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The putative ?-glucosidase BGL3I regulates cellulase induction in Trichoderma reesei.

ABSTRACT: Background:The filamentous fungus Trichoderma reesei (anamorph of Hypocrea jecorina) displays increased cellulase expression while growing on inducers such as lactose or cellulose. However, the mechanism of cellulase induction in T. reesei is not yet completely characterized. Here, a protein annotated as ?-glucosidase (BGL3I) was found to be involved in cellulase induction in T. reesei. The effects of BGL3I on cellulase production have not yet been fully understood. Results:Deletion of the bgl3i gene had no influence on the growth of T. reesei, but significantly increased its cellulase activities. Deletion of bgl3i also resulted in decreased extracellular galactosidase activity, but significantly increased transcription of lactose permeases, which might be involved in lactose transport. Furthermore, deletion of bgl3i enhanced the transcription levels of intracellular ?-glucosidases cel1a, cel1b and the regulator xyr1, which are all essential for lactose induction in T. reesei. BGL3I was found to have a relatively high ability to hydrolyze sophorose, which is proposed to be the strongest natural inducer of cellulase synthesis in T. reesei. Conclusions:BGL3I may take part in the complex regulating system of cellulase induction. The deletion of bgl3i offers a new strategy to improve T. reesei strain performance.

SUBMITTER: Zou G

PROVIDER: S-EPMC6240962 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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The putative β-glucosidase BGL3I regulates cellulase induction in Trichoderma reesei.

Zou Gen G Jiang Yanping Y Liu Rui R Zhu Zhihua Z Zhou Zhihua Z

Biotechnology for biofuels 20181119

<h4>Background</h4>The filamentous fungus Trichoderma reesei (anamorph of Hypocrea jecorina) displays increased cellulase expression while growing on inducers such as lactose or cellulose. However, the mechanism of cellulase induction in T. reesei is not yet completely characterized. Here, a protein annotated as β-glucosidase (BGL3I) was found to be involved in cellulase induction in T. reesei. The effects of BGL3I on cellulase production have not yet been fully under ...[more]

PMID: 30473732

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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:BackgroundThe filamentous fungus Trichoderma reesei (anamorph of Hypocrea jecorina) produces increased cellulase expression when grown on cellulose or its derivatives as a sole carbon source. It has been believed that β-glucosidases of T. reesei not only metabolize cellobiose but also contribute in the production of inducers of cellulase gene expression by their transglycosylation activity. The cellulase hyper-producing mutant PC-3-7 developed in Japan has enhanced cellulase production ability when cellobiose is used as the inducer. The comparative genomics analysis of PC-3-7 and its parent revealed a single-nucleotide mutation within the bgl2 gene encoding intracellular β-glucosidase II (BGLII/Cel1a), giving rise to an amino acid substitution in PC-3-7, which could potentially account for the enhanced cellulase expression when these strains are cultivated on cellulose and cellobiose.ResultsTo analyze the effects of the BGLII mutation in cellulase induction, we constructed both a bgl2 revertant and a disruptant. Enzymatic analysis of the transformant lysates showed that the strain expressing mutant BGLII exhibited weakened cellobiose hydrolytic activity, but produced some transglycosylation products, suggesting that the SNP in bgl2 strongly diminished cellobiase activity, but did not result in complete loss of function of BGLII. The analysis of the recombinant BGLII revealed that transglycosylation products might be oligosaccharides, composed probably of glucose linked β-1,4, β-1,3, or a mixture of both. PC-3-7 revertants of bgl2 exhibited reduced expression and inducibility of cellulase during growth on cellulose and cellobiose substrates. Furthermore, the effect of this bgl2 mutation was reproduced in the common strain QM9414 in which the transformants showed cellulase production comparable to that of PC-3-7.ConclusionWe conclude that BGLII plays an important role in cellulase induction in T. reesei and that the bgl2 mutation in PC-3-7 brought about enhanced cellulase expression on cellobiose. The results of the investigation using PC-3-7 suggested that other mutation(s) in PC-3-7 could also contribute to cellulase induction. Further investigation is essential to unravel the mechanism responsible for cellulase induction in T. reesei.

Project description:BackgroundThe filamentous fungus Trichoderma reesei is the main industrial cellulolytic enzyme 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 and transcriptome sequencing) 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 NG 14 and RUT C30. In this follow-up work, we examine how these mutations affect phenotype as regards the transcriptome and cultivation behaviour.ResultsWe performed kinetic transcriptome analysis of the NG 14 and RUT C30 strains of early enzyme production induced by lactose using bioreactor cultivations close to an industrial cultivation regime. RUT C30 exhibited both earlier onset of protein production (3 h) and higher steady-state productivity. A rather small number of genes compared to previous studies were regulated (568), most of them being specific to the NG 14 strain (319). Clustering analysis highlighted similar behaviour for some functional categories and allowed us to distinguish between induction-related genes and productivity-related genes. Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting that these clusters contain genes playing a role in strain performance.ConclusionsThis is the first kinetic analysis of a transcriptomic study carried out under conditions approaching industrial ones with two related strains of T. reesei showing distinctive cultivation behaviour. 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 suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei. Genes from two specific clusters might be potential targets for such genetic engineering.

Dataset Information

The putative ?-glucosidase BGL3I regulates cellulase induction in Trichoderma reesei.

Publications

The putative β-glucosidase BGL3I regulates cellulase induction in <i>Trichoderma reesei</i>.

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