scholarly journals Disruption of alpha-tubulin releases carbon catabolite repression and enhances enzyme production in Trichoderma reesei even in the presence of glucose

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Nozomu Shibata ◽  
Hiroshi Kakeshita ◽  
Kazuaki Igarashi ◽  
Yasushi Takimura ◽  
Yosuke Shida ◽  
...  
Keyword(s):  
Trichoderma Reesei ◽  
Catabolite Repression ◽  
Enzyme Production ◽  
Protein Production ◽  
Carbon Catabolite Repression ◽  
Transcriptional Regulator ◽  
Secretory Protein ◽  
Sugar Transporter ◽  
Alpha Tubulin ◽  
High Enzyme

Abstract Background Trichoderma reesei is a filamentous fungus that is important as an industrial producer of cellulases and hemicellulases due to its high secretion of these enzymes and outstanding performance in industrial fermenters. However, the reduction of enzyme production caused by carbon catabolite repression (CCR) has long been a problem. Disruption of a typical transcriptional regulator, Cre1, does not sufficiently suppress this reduction in the presence of glucose. Results We found that deletion of an α-tubulin (tubB) in T. reesei enhanced both the amount and rate of secretory protein production. Also, the tubulin-disrupted (ΔtubB) strain had high enzyme production and the same enzyme profile even if the strain was cultured in a glucose-containing medium. From transcriptome analysis, the ΔtubB strain exhibited upregulation of both cellulase and hemicellulase genes including some that were not originally induced by cellulose. Moreover, cellobiose transporter genes and the other sugar transporter genes were highly upregulated, and simultaneous uptake of glucose and cellobiose was also observed in the ΔtubB strain. These results suggested that the ΔtubB strain was released from CCR. Conclusion Trichoderma reesei α-tubulin is involved in the transcription of cellulase and hemicellulase genes, as well as in CCR. This is the first report of overcoming CCR by disrupting α-tubulin gene in T. reesei. The disruption of α-tubulin is a promising approach for creating next-generation enzyme-producing strains of T. reesei.

scholarly journals Genetic Modification of Carbon Catabolite Repression in Trichoderma reesei for Improved Protein Production

2009 ◽  
Vol 75 (14) ◽  
pp. 4853-4860 ◽  
Author(s):  
Tiina Nakari-Setälä ◽  
Marja Paloheimo ◽  
Jarno Kallio ◽  
Jari Vehmaanperä ◽  
Merja Penttilä ◽  
...  
Keyword(s):  
Trichoderma Reesei ◽  
Catabolite Repression ◽  
Enzyme Production ◽  
Null Allele ◽  
Carbon Catabolite Repression ◽  
Regulatory Gene ◽  
Hydrolytic Enzymes ◽  
Strain Engineering ◽  
Mutant Variant ◽  
Mutant Strains

ABSTRACT The cellulase and hemicellulase genes of the filamentous fungus Trichoderma reesei have been shown to be under carbon catabolite repression mediated by the regulatory gene cre1. In this study, strains were constructed in which the cre1 gene was either completely removed or replaced by a truncated mutant variant, cre1-1, found previously in the Rut-C30 mutant strain with enhanced enzyme production capability. The T. reesei transformants with either deletion or truncation of cre1 had clearly altered colony morphology compared with the parental strains, forming smaller colonies and fewer aerial hyphae and spores. Liquid cultures in a medium with glucose as a carbon source showed that the transformants were derepressed in cellulase and hemicellulase production. Interestingly, they also produced significantly elevated levels of these hydrolytic enzymes in fermentations carried out in a medium inducing the hydrolase genes. This suggests that cre1 acts as a modulator of cellulase and hemicellulase gene expression under both noninducing and inducing conditions. There was no phenotypic difference between the Δcre1 and cre1-1 mutant strains in any of the experiments done, indicating that the cre1-1 gene is practically a null allele. The results of this work indicate that cre1 is a valid target gene in strain engineering for improved enzyme production in T. reesei.

Carbon catabolite repression of xylanase I (xyn1) gene expression in Trichoderma reesei

1996 ◽  
Vol 21 (6) ◽  
pp. 1273-1281 ◽  
Author(s):  
Robert L. Mach ◽  
Joseph Strauss ◽  
Susanne Zeilinger ◽  
Martin Schindler ◽  
Christian P. Kubicek
Keyword(s):  
Gene Expression ◽  
Trichoderma Reesei ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression

Defining the genome-wide role of CRE1 during carbon catabolite repression in Trichoderma reesei using RNA-Seq analysis

2014 ◽  
Vol 73 ◽  
pp. 93-103 ◽  
Author(s):  
Amanda Cristina Campos Antoniêto ◽  
Lílian dos Santos Castro ◽  
Rafael Silva-Rocha ◽  
Gabriela Felix Persinoti ◽  
Roberto Nascimento Silva
Keyword(s):  
Trichoderma Reesei ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Rna Seq ◽  
Genome Wide

scholarly journals Carbon catabolite repression involves physical interaction of the transcription factor CRE1/CreA and the Tup1–Cyc8 complex in Penicillium oxalicum and Trichoderma reesei

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Yueyan Hu ◽  
Mengxue Li ◽  
Zhongjiao Liu ◽  
Xin Song ◽  
Yinbo Qu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Filamentous Fungi ◽  
Transcriptional Repression ◽  
Catabolite Repression ◽  
Enzyme Production ◽  
Carbon Catabolite Repression ◽  
Penicillium Oxalicum ◽  
Physical Interaction ◽  
Corepressor Complex ◽  
Encoding Gene

Abstract Background Cellulolytic enzyme production in filamentous fungi requires a release from carbon catabolite repression (CCR). The protein CRE1/CreA (CRE = catabolite responsive element) is a key transcription factor (TF) that is involved in CCR and represses cellulolytic gene expression. CRE1/CreA represents the functional equivalent of Mig1p, an important Saccharomyces cerevisiae TF in CCR that exerts its repressive effect by recruiting a corepressor complex Tup1p–Cyc8p. Although it is known from S. cerevisiae that CRE1/CreA might repress gene expression via interacting with the corepressor complex Tup1–Cyc8, this mechanism is unconfirmed in other filamentous fungi, since the physical interaction has not yet been verified in these organisms. The precise mechanism on how CRE1/CreA achieves transcriptional repression after DNA binding remains unknown. Results The results from tandem affinity purification and bimolecular fluorescence complementation revealed a direct physical interaction between the TF CRE1/CreA and the complex Tup1–Cyc8 in the nucleus of cellulolytic fungus Trichoderma reesei and Penicillium oxalicum. Both fungi have the ability to secrete a complex arsenal of enzymes to synergistically degrade lignocellulosic materials. In P. oxalicum, the protein PoCyc8, a subunit of complex Tup1–Cyc8, interacts directly with TF PoCreA and histone H3 lysine 36 (H3K36) methyltransferase PoSet2 in the nucleus. The di-methylation level of H3K36 in the promoter of prominent cellulolytic genes (cellobiohydrolase-encoding gene Pocbh1/cel7A and endoglucanase-encoding gene Poegl1/cel7B) is positively correlated with the expression levels of TF PoCreA. Since the methylation of H3K36 was also demonstrated to be a repression marker of cellulolytic gene expression, it appears feasible that the cellulolytic genes are repressed via PoCreA-Tup1–Cyc8-Set2-mediated transcriptional repression. Conclusion This study verifies the long-standing conjecture that the TF CRE1/CreA represses gene expression by interacting with the corepressor complex Tup1–Cyc8 in filamentous fungi. A reasonable explanation is proposed that PoCreA represses gene expression by recruiting complex PoTup1–Cyc8. Histone methyltransferase Set2, which methylates H3K36, is also involved in the regulatory network by interacting with PoCyc8. The findings contribute to the understanding of CCR mechanism in filamentous fungi and could aid in biotechnologically relevant enzyme production.

scholarly journals Mutations in Catabolite Control Protein CcpA Separating Growth Effects from Catabolite Repression

1999 ◽  
Vol 181 (13) ◽  
pp. 4125-4128 ◽  
Author(s):  
Elke Küster ◽  
Tanja Hilbich ◽  
Michael K. Dahl ◽  
Wolfgang Hillen
Keyword(s):  
Growth Rate ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Transcriptional Regulator ◽  
Growth Effect ◽  
Single Amino Acid ◽  
Chromosomal Deletion ◽  
Regulatory Pathways ◽  
Amino Acid Mutations ◽  
Control Protein

ABSTRACT Carbon catabolite repression in Bacillus megaterium is mediated by the transcriptional regulator CcpA. A chromosomal deletion of ccpA eliminates catabolite repression and reduces the growth rate on glucose. We describe four single-amino-acid mutations in CcpA which separate the growth effect from catabolite repression, suggesting distinct regulatory pathways for these phenotypes.

scholarly journals Redesigning transcription factor Cre1 for alleviating carbon catabolite repression in Trichoderma reesei

2020 ◽  
Vol 5 (3) ◽  
pp. 230-235 ◽  
Author(s):  
Lijuan Han ◽  
Kuimei Liu ◽  
Wei Ma ◽  
Yi Jiang ◽  
Shaoli Hou ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Trichoderma Reesei ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression

scholarly journals Trichoderma reesei CRE1-mediated Carbon Catabolite Repression in Response to Sophorose Through RNA Sequencing Analysis

2016 ◽  
Vol 17 (2) ◽  
pp. 119-131 ◽  
Author(s):  
Amanda Cristina Campos Antoniêto ◽  
Renato Graciano de Paula ◽  
Lílian dos Santos Castro ◽  
Rafael Silva-Rocha ◽  
Gabriela Felix Persinoti ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Trichoderma Reesei ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Sequencing Analysis

scholarly journals Random and combinatorial mutagenesis for improved total production of secretory target protein in Escherichia coli

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
David Gonzalez-Perez ◽  
James Ratcliffe ◽  
Shu Khan Tan ◽  
Mary Chen May Wong ◽  
Yi Pei Yee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Protein Secretion ◽  
Protein Production ◽  
Secretory Protein ◽  
Target Protein ◽  
Carrier Protein ◽  
Signal Peptides ◽  
Carrier Proteins ◽  
E Coli ◽  
Combinatorial Mutagenesis

AbstractSignal peptides and secretory carrier proteins are commonly used to secrete heterologous recombinant protein in Gram-negative bacteria. The Escherichia coli osmotically-inducible protein Y (OsmY) is a carrier protein that secretes a target protein extracellularly, and we have previously applied it in the Bacterial Extracellular Protein Secretion System (BENNY) to accelerate directed evolution. In this study, we reported the first application of random and combinatorial mutagenesis on a carrier protein to enhance total secretory target protein production. After one round of random mutagenesis followed by combining the mutations found, OsmY(M3) (L6P, V43A, S154R, V191E) was identified as the best carrier protein. OsmY(M3) produced 3.1 ± 0.3 fold and 2.9 ± 0.8 fold more secretory Tfu0937 β-glucosidase than its wildtype counterpart in E. coli strains BL21(DE3) and C41(DE3), respectively. OsmY(M3) also produced more secretory Tfu0937 at different cultivation temperatures (37 °C, 30 °C and 25 °C) compared to the wildtype. Subcellular fractionation of the expressed protein confirmed the essential role of OsmY in protein secretion. Up to 80.8 ± 12.2% of total soluble protein was secreted after 15 h of cultivation. When fused to a red fluorescent protein or a lipase from Bacillus subtillis, OsmY(M3) also produced more secretory protein compared to the wildtype. In this study, OsmY(M3) variant improved the extracellular production of three proteins originating from diverse organisms and with diverse properties, clearly demonstrating its wide-ranging applications. The use of random and combinatorial mutagenesis on the carrier protein demonstrated in this work can also be further extended to evolve other signal peptides or carrier proteins for secretory protein production in E. coli.

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