from the transcriptome of Alternaria tenuissima, a pathogenic fungus causing foliar disease in tomato, using next-generation sequencing (NGS) technology. In total, 24,670 unigenes were annotated using five databases, including NCBI non-redundant protein, Swiss-Prot, euKaryotic Orthologous Groups, Kyoto Encyclopedia of Genes and Genomes, and the Gene Ontology. A total of 1,140 simple sequence repeats were also identified for use as molecular markers. Sixteen of the simple sequence repeat loci were selected to study the population structure of A. tenuissima. A population genetic analysis of 191 A. tenuissima isolates, sampled from four geographic regions in China, indicated that A. tenuissima had a high level of genetic diversity, and that the selected simple sequence repeat markers could reliably capture the genetic variation. The null hypothesis of random mating was rejected for all four geographic regions in China. Isolation by distance was observed for the entire data set, but not within clusters, which is indicative of barriers to gene flow among geographic regions. The analyses of Bayesian and principal coordinates, however, did not separate four geographic regions into four separate genetic clusters. The different levels of historical migration rates suggest that isolation by distance did not represent a major biological obstacle to the spread of A. tenuissima. The potential epidemic spread of A. tenuissima in China may occur through the transport of plant products or other factors. The presented results provide a basis for a comprehensive understanding of the population genetics of A. tenuissima in China.
