The transcription factor ACE3 controls cellulase activities and lactose metabolism via two additional regulators in the fungus Trichoderma reesei

ABSTRACTThe industrially used ascomyceteTrichoderma reeseisecretes a typical yellow pigment during cultivation, while otherTrichodermaspecies do not. A comparative genomic analysis suggested that a putative secondary metabolism cluster, containing two polyketide-synthase encoding genes, is responsible for the yellow pigment synthesis. This cluster is conserved in a set of rather distantly related fungi, includingAcremonium chrysogenumandPenicillium chrysogenum. In an attempt to silence the cluster inT. reesei, two genes of the cluster encoding transcription factors were individually deleted. For a complete genetic proof-of-function, the genes were reinserted into the genomes of the respective deletion strains. The deletion of the first transcription factor (termed yellow pigment regulator 1 [Ypr1]) resulted in the full abolishment of the yellow pigment formation and the expression of most genes of this cluster. A comparative high-pressure liquid chromatography (HPLC) analysis of supernatants of theypr1deletion and its parent strain suggested the presence of several yellow compounds inT. reeseithat are all derived from the same cluster. A subsequent gas chromatography/mass spectrometry analysis strongly indicated the presence of sorbicillin in the major HPLC peak. The presence of the second transcription factor, termed yellow pigment regulator 2 (Ypr2), reduces the yellow pigment formation and the expression of most cluster genes, including the gene encoding the activator Ypr1.IMPORTANCETrichoderma reeseiis used for industry-scale production of carbohydrate-active enzymes. During growth, it secretes a typical yellow pigment. This is not favorable for industrial enzyme production because it makes the downstream process more complicated and thus increases operating costs. In this study, we demonstrate which regulators influence the synthesis of the yellow pigment. Based on these data, we also provide indication as to which genes are under the control of these regulators and are finally responsible for the biosynthesis of the yellow pigment. These genes are organized in a cluster that is also found in other industrially relevant fungi, such as the two antibiotic producersPenicillium chrysogenumandAcremonium chrysogenum. The targeted manipulation of a secondary metabolism cluster is an important option for any biotechnologically applied microorganism.

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A novel transcription factor specifically regulates GH11 xylanase genes in Trichoderma reesei

Biotechnology for Biofuels ◽

10.1186/s13068-017-0878-x ◽

2017 ◽

Vol 10 (1) ◽

Cited By ~ 14

Author(s):

Rui Liu ◽

Ling Chen ◽

Yanping Jiang ◽

Gen Zou ◽

Zhihua Zhou

Keyword(s):

Transcription Factor ◽

Trichoderma Reesei

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Understanding the Role of Trichoderma reesei Vib1 in Gene Expression during Cellulose Degradation

Journal of Fungi ◽

10.3390/jof7080613 ◽

2021 ◽

Vol 7 (8) ◽

pp. 613

Author(s):

Xiuzhen Chen ◽

Bingran Song ◽

Minglu Liu ◽

Lina Qin ◽

Zhiyang Dong

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Trichoderma Reesei ◽

Cellulose Degradation ◽

Strain Improvement ◽

Cellulase Production ◽

Oxidoreductase Activity ◽

Expression Of Genes ◽

Genes Encoding ◽

Almost All

Vib1, a member of the Ndt80/PhoG-like transcription factor family, has been shown to be essential for cellulase production of Trichoderma reesei. Here, we combined transcriptomic and genetic analyses to gain mechanistic insights into the roles of Vib1 during cellulose degradation. Our transcriptome analysis showed that the vib1 deletion caused 586 genes with decreased expression and 431 genes with increased expression on cellulose. The downregulated genes were enriched for Gene Ontology terms associated with carbohydrate metabolism, transmembrane transport, oxidoreductase activity, and transcription factor activity. Of the 258 genes induced by cellulose, 229 showed no or decreased expression in Δvib1 on cellulose, including almost all (hemi)cellulase genes, crucial sugar transporter genes (IDs:69957, 3405), and the genes encoding main transcriptional activators Xyr1 and Ace3. Additionally, Vib1 also regulated the expression of genes involved in secondary metabolism. Further comparison of the transcriptomes of Δvib1 and Δxyr1 in cellulose revealed that the genes regulated by Vib1 had much overlap with Xyr1 targets especially for the gene set induced by cellulose, presumably whose expression requires the cooperativity between Vib1 and Xyr1. Genetic evidence indicated that Vib1 regulates cellulase gene expression partially via Xyr1. Our results will provide new clues for strain improvement.

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ACE4, a novel transcriptional activator involved in the regulation of cellulase genes on cellulose in Trichoderma reesei

Applied and Environmental Microbiology ◽

10.1128/aem.00593-21 ◽

2021 ◽

Author(s):

Yumeng Chen ◽

Aibo Lin ◽

Pei Liu ◽

Xingjia Fan ◽

Chuan Wu ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Transcription Factors ◽

Trichoderma Reesei ◽

Transcriptional Activator ◽

Cellulase Production ◽

Comparative Genomic ◽

Transcriptional Level ◽

Cellulase Gene ◽

Cellulase Expression

The filamentous fungus Trichoderma reesei is a model strain for cellulase production. Cellulase gene expression in T. reesei is controlled by multiple transcription factors. Here, we identified by comparative genomic screening a novel transcriptional activator ACE4 ( A ctivator of c ellulase e xpression 4) that positively regulates cellulase gene expression on cellulose in T. reesei . Disruption of the ace4 gene significantly decreased expression of four main cellulase genes, and the essential cellulase transcription factor encoding gene ace3 . Overexpression of ace4 increased cellulase production by approximately 22% compared to that in the parental strain. Further investigations using electrophoretic mobility shift assays, DNase I footprinting assays, and chromatin immunoprecipitation assays indicated that ACE4 directly binds to the promoter of cellulase genes by recognizing the two adjacent 5′-GGCC-3′ sequences. Additionally, ACE4 directly binds to the promoter of ace3 and, in turn, regulates the expression of ACE3 to facilitate cellulase production. Collectively, these results demonstrate an important role for ACE4 in regulating cellulase gene expression, which will contribute to understanding the mechanism underlying cellulase expression in T. reesei . IMPORTANCE T. reesei is commonly utilized in industry to produce cellulases, enzymes that degrade lignocellulosic biomass for the production of bioethanol and bio-based products. T. reesei is capable of rapidly initiating the biosynthesis of cellulases in the presence of cellulose, which has made it useful as a model fungus for studying gene expression in eukaryotes. Cellulase gene expression is controlled through multiple transcription factors at the transcriptional level. However, the molecular mechanisms by which transcription is controlled remain unclear. In the present study, we identified a novel transcription factor, ACE4, which regulates cellulase expression on cellulose by binding to the promoters of cellulase genes and the cellulase activator ace3 . Our study not only expands the general functional understanding of the novel transcription factor ACE4 but also provides evidence for the regulatory mechanism mediating gene expression in T. reesei .

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Combined strategy of transcription factor manipulation and β-glucosidase gene overexpression in Trichoderma reesei and its application in lignocellulose bioconversion

Journal of Industrial Microbiology & Biotechnology ◽

10.1007/s10295-018-2041-5 ◽

2018 ◽

Vol 45 (9) ◽

pp. 803-811 ◽

Cited By ~ 2

Author(s):

Ying Xia ◽

Lirong Yang ◽

Liming Xia

Keyword(s):

Transcription Factor ◽

Trichoderma Reesei ◽

Gene Overexpression ◽

Combined Strategy

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Systems Analysis of Lactose Metabolism in Trichoderma reesei Identifies a Lactose Permease That Is Essential for Cellulase Induction

PLoS ONE ◽

10.1371/journal.pone.0062631 ◽

2013 ◽

Vol 8 (5) ◽

pp. e62631 ◽

Cited By ~ 82

Author(s):

Christa Ivanova ◽

Jenny A. Bååth ◽

Bernhard Seiboth ◽

Christian P. Kubicek

Keyword(s):

Trichoderma Reesei ◽

Systems Analysis ◽

Lactose Permease ◽

Cellulase Induction ◽

Lactose Metabolism

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Precision engineering of the transcription factor Cre1 in Hypocrea jecorina (Trichoderma reesei) for efficient cellulase production in the presence of glucose

10.1101/2020.03.08.982249 ◽

2020 ◽

Author(s):

Lijuan Han ◽

Yinshuang Tan ◽

Wei Ma ◽

Kangle Niu ◽

Shaoli Hou ◽

...

Keyword(s):

Transcription Factor ◽

Trichoderma Reesei ◽

Parent Strain ◽

Zinc Finger Protein ◽

Cellulase Production ◽

Fungal Species ◽

Cellulolytic Enzymes ◽

Phosphorylation Sites ◽

Precision Engineering ◽

C Terminus

SummaryIn Trichoderma reesei, carbon catabolite repression (CCR) significantly downregulates the transcription of cellulolytic enzymes, which is usually mediated by the zinc finger protein Cre1. It was found that there is a conserved region at the C-terminus of Cre1/CreA in several cellulase-producing fungi that contains up to three continuous S/T phosphorylation sites. Here, S387, S388, T389, and T390 at the C-terminus of Cre1 in T. reesei were mutated to valine for mimicking an unphosphorylated state, thereby generating the transformants Tr_Cre1S387V, Tr_Cre1S388V, Tr_Cre1T389V, and Tr_Cre1T390V, respectively. Transcription of cel7a in Tr_ Cre1S388V was markedly higher than that of the parent strain when grown in glucose-containing media. Under these conditions, both filter paperase (FPase) and p-nitrophenyl-β-D-cellobioside (pNPCase) activities, as well as soluble proteins from Tr_Cre1S388V were significantly increased by up to 2- to 3-fold compared with that of other transformants and the parent strain. To our knowledge, this is the first report demonstrating an improvement of cellulase production in fungal species under CCR by mimicking dephosphorylation at the C-terminus of Cre1. Taken together, we developed a precision engineering strategy based on the modification of phosphorylation sites of Cre1 transcription factor to enhance the production of cellulase in fungal species under CCR.

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