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Effects of manR disruption and overexpression on Aspergillus oryzae gene expression

ABSTRACT: Fungal endo-β-mannanases (β-mannanases) are widely used as industrial enzymes; however, no transcriptional regulator of β-mannanases has been identified in fungi or other eukaryotic cells to date. To identify a transcriptional regulator of β-mannanases in Aspergillus oryzae, a gene-disruptant library of transcriptional regulators was screened for mutants exhibiting reduced β-mannanase activity by using konjac glucomannan as the substrate, and ManR, a Zn(II)2Cys6 type DNA binding protein was identified. Moreover, a manR-overexpressing strain showed significantly increased β-mannanase activity. DNA microarray analysis of the manR-disruptant strain and the manR-overexpressing strain further indicated that when konjac glucomannan is used as the carbon source, ManR positively regulates the gene expression of not only β-mannanase, but also the enzymes involved in the degradation of galactomannans and glucomannans such as α-galactosidase, β-mannosidase, acetylmannan esterase, and β-glucosidase. Therefore, we conclude that ManR is a positive regulator of the β-mannan utilization system in A. oryzae.

ORGANISM(S): Aspergillus oryzae RIB40

PROVIDER: GSE40590 | GEO | 2012/11/08

SECONDARY ACCESSION(S): PRJNA174354

REPOSITORIES: GEO

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Effects of manR disruption and overexpression on Aspergillus oryzae gene expression

Project description:Fungal endo-M-NM-2-mannanases (M-NM-2-mannanases) are widely used as industrial enzymes; however, no transcriptional regulator of M-NM-2-mannanases has been identified in fungi or other eukaryotic cells to date. To identify a transcriptional regulator of M-NM-2-mannanases in Aspergillus oryzae, a gene-disruptant library of transcriptional regulators was screened for mutants exhibiting reduced M-NM-2-mannanase activity by using konjac glucomannan as the substrate, and ManR, a Zn(II)2Cys6 type DNA binding protein was identified. Moreover, a manR-overexpressing strain showed significantly increased M-NM-2-mannanase activity. DNA microarray analysis of the manR-disruptant strain and the manR-overexpressing strain further indicated that when konjac glucomannan is used as the carbon source, ManR positively regulates the gene expression of not only M-NM-2-mannanase, but also the enzymes involved in the degradation of galactomannans and glucomannans such as M-NM-1-galactosidase, M-NM-2-mannosidase, acetylmannan esterase, and M-NM-2-glucosidase. Therefore, we conclude that ManR is a positive regulator of the M-NM-2-mannan utilization system in A. oryzae. manR disruptant, manR-overexpressing strain and A. oryzae RkuptrP2-1M-bM-^HM-^FAF/P (derivative of A. oryzae RIB40) were cultivated in minimal medium containing 1% konjac glucomannan as the sole carbon source. After 6 h cultivation, total RNAs from the mycelia were extracted, and DNA microarray analysis was carried out. The analysis of manR disruptant was conducted with 4 biological replications, the analysis of manR overexpressing strain was conducted with 3 biological replications.

2012-11-08 | E-GEOD-40590 | biostudies-arrayexpress

Expression profiling of Cellvibrio japonicus using plant mannans

Project description:Plant mannans are a component of lignocellulose that may possess diverse compositions changing both in terms of its backbone and side-chain substitutions. Consequently, the degradation of mannan substrates requires a cadre of enzymes for complete reduction to substituent monosaccharides, specifically mannose, galactose, and/or glucose. One bacterium that possesses this suite of enzymes is the Gram-negative saprophyte Cellvibrio japonicus, which has ten predicted mannanases from the Glycoside Hydrolase (GH) families 5, 26, and 27. Here we describe a systems biology approach to identify and characterize the essential components of the mannan degradation apparatus in this bacterium. Transcriptomic analysis uncovered significant changes in gene expression for most mannanases, as well as many genes that encode Carbohydrate Active Enzymes (CAZymes) when mannan substrates were actively being degraded. A comprehensive mutational analysis characterized 54 CAZyme genes in the context of mannan utilization. Growth analysis of the mutant strains indicated that the man26C, aga27A, and man5D genes, which encode a mannobiohydrolase, α- galactosidase, and mannosidase, respectively, were influential to the deconstruction of galactomannan. Furthermore, our updated model of mannan degradation in C. japonicus proposes that the removal of galactose sidechains from substituted mannans constitutes a crucial step for the efficient and complete degradation of this hemicellulose by bacteria.

2022-06-28 | GSE206864 | GEO

Protein production by Aspergillus oryzae

Project description:The filamentous fungus Aspergillus oryzae is an important microbial cell factory for industrial production of useful enzymes, such as α-amylase. In order to optimize the industrial enzyme production process, there is a need to understand fundamental processes underlying protein production, here under how protein production links to metabolism through global regulatory structures. In this study, two α-amylase-producing strains of A. oryzae, a wild type strain and a transformant strain containing additional copies of the α-amylase gene, were characterized at a systematic level. Based on integrated analysis of ome-data together with genome-scale metabolic network and flux calculation, we identified key genes, key enzymes, key proteins, and key metabolites involved in the processes of protein synthesis and secretion, nucleotide metabolism, and amino acid metabolism that can be the potential targets for improving industrial protein production. Keywords: Two Aspergillus oryzae strains and two different carbon sources Two carbon sources (glucose, maltose) with three biological replicates for A. oryzae strain A1560 and strain CF1.1

2012-02-12 | E-GEOD-22566 | biostudies-arrayexpress

Protein production by Aspergillus oryzae

2012-02-12 | GSE22566 | GEO

Expression analysis of Aspergillus nidulans AN1599 (PbcR) overexpression mutant

Project description:Investigation of whole genome gene expression level changes in Aspergillus nidulans AN1599 (PbcR) overexpression mutant, compared to the FGSC A4 wild-type strain. Overexpression of the Zn(II)2Cys6 –type transcription factor, AN1599.4 (PbcR, pimaradiene biosynthetic cluster regulator), activates a secondary metabolite gene cluster in Aspergillus nidulans. Activation of the pathway in Aspergillus nidulans lead to a production of ent-pimara-8(14),15-diene.

2012-04-16 | GSE32954 | GEO

Expression analysis of Aspergillus oryzae grown on different carbon sources

2013-07-15 | GSE48864 | GEO

Project description:β-Mannan utilization by Roseburia intestinalis

| PRJNA516396 | ENA

Cap analysis gene expression (CAGE) of Aspergillus luchuensis mut. kawachii control and sirtuin D disruptant strains

Project description:Sirtuin is considered to play a significant role in the growth phase-dependent gene expression. In this study, we characterized sirtuin in the white koji fungus, Aspergillus kawachii, to examine their role on regulation of amylolytic enzymes and citric acid productions during the solid-state culture (koji). Characterization of rice-koji made using five putative sirtuin gene disruptants indicated that they are involved in amylolytic activity and acidity of rice koji; especially the sirD disruptant showed lower levels of α-amylase activity and citric acid production per mycelial weight in the koji compared the control strain. In addition, the sirD disruptant also showed a change in mycelial pigmentation, higher sensitivity to cell wall biogenesis inhibitor such as calcofluor white and Congo red, and reduced conidia formation, indicating that SirD is required for secondary metabolism, cell wall integrity, and conidial development. The cap analysis gene expression (CAGE) indicated that the transcriptional changes related to the characteristic phenotype of the sirD disruptant (e.g., a reduced transcript level of acid-stable α-amylase gene and a citric acid exporter) in rice koji. These results indicated that SirD has a significant role on the global transcriptional regulation including productions of α-amylase and citric acid in A. kawachii during the solid-state fermentation process.

2019-06-15 | GSE132729 | GEO

Transcription profiling of RaxR-regulon members of Xanthomonas oryzae pv. oryzae PXO99A

Project description:Xanthomonas oryzae pv. oryzae strain PXO99A, so called Xoo, is disable to infect in rice cultivar carrying Xa21 gene. Disrupted mutant of raxR gene, response regulator of two-component regulatory system (TCS), in Xoo was previously shown to partially retrieve back the bacterial capability to establish in Xa21 rice. RaxR was shown to mediate the expression of other rax gene operon members and also its expression is changed dependent on cell population density. In this study, we investigated the regulatory mechanisms mediated by RaxR using whole-genome transcriptional profiling analysis in comparison of (i) PXO99R (PXO99 strain lacking RaxR) vs. PXO99, (ii) PXO99Rox (PXO99 strain overexpressing RaxR) vs. PXO99, and (iii) PXO99Rox vs. PXO99R. As a result of array analysis, we revealed that RaxR is not only required for AvrXa21 activitiy, it also plays roles in regulatory functions, for example, pathogenicity, motility, and stress tolerance. Then, we generated knock out mutants of RaxR regulon members to validate regulatory functions of RaxR and to extend other biological impacts of RaxR beyond the Xoo AvrXa21 activity. The combined interpretation from array analysis and mutant functional validation presents the complexity of regulatory pathways between AvrXa21 activity and other biological activities in Xoo. Keywords: Comparative transcription profiling between modified genetic mutant and wild type

2009-01-31 | GSE14498 | GEO

Expression analysis of Aspergillus oryzae grown on different carbon sources

Project description:In this study, we focused on chemically defined inducers or substrates to drive expression of cellulases, hemicellulases and accessory enzymes in the model filamentous fungus Aspergillus oryzae. Cellohexaose (O-CHE), mannohexaose (O-MHE), xylopentaose (O-XPE), arabinoheptaose (O-AHP), 1,3:1,4-M-NM-2-glucohexaose (O-BGHEXA), 63-M-NM-1-D-glucosyl-maltotriosyl-maltotriose (O-GMH), 61-M-NM-1-D-galactosyl-mannotriose (O-GM3), xyloglucan (X3Glc4-borohydride reduced; O-X3G4R), turanose (TYR) and sophorose (SOP) were used to induce the plant polysaccharide degradation machinery of A. oryzae. The strain used in this study was the A. oryzae sequenced strain RIB40, obtained from IBT culture collection at Technical University of Denmark. To obtain a global view of the A. oryzae transcriptome activated for plant biomass conversion, mRNA from growth after 2 h on 10 different carbohydrate active enzyme inducers (di- and M-bM-^@M-^Soligo saccharides) was subjected to custom-designed Agilent microarray analysis.

2013-07-15 | E-GEOD-48864 | biostudies-arrayexpress

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