Project description:We have examined and compared the transcriptome of T. reesei growing on wheat straw and lactose as carbon sources under otherwise similar conditions. Gene expression on wheat straw exceeded that on lactose, and 1619 genes were found to be only induced on wheat straw but not on lactose. They comprised 30 % of the CAZome, but were also enriched in genes associated with phospholipid metabolism, DNA synthesis and repair and iron homeostatis. Two thirds of the CAZome was expressed both on wheat straw as well as on lactose, but 60 % of it at least >2-fold higher on the former. Major wheat straw specific genes comprised xylanases, chitinases and M-CM-^_-mannosidases. Interestingly, the latter two CAZyme families were significantly higher expressed in a strain in which xyr1 encoding the major regulator of cellulase and hemicellulase biosynthesis is non-functional, demonstrating that XYR1 is a repressor of these genes. We used two biological replicas of four T. reesei strains growing on glucose, lactose, and on wheat straw
Project description:We have examined and compared the transcriptome of T. reesei growing on wheat straw and lactose as carbon sources under otherwise similar conditions. Gene expression on wheat straw exceeded that on lactose, and 1619 genes were found to be only induced on wheat straw but not on lactose. They comprised 30 % of the CAZome, but were also enriched in genes associated with phospholipid metabolism, DNA synthesis and repair and iron homeostatis. Two thirds of the CAZome was expressed both on wheat straw as well as on lactose, but 60 % of it at least >2-fold higher on the former. Major wheat straw specific genes comprised xylanases, chitinases and ß-mannosidases. Interestingly, the latter two CAZyme families were significantly higher expressed in a strain in which xyr1 encoding the major regulator of cellulase and hemicellulase biosynthesis is non-functional, demonstrating that XYR1 is a repressor of these genes.
Project description:Lactose (1,4-0-M-CM-^_-d-galactopyranosyl-d-glucose), a by-product from cheese manufacture or whey processing industries, is known to induce the formation of plant biomass hydrolyzing enzymes needed for the biorefinery industry in the fungus Trichoderma reesei, but the reason for this induction and the underlying mechanism are not fully understood. Here, we used systems analysis of the Trichoderma reesei transcriptome during utilization of lactose. We found that the respective CAZome encoded glycosyl hydrolases specifically tailored for the attack of monocotyledon xyloglucan. In addition, genes for a high number of putative transporters of the major facilitator superfamily were also induced. Systematic knock out of them identified a gene whose knock-out completely impaired lactose utilization and cellulase induction in Trichoderma reesei. These data shed new light on the mechanism by which Trichoderma reesei metabolizes lactose and illuminate the key role of M-CM-^_-D-galactosides in habitat specificity of this fungus. We used two biological replicas of Trichoderma reesei growing on lactose, glucose and glycerol
Project description:Lactose (1,4-0-ß-d-galactopyranosyl-d-glucose), a by-product from cheese manufacture or whey processing industries, is known to induce the formation of plant biomass hydrolyzing enzymes needed for the biorefinery industry in the fungus Trichoderma reesei, but the reason for this induction and the underlying mechanism are not fully understood. Here, we used systems analysis of the Trichoderma reesei transcriptome during utilization of lactose. We found that the respective CAZome encoded glycosyl hydrolases specifically tailored for the attack of monocotyledon xyloglucan. In addition, genes for a high number of putative transporters of the major facilitator superfamily were also induced. Systematic knock out of them identified a gene whose knock-out completely impaired lactose utilization and cellulase induction in Trichoderma reesei. These data shed new light on the mechanism by which Trichoderma reesei metabolizes lactose and illuminate the key role of ß-D-galactosides in habitat specificity of this fungus.
Project description:The filamentous fungus Trichoderma reesei is the main industrial cellulytic enzymes producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome sequencing, transcriptome) can shed light on the mechanisms of cellulase production and help identify genes linked to performance. Previously, our group published comparative genome sequencing of the lower and higher producer strains NG14 and RUT C30. In this follow-up work, we examined how these mutations affect phenotype at the level of the transcriptome and cultivation behavior. We performed kinetic transcriptome analysis of the NG14 and RUT C30 strains of early cellulase production induced by lactose using bioreactor cultivations close to industrial conditions. RUT C30 exhibited both earlier onset of cellulase production and higher steady-state productivity. A rather low number of genes were regulated, most of them being specific to the NG14 strains. Clustering of these genes highlighted similar behavior for some functional categories, and allowed to distinguish between induction-related genes and productivity-related genes. The lower number of genes regulated in RUT C30 could not formally be linked to the relief in catabolic repression that is characteristic of this strain. Cross-comparison of our transcriptome data with mutations previously identified revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting these clusters contain genes playing a role in strain performance. This is the first kinetic transcriptome study carried out in industry-relevant conditions with two related strains of T. reesei showing distinctive performances. Our study sheds some light on some of the events occurring in these strains following induction by lactose. The fact that few regulated genes have been affected by mutagenesis suggest that the induction mechanism is essentially intact and that there is room for further improvement of T. reesei. We also provide some potential target for further genetic improvement of these strains.
Project description:The filamentous fungus Trichoderma reesei is the main industrial cellulytic enzymes producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome sequencing, transcriptome) can shed light on the mechanisms of cellulase production and help identify genes linked to performance. Previously, our group published comparative genome sequencing of the lower and higher producer strains NG14 and RUTM-BM- C30. In this follow-up work, we examined how these mutations affect phenotype at the level of the transcriptome and cultivation behavior.M-BM- We performed kinetic transcriptome analysis of the NG14 and RUTM-BM- C30 strains of early cellulase production induced by lactose using bioreactor cultivations close to industrial conditions. RUTM-BM- C30 exhibited both earlier onset of cellulase production and higher steady-state productivity. A rather lowM-BM- number of genes were regulated, most of them being specific to the NG14 strains. Clustering of these genes highlighted similar behavior for some functional categories, and allowed to distinguish between induction-related genes and productivity-related genes. The lower number of genes regulated in RUTM-BM- C30 could not formally be linked to the relief in catabolic repression that is characteristic of this strain. Cross-comparison of our transcriptome data with mutations previously identified revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting these clusters contain genes playing a role in strain performance. This is the first kinetic transcriptome study carried out in industry-relevant conditions with two related strains of T. reesei showing distinctive performances. Our study sheds some light on some of the events occurring in these strains following induction by lactose. The fact that few regulated genes have been affected by mutagenesis suggest that the induction mechanism is essentially intact and that there is room for further improvement of T. reesei. We also provide some potential target for further genetic improvement of these strains. Two biological pool by condition in dye switch. For the two biological replicates on each four experiments we apply on the pretreated results the linear modeling approach implemented by lmFit and the empirical Bayes statistics implemented by eBayes from the limma R package (Smyth 2004). We select the list of statistically regulated genes using a 5% significance threshold.
Project description:The ascomycete Trichoderma reesei is an industrial producer of cellulolytic and hemicellulolytic enzymes and also serves as a model for investigations on these enzymes and their genes. The strain QM9978 has a cellulase negative phenotype and therefore presents a valuable tool for understanding the mechanisms underlying cellulase regulation. A transcriptomic analyses of the cellulase negative strain QM9978 and the original strain QM6a have been performed to identify the genetic differences between QM6a and QM9978 leading to the cellulase-negative phenotype
Project description:The ascomycete Trichoderma reesei is one of the most well studied cellulolytic fungi and widely used in the biotechnology industry, as in the production of second-generation bioethanol. Carbon catabolite repression (CCR) mechanism adopted by T. reesei is mediated by the transcription factor CRE1 and consists in the repression of genes related to the production of cellulase when a readily available carbon source is present in the medium. Using RNA sequencing this study aims to contribute to understanding of CCR during growth in cellulose and glucose, by comparing the mutant strain of T. reesei Δcre1 with its parental, QM9414.
Project description:Systemic analysis of lactose metabolism in Trichoderma reesei identifies a lactose permease that is essential for cellulase induction
Project description:The mitosporic fungus Trichoderma reesei is an industrial producer of enzymes for degradation of lignocellulosic polysaccharides to soluble monomers that can be fermented to biofuels. The genes encoding these enzymes in T. reesei have recently been shown to be clustered in the genome. Here we will show that the expression of these genes is epigenetically controlled at the heterochromatin level by the protein methyltransferase LAE1. Deletion of lae1 led to a loss of expression of the major cellulase and hemicellulase encoding genes, and resulted in an inability to grow on cellulose. The cellulase null phenotype was also seen with known soluble inducers of enzymes active on cellulose. In contrast, introduction of a second copy of lae1 or its enhanced expression under a strong constitutive promoter resulted in increased levels of cellulases. Thus, our data provides an experiment-based explanation for the advantage for clustering of cellulases in the genome of T. reesei, and imply that the heterochromatin structure is a major determinant of cellulase gene expression and hence an attractive target for strain improvement. We used two biological replicas of four T. reesei strains growing on lactose, the parent strain (QM9414), a delta-lae1, and two overexpressing strains (tef1:lae1 mutant 1 and mutant 2).