Loss of Aspergillus oryzae amyR function indirectly affects hemicellulolytic and cellulolytic enzyme production

Watanabe, Jun; Tanaka, Hisaki; Mogi, Yoshinobu; Yamazaki, Tatsuo; Suzuki, Kazushi; Watanabe, Takeshi; Akita, Osamu

doi:10.1016/j.jbiosc.2010.12.006

Cited by 24 publications

(12 citation statements)

References 23 publications

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“…Therefore, P . oxalicum amyR (PDE_03964), an Aspergillus oryzae amyR homolog [ 41 ], was considered tightly associated with cellulolytic enzyme production. The strain with the deletion of amyR exhibited visible varying halos on cellulose and starch plates, as well as an identical phenotype on glucose relative to its parental strain ( Fig 9A ).…”

Section: Resultsmentioning

confidence: 99%

Synergistic and Dose-Controlled Regulation of Cellulase Gene Expression in Penicillium oxalicum

et al. 2015

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Filamentous fungus Penicillium oxalicum produces diverse lignocellulolytic enzymes, which are regulated by the combinations of many transcription factors. Here, a single-gene disruptant library for 470 transcription factors was constructed and systematically screened for cellulase production. Twenty transcription factors (including ClrB, CreA, XlnR, Ace1, AmyR, and 15 unknown proteins) were identified to play putative roles in the activation or repression of cellulase synthesis. Most of these regulators have not been characterized in any fungi before. We identified the ClrB, CreA, XlnR, and AmyR transcription factors as critical dose-dependent regulators of cellulase expression, the core regulons of which were identified by analyzing several transcriptomes and/or secretomes. Synergistic and additive modes of combinatorial control of each cellulase gene by these regulatory factors were achieved, and cellulase expression was fine-tuned in a proper and controlled manner. With one of these targets, the expression of the major intracellular β-glucosidase Bgl2 was found to be dependent on ClrB. The Bgl2-deficient background resulted in a substantial gene activation by ClrB and proved to be closely correlated with the relief of repression mediated by CreA and AmyR during cellulase induction. Our results also signify that probing the synergistic and dose-controlled regulation mechanisms of cellulolytic regulators and using it for reconstruction of expression regulation network (RERN) may be a promising strategy for cellulolytic fungi to develop enzyme hyper-producers. Based on our data, ClrB was identified as focal point for the synergistic activation regulation of cellulase expression by integrating cellulolytic regulators and their target genes, which refined our understanding of transcriptional-regulatory network as a “seesaw model” in which the coordinated regulation of cellulolytic genes is established by counteracting activators and repressors.

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Section: Resultsmentioning

confidence: 99%

Synergistic and Dose-Controlled Regulation of Cellulase Gene Expression in Penicillium oxalicum

et al. 2015

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“…In A. oryzae , mutation in amyR causes down-regulation of amylases and up-regulation of cellulolytic enzymes only in the presence of starch, and a CCR-involved mechanism is proposed to account for the increased cellulolytic enzymes29. A CCR-involved mechanism is logic as AmyR activates the synthesis of amylases that convert starch to glucose, and the latter inhibits the production of cellulolytic enzymes through CCR.…”

Section: Discussionmentioning

confidence: 99%

Long-term strain improvements accumulate mutations in regulatory elements responsible for hyper-production of cellulolytic enzymes

Liu

Zhang

Qin

et al. 2013

Sci Rep

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Long-term strain improvements through repeated mutagenesis and screening have generated a hyper-producer of cellulases and hemicellulases from Penicillium decumbens 114 which was isolated 30 years ago. Here, the genome of the hyper-producer P. decumbens JU-A10-T was sequenced and compared with that of the wild-type strain 114-2. Further, the transcriptomes and secretomes were compared between the strains. Selective hyper-production of cellulases and hemicellulases but not all the secreted proteins was observed in the mutant, making it a more specific producer of lignocellulolytic enzymes. Functional analysis identified that changes in several transcriptional regulatory elements played crucial roles in the cellulase hyper-producing characteristics of the mutant. Additionally, the mutant showed enhanced supply of amino acids and decreased synthesis of secondary metabolites compared with the wild-type. The results clearly point out that we can target gene regulators and promoters with minimal alterations of the genetic content but maximal effects in genetic engineering.

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“…As the main agents of plant decay in the environment, filamentous fungi such as Trichoderma reesei [2, 3], Aspergillus spp. [4, 5], and Neurospora crassa [6, 7] can express a complex mixture of enzymes to synergistically deconstruct the polysaccharides in plant cell walls. Commercial biofuel production remains hindered by the high cost of enzyme production for biomass conversion [8].…”

Section: Introductionmentioning

confidence: 99%

Combining manipulation of transcription factors and overexpression of the target genes to enhance lignocellulolytic enzyme production in Penicillium oxalicum

Gao

Xia

et al. 2017

Biotechnol Biofuels

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BackgroundLignocellulolytic enzymes are the main enzymes to saccharify lignocellulose from renewable plant biomass in the bio-based economy. The production of these enzymes is transcriptionally regulated by multiple transcription factors. We previously engineered Penicillium oxalicum for improved cellulase production via manipulation of three genes in the cellulase expression regulatory network. However, the potential of combinational engineering of multiple regulators and their targets at protein abundance and activity levels has not been fully explored.ResultsHere, we verified that a point mutation XlnRA871V in transcription factor XlnR enhanced the expression of lignocellulolytic enzymes, particularly hemicellulases, in P. oxalicum. Then, overexpression of XlnRA871V with a constitutive PDE_02864 promoter was combined with the overexpression of cellulase transcriptional activator ClrB and deletion of carbon catabolite repressor CreA. The resulted strain RE-7 showed 8.9- and 51.5-fold increased production of cellulase and xylanase relative to the starting strain M12, respectively. Further overexpression of two major cellulase genes cbh1-2 and eg1 enabled an additional 13.0% improvement of cellulase production. In addition, XlnRA871V led to decreased production of β-glucosidase and amylase, which could be attributed to the reduced transcription of corresponding enzyme-encoding genes.ConclusionsThe results illustrated that combinational manipulation of the involved transcription factors and their target genes was a viable strategy for efficient production of lignocellulolytic enzymes in filamentous fungi. The striking negative effect of XlnRA871V mutation on amylase production was also highlighted.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-017-0783-3) contains supplementary material, which is available to authorized users.

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Loss of Aspergillus oryzae amyR function indirectly affects hemicellulolytic and cellulolytic enzyme production

Cited by 24 publications

References 23 publications

Synergistic and Dose-Controlled Regulation of Cellulase Gene Expression in Penicillium oxalicum

Synergistic and Dose-Controlled Regulation of Cellulase Gene Expression in Penicillium oxalicum

Long-term strain improvements accumulate mutations in regulatory elements responsible for hyper-production of cellulolytic enzymes

Combining manipulation of transcription factors and overexpression of the target genes to enhance lignocellulolytic enzyme production in Penicillium oxalicum

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