Abstract
Aspergillus aculeatus produces cellulolytic enzymes in the presence of their substrates. We screened a library of 12,000 A. aculeatus T-DNA-inserted mutants to identify a regulatory factor involved in the expression of their enzyme genes in response to inducers. We found one mutant that reduced the expression of FIII-avicelase (chbI) in response to cellulose. T-DNA was inserted into a putative protein kinase gene similar to AN10082 in A. nidulans, serine-arginine protein kinase F, SrpkF. Fold increases in srpkF gene expression in response to various carbon sources were 2.3 (D-xylose), 44 (Avicel®), 59 (Bacto™ Tryptone), and 98 (no carbon) compared with D-glucose. Deletion of srpkF in A. aculeatus resulted in a significant reduction in cellulose-responsive expression of chbI, hydrocellulase (cel7b), and FIb-xylanase (xynIb) genes at an early induction phase. Further, the srpkF-overexpressing strain showed upregulation of the srpkF gene from four- to nine-fold higher than in the control strain. srpkF overexpression upregulated cbhI and cel7b in response to cellobiose and the FI-carboxymethyl cellulase gene (cmc1) and xynIb in response to D-xylose. However, the srpkF deletion did not affect the expression of xynIb in response to D-xylose due to the less expression of srpkF under the D-xylose condition. Our data demonstrate that SrpkF is primarily involved in cellulose-responsive expression, though it has a potential to stimulate gene expression in response to both cellobiose and D-xylose in A. aculeatus.
Highlights
Filamentous fungi are prominent producers of enzymes that degrade lignocellulose (Payne et al 2015)
We identified a putative protein kinase, serine–arginine protein kinase F (SrpkF), which participates in the ManR- and XlnR-dependent signaling pathways to induce gene expression in response to cellulosic carbon sources
In this study, we demonstrated that deletion of srpkF resulted in the reduction of cellulase and xylanase gene expression in response to cellulosic carbon sources
Summary
Filamentous fungi are prominent producers of enzymes that degrade lignocellulose (Payne et al 2015). Trichoderma reesei is well known to produce copious amounts of cellulolytic enzymes (Bischof et al 2016). T. reesei glycoside hydrolases are being continuously improved to utilize lignocellulose for feedstock and generate biobased products. B-Glucosidase from A. aculeatus showed high compatibility in T. reesei, suggesting that A. aculeatus produces promising enzymes to liberate glucose from cellulose. Cellulolytic enzymes of A. aculeatus are not utilized in industry because of low production levels. We aim to understand the regulatory mechanisms of the associated gene expressions and apply this knowledge to improving enzyme production in A. aculeatus
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