Genomic sequencing, genome-scale metabolic network reconstruction, and in silico flux analysis of the grape endophytic fungus Alternaria sp. MG1

Abstract

BackgroundAlternaria 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.ResultsFirstly, 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.ConclusionsThis 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 materialThe online version of this article (10.1186/s12934-019-1063-7) contains supplementary material, which is available to authorized users.

The Evolution of Flavonoid Biosynthesis: A Bryophyte Perspective

“…In contrast, sequences with similarity to 4CL do occur in rhodophyte and chlorophyte genomes (Labeeuw et al, 2015;de Vries et al, 2017), suggesting the existence of this enzyme in a shared ancestor of land plants and algae before the ancestral divergence of the red algae (Labeeuw et al, 2015). A further aspect yet to be addressed is the presence in fungi of genes with significant sequence similarity to those of the phenylpropanoid pathway (Bilska et al, 2018;Lu et al, 2019). As the separation of fungi and plants is thought to have occurred during the early stages of eukaryote divergence (Burki, 2014;Burki et al, 2020), it is possible that these may represent cases of convergent evolution.…”

“…Notably, in fungal species there have been recent reports of both the production of flavonoids and the presence of genes with significant sequence similarity to those of the phenylpropanoid pathway of land plants. Findings include the detection of a range of phenylpropanoids, including flavonoids, in Fusarium (Bilska et al, 2018) and detection of flavonoids and candidate genes for stilbene production in Alternaria (Lu et al, 2019). The significance of phenylpropanoid biosynthesis being present in fungi for the current proposals on the evolutionary origins of flavonoid biosynthesis has yet to be addressed.…”

Genetic engineering of the precursor supply pathway for the overproduction of the nC14-surfactin isoform with promising MEOR applications

“…Based on these characteristics, a number of studies have shown that surfactin-based extraction has good prospects in MEOR [ 11 ]. The industrial application of surfactin is not limited by yield, since the surfactin production of engineered strains reached 10–20 g/L [ 12 ]. At present, the point of concern is the heterogeneity of surfactin structure, which closely affects the functional activity of surfactin preparations.…”

“…At present, strategies proposed to modify the structure of surfactin are all based on the metabolism of branched-chain amino acids, because some precursors for the synthesis of branched chain fatty acids presented in surfactin are derived from the branched chain amino acids valine, leucine and isoleucine [ 12 ]. For example, the proportion of surfactin with even β-hydroxy fatty acid components C 14 and C 16 increased with the addition of Arg, Gln or Val, whereas the addition of Cys, His, Ile, Leu, Met, and Ser enhanced the proportion of odd β-hydroxy fatty acids in B. subtilis TD7 [ 20 ].…”

Applications of microbial co‐cultures in polyketides production

“…First, high-throughput biotechnology has been used to study the mechanisms for microbial co-culture to produce polyketides, such as genome, transcriptome, metabolome and proteome. Besides, constructing genome-scale metabolic model has also been developed to predict the metabolic flow of co-culture system, which is helpful to clarify the interaction between micro-organisms and provides theoretical basis for the development of artificial intervention to the co-culture system (Lu et al 2019). In addition, cell fusion and gene engineering biotechnology can also be used to construct engineering cell, change the metabolic flow of micro-organisms or construct new metabolic pathways.…”