The Wide-Domain Carbon Catabolite Repressor CreA Indirectly Controls Expression of the Aspergillus nidulans xlnB Gene, Encoding the Acidic Endo-β-(1,4)-Xylanase X ₂₄

Abstract

The Aspergillus nidulans xlnB gene, which encodes the acidic endo-β-(1,4)-xylanase X24, is expressed when xylose is present as the sole carbon source and repressed in the presence of glucose. That the mutation creAd30results in considerably elevated levels of xlnB mRNA indicates a role for the wide-domain repressor CreA in the repression of xlnB promoter (xlnBp) activity. Functional analyses of xlnBp::goxC reporter constructs show that none of the four CreA consensus target sites identified in xlnBp are functional in vivo. The CreA repressor is thus likely to exert carbon catabolite repression via an indirect mechanism rather than to influence xlnB expression by acting directly on xlnB.

Enhanced glycosyl hydrolase production in Aspergillus nidulans using transcription factor engineering approaches

“…The maximum activity produced by AR137 and AR138 was achieved between 72 and 96 hours after mycelial transfer to inducing conditions, whereas that of the xlnB p ::rhaA strains (AR142 and AR143) was achieved after 120 hours. This concords with previous observations in which the xlnA transcript was seen to accumulate to a greater level under inducing conditions than that of xlnB [18,[25][26][27]. Therefore, in a wild type genetic background under these experimental conditions the xlnA gene promoter is more effective for heterologous protein production than xlnB p .…”

“…In order to investigate the potential of the A. nidulans xylanolytic system (XlnR-xln p ) for the production of heterologous proteins, the well-characterized promoters of the xlnA and xlnB genes (xlnA p and xlnB p ) were chosen [18,[25][26][27] and the rhaA gene from A. aculeatus that encodes an extracellular α-L-rhamnosidase of industrial interest [37] was used as a reporter.…”

“…For efficient induction, xlnA p requires both xylose and alkaline pH whereas xlnB p requires xylose and acidic pH [25]. Previous studies that have demonstrated the roles of PacC, CreA and XlnR in the regulation of the A. nidulans xylanolytic system led to the identification of genetic backgrounds in which the expression of xlnA or xlnB was strongly enhanced compared to controls [18,[25][26][27]. Notable examples are creA d 30 and gpdA p ::xlnR (xlnA and xlnB overexpressed), pacC c 14 (xlnA overexpressed) and palA1 (xlnB overexpressed because the proteolytic activation of PacC is impaired) mutant strains.…”

“…In this context, we are exploring the potential of other regulated A. nidulans promoters, such as those of the xylanolytic genes xlnA (encodes xylanase X 22 ) and xlnB (encodes xylanase X 24 ), with the aim of obtaining improvements in expression levels and/or production times of both endogenous and heterologous high-value plant cell wall degrading enzymes. The biotic and abiotic factors influencing the expression of these promoters (xlnA p and xlnB p ) and the transcriptional repressors and activators associated with them have been thoroughly characterized [18,[25][26][27]]. An additional advantage of these alternative promoters is the almost complete lack of basal level of expression in the absence of the inducer.…”

“…The activities of the xlnA and xlnB promoters are tightly regulated by at least three mechanisms: ambient pH regulation mediated by the wide-domain transcriptional factor PacC, carbon catabolite repression (CCR) mediated by the wide-domain repressor CreA and specific induction in the presence of xylan or xylose controlled by the Zn(II) 2 Cys 6 protein XlnR, which is itself transcriptionally regulated by CreA [18,[25][26][27]. For efficient induction, xlnA p requires both xylose and alkaline pH whereas xlnB p requires xylose and acidic pH [25].…”