Independent duplications of α-amylase in different strains of Aspergillus oryzae

Hunter, A. J.; Jin, Bo; Kelly, Joseph F.

doi:10.1016/j.fgb.2011.01.006

Cited by 23 publications

(25 citation statements)

References 31 publications

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“…In contrast to the case for the creB disruptant in T. reesei, improved growth was not observed on maltose, likely because secreted ␣-glucosidase activity would not limit growth of A. oryzae RIB40, which was selected for strong starch degradation (21).…”

Section: Resultsmentioning

confidence: 62%

Deletion of creB in Aspergillus oryzae Increases Secreted Hydrolytic Enzyme Activity

Hunter

Morris

Jin

et al. 2013

Appl Environ Microbiol

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dAspergillus oryzae has been used in the food and beverage industry for centuries, and industrial strains have been produced by multiple rounds of selection. Targeted gene deletion technology is particularly useful for strain improvement in such strains, particularly when they do not have a well-characterized meiotic cycle. Phenotypes of an Aspergillus nidulans strain null for the CreB deubiquitinating enzyme include effects on growth and repression, including increased activity levels of various enzymes. We show that Aspergillus oryzae contains a functional homologue of the CreB deubiquitinating enzyme and that a null strain shows increased activity levels of industrially important secreted enzymes, including cellulases, xylanases, amylases, and proteases, as well as alleviated inhibition of spore germination on glucose medium. Reverse transcription-quantitative PCR (RTqPCR) analysis showed that the increased levels of enzyme activity in both Aspergillus nidulans and Aspergillus oryzae are mirrored at the transcript level, indicating transcriptional regulation. We report that Aspergillus oryzae DAR3699, originally isolated from soy fermentation, has a similar phenotype to that of a creB deletion mutant of the RIB40 strain, and it contains a mutation in the creB gene. Collectively, the results for Aspergillus oryzae, Aspergillus nidulans, Trichoderma reesei, and Penicillium decumbens show that deletion of creB may be broadly useful in diverse fungi for increasing production of a variety of enzymes.A spergillus oryzae is a multicellular fungus that has been used for centuries for the production of Asian foods and beverages, including sake (rice wine) and shoyu (fermented soybean). Today it is also used industrially as a source of secreted enzymes, including cellulases, amylases, proteases, ␤-galactosidase, and lipase, and as a host for the production of heterologous proteins (1, 2).creB was identified in the model filamentous fungus Aspergillus nidulans in a screen for mutations that alleviate carbon catabolite repression (CCR) (3). CCR is a mechanism by which genes for the utilization of nonpreferred carbon sources are repressed in the presence of preferred carbon sources; organisms thus avoid wasting energy producing enzymes for the degradation of complex carbon sources when more readily metabolized carbon sources are available. A. nidulans creB mutants grow well and alleviate CCR of various enzymes, including acetyl-coenzyme A (acetyl-CoA) synthetase, isocitrate lyase, acetamidase, and alcohol dehydrogenase (3, 4). In addition, A. nidulans creB mutants show a pleiotropic range of phenotypes apparently unrelated to CCR, including slightly reduced conidiation, reduced growth with quinate, proline, or glucuronate as the sole carbon source, enhanced growth on acetamide or acrylamide, resistance to molybdate, hypersensitivity to acriflavine, and reduced acidification of liquid growth media (3-5). creB encodes a deubiquitinating enzyme (DUB); DUBs are cysteine proteases that specifically cleave ubiquitin from ubiqui...

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Section: Resultsmentioning

confidence: 62%

Deletion of creB in Aspergillus oryzae Increases Secreted Hydrolytic Enzyme Activity

Hunter

Morris

Jin

et al. 2013

Appl Environ Microbiol

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“…Gene duplication of -amylase genes likely occurred during the domestication of A. oryzae, and recently Hunter et al suggested that the amyB and amyC genes arose by the activity of transposable elements in the downstream regions. 24) Our data provide the idea that the ability to express large amounts of -amylases in A. oryzae is attributed to duplicated -amylase genes such as amyB and amyC. Total RNAs were extracted from the -amylase gene disruptants and the control strain after culture in MPY liquid medium at 30 C for 24 h. Synthesized cDNAs were subjected to real-time PCR analysis.…”

Section: Discussionmentioning

confidence: 88%

“…24) The A. oryzae amyA gene is present in the conserved gene cluster together with genes for -glucosidase AgdA and transcription factor AmyR. Gene duplication of -amylase genes likely occurred during the domestication of A. oryzae, and recently Hunter et al suggested that the amyB and amyC genes arose by the activity of transposable elements in the downstream regions.…”

Section: Discussionmentioning

confidence: 99%

Contribution Ratios ofamyA,amyB,amyCGenes to High-Level α-Amylase Expression inAspergillus oryzae

Nemoto

Maruyama

Kitamoto

2012

Bioscience, Biotechnology, and Biochemistry

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Aspergillus oryzae strains express α-amylases abundantly, and the genome reference strain RIB40 has three α-amylase genes (amyA, amyB, and amyC). However, there is no information on the contribution ratios of individual α-amylase genes to total expression. In this study, we generated single, double, and triple disruptants of α-amylase genes by employing a strain (ΔligD) with high gene-targeting efficiency and pyrG marker recycling in A. oryzae. All the disruptants showed reduced activities of α-amylases, and the triple disruptant completely lost activity. Comparative analyses of the activities and mRNA amounts of the α-amylases suggest that the contribution of amyA to the α-amylase expression is smaller than those of amyB and amyC. The present study suggests that the ability to express a large amount of α-amylases in A. oryzae is attributed to gene duplication of genes such as amyB and amyC.

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“…Notable examples of gene duplication being associated with microbial domestication include those of α-amylase in Aspergillus oryzae , which is instrumental in starch saccharification during the production of sake (Hunter et al 2011; Gibbons et al 2012), and of the MAL1 and MAL3 loci in beer-associated strains of S. cerevisiae that metabolize maltose—the most abundant sugar in the beer wort (Gallone et al 2016; Gonçalves et al 2016). Beer strains of S. cerevisiae often contain additional duplicated genes associated with maltose metabolism, including MPH2 and MPH3 , two maltose permeases, and the putative maltose-responsive transcription factor, YPR196W (Gonçalves et al 2016).…”

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confidence: 99%

Extensive Copy Number Variation in Fermentation-Related Genes AmongSaccharomyces cerevisiaeWine Strains

Steenwyk

Rokas

2017

G3 Genes|Genomes|Genetics

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Due to the importance of Saccharomyces cerevisiae in wine-making, the genomic variation of wine yeast strains has been extensively studied. One of the major insights stemming from these studies is that wine yeast strains harbor low levels of genetic diversity in the form of single nucleotide polymorphisms (SNPs). Genomic structural variants, such as copy number (CN) variants, are another major type of variation segregating in natural populations. To test whether genetic diversity in CN variation is also low across wine yeast strains, we examined genome-wide levels of CN variation in 132 whole-genome sequences of S. cerevisiae wine strains. We found an average of 97.8 CN variable regions (CNVRs) affecting 4% of the genome per strain. Using two different measures of CN diversity, we found that gene families involved in fermentation-related processes such as copper resistance (CUP), flocculation (FLO), and glucose metabolism (HXT), as well as the SNO gene family whose members are expressed before or during the diauxic shift, showed substantial CN diversity across the 132 strains examined. Importantly, these same gene families have been shown, through comparative transcriptomic and functional assays, to be associated with adaptation to the wine fermentation environment. Our results suggest that CN variation is a substantial contributor to the genomic diversity of wine yeast strains, and identify several candidate loci whose levels of CN variation may affect the adaptation and performance of wine yeast strains during fermentation.

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Independent duplications of α-amylase in different strains of Aspergillus oryzae

Cited by 23 publications

References 31 publications

Deletion of creB in Aspergillus oryzae Increases Secreted Hydrolytic Enzyme Activity

Deletion of creB in Aspergillus oryzae Increases Secreted Hydrolytic Enzyme Activity

Contribution Ratios ofamyA,amyB,amyCGenes to High-Level α-Amylase Expression inAspergillus oryzae

Extensive Copy Number Variation in Fermentation-Related Genes AmongSaccharomyces cerevisiaeWine Strains

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