Project description:The thermophilic filamentous fungi Myceliophthora thermophila (Sporotrichum thermophile) has an ability to decompose cellulolytic biomass. To identify the genes and proteins involved in this process, we explored the transcriptomes of M. thermophila grown at 45 °C on different agricultural straws (oat, triticale, canola, flax straws).
Project description:The thermophilic filamentous fungi Myceliophthora thermophila (Sporotrichum thermophile) and Thielavia terrestris are proficient decomposers of cellulose, suggesting that they will be a rich source of thermostable industrial enzymes for lignocellulose degradation. To identify the genes and proteins involved in this process, we explored the transcriptomes of M. thermophila and T. terrestris growing at 45 ºC on either glucose, alfalfa, or barley straw by short-read sequencing of extracted mRNA. To better understand the adaptations that allow these fungi to grow at elevated temperatures, we compared their transcriptomes when growing at 34C to their transcritomes at 45C, and also to the transcriptome of the related fungus Chaetomium globosum, which does not grow at 45C. RNA was extracted from cultures in early growth stage growing with glucose, alfalfa, or barley straw as carbon source at 34C or 45C (M. thermophila and T. terrestris); duplicate cultures were sampled in some conditions.
Project description:Transcriptional landscape of MyceliopTranscriptional profiling of Myceliophthora thermophila on galactose and metabolic engineering for improved galactose utilization thora thermophila responding to soluble starch and the role of regulator AmyR on polysaccharide degradation Efficient biological conversion of all sugars from lignocellulosic biomass is necessary for cost-effective production of biofuels and commodity chemicals. Galactose is the next most abundant sugar in many hemicelluloses and it will be important to capture this carbon for an efficient bioconversion process of plant biomass. Thermophilic fugus Myceliophthora thermophila has been used as cell factory to produce biochemicals directly from renewable polysaccharides. This study determined the transcriptomic profiles of M. thermophila responding to galactose and identified the gene involved in the oxido-reductive pathway for galactose degradation, of which hexokinase might be the rate-limiting enzyme in this thermophilic fungus. Galactokinase was necessary for high induction of galactose transporter and disruption of galK resulted in decreased galactose utilization. Both of galactokinase deletion and sugar transporter overexpression could further activate the oxido-reductive pathway and led to improved galactose utilization. In addition, disruption of the Leloir pathway brought about reduced tolerance to cell wall perturbation, oxidative stress and high osmolality. In order to accelerate galactose consumption, the third galactose-degradation pathway- De Ley-Doudoroff pathway was successfully integrated into M. thermophila and the consumption rate of galactose was increased by 57%. This study will be benefit for the rational design of fungal strains to produce biofuels and biochemical from plant biomass, especially, marine plant biomass with the abundant of galactose, such as red seaweed and molasses.
Project description:The thermophilic filamentous fungi Myceliophthora thermophila (Sporotrichum thermophile) and Thielavia terrestris are proficient decomposers of cellulose, suggesting that they will be a rich source of thermostable industrial enzymes for lignocellulose degradation. To identify the genes and proteins involved in this process, we explored the transcriptomes of M. thermophila and T. terrestris growing at 45 ºC on either glucose, alfalfa, or barley straw by short-read sequencing of extracted mRNA. To better understand the adaptations that allow these fungi to grow at elevated temperatures, we compared their transcriptomes when growing at 34C to their transcritomes at 45C, and also to the transcriptome of the related fungus Chaetomium globosum, which does not grow at 45C.
Project description:Thermothelomyces thermophilus, formerly known as Myceliophthora thermophila, is used in industry to produce lignocellulolytic enzymes and heterologous proteins. However, the transcriptional network driving the expression of these proteins remains elusive. As a first step to systematically uncover this network, we investigated growth, protein secretion, and transcriptomic fingerprints of strains deficient in the cellulolytic transcriptional regulators Clr1, Clr2, and Clr4, respectively. The genes encoding Clr1, Clr2, and Clr4 were individually deleted using split marker or the CRISPR/Cas12a technology and the resulting strains as well as the parental strain were cultivated in bioreactors under chemostat conditions using glucose as carbon source. During steady state conditions, cellulose was added instead of glucose to study the genetic and cellular responses in all four strains to the shift in carbon source availability. Notably, the clr1 and clr2 deletion strains were unable to continue to grow on cellulose, demonstrating a key role of both regulators in cellulose catabolism. Their transcriptomic fingerprints uncovered not only a lack of cellulase gene expression but also reduced expression of genes predicted to encode hemicellulases, pectinases, and esterases. In contrast, the growth of the clr4 deletion strain was very similar compared to the parental strain. However, a much stronger expression of cellulases, hemicellulases, pectinases, and esterases was observed. The data gained in this study suggest that both transcriptional regulators Clr1 and Clr2 activate the expression of genes predicted to encode cellulases as well as hemicellulases, pectinases, and esterases. They further suggest that Clr1 controls the basal expression of cellulases and initiates the main lignocellulolytic response to cellulose via induction of clr2 expression. In contrast, Clr4 seems to act as a repressor of the lignocellulolytic response presumably via controlling clr2 expression. Comparative transcriptomics in all four strains revealed potentially new regulators in carbohydrate catabolism and lignocellulolytic enzyme expression that define a candidate gene list for future analyses.
Project description:Myceliophthora thermophila is a thermophilic fungus with great biotechnological characteristics for industrial applications, which can degrade and utilize all major polysaccharides in plant biomass. Nowadays, it has been developing into a platform for production of enzyme, commodity chemicals and biofuels. Therefore, an accurate genome-scale metabolic model would be an accelerator for this fungus becoming a universal chassis for biomanufacturing. Here we present a genome-scale metabolic model for M. thermophila constructed using an auto-generating pipeline with consequent thorough manual curation. Temperature plays a basic and critical role for the microbe growth. we are particularly interested in the genome wide response at metabolic layer of M. thermophilia as it is a thermophlic fungus. To study the effects of temperature on metabolic characteristics of M. thermophila growth, the fungus was cultivated under different temperature. The metabolic rearrangement predicted using context-specific GEMs integrating transcriptome data.The developed model provides new insights into thermophilic fungi metabolism and highlights model-driven strain design to improve biotechnological applications of this thermophilic lignocellulosic fungus.
Project description:Producing the fuels and chemicals from renewable plant biomass has been thought as a feasible way for global sustainable development. However, the economical efficiency of biorefinery remains challenges. Here a cellulolytic thermophilic fungus, Myceliophthora thermophila, was constructed into a platform through metabolic engineering, which can efficiently convert lignocellulose to important bulk chemicals for polymers, four carbon 1, 4-diacids (malic and succinic acid), directly from lignocellulose without any extra enzymes addition or complicated pretreatment, with titer of over 200 g/L on cellulose and 110 g/L on plant biomass (corncob) during fed-batch fermentation. Our study represents a milestone of consolidated bioprocessing technology (CBP) and offers a new promising system for cost-effectively production of biomass-based chemicals and potentially fuels.
Project description:An efficient Agrobacterium-mediated genetic transformation based on the plant binary vector pPZP-RCS2 was carried out for the multiple heterologous protein production in filamentous fungus Thermothelomyces thermophilus F-859 (formerly Myceliophthora thermophila F-859). The engineered fungus Th. thermophilus was able to produce plant storage proteins of Zea mays (α-zein Z19) and Amaranthus hypochondriacus (albumin A1) to enrich fungal biomass by valuable nutritional proteins and improved amino acid content. The mRNA levels of z19 and a1 genes were significantly dependent on their driving promoters: the promoter of tryptophan synthase (PtrpC) was more efficient to express a1, while the promoter of translation elongation factor (Ptef) provided much higher levels of z19 transcript abundance. In general, the total recombinant proteins and amino acid contents were higher in the Ptef-containing clones. This work describes a new strategy to improve mycoprotein nutritive value by overexpression of plant storage proteins.
Project description:The aim of this study is to investigate the role of the transcription factor Xyr1 in the regulation of cellulolytic and xylanolytic genes of the filamantous fungus Myceliophthora thermophila