Deletion of the middle region of the transcription factor ClrB in Penicillium oxalicum enables cellulase production in the presence of glucose

Abstract

Enzymes that degrade lignocellulose to simple sugars are of great interest in research and for biotechnology because of their role in converting plant biomass to fuels and chemicals. The synthesis of cellulolytic enzymes in filamentous fungi is tightly regulated at the transcriptional level, with the transcriptional activator ClrB/CLR-2 playing a critical role in many species. In <i>Penicillium oxalicum</i>, <i>clrB</i> overexpression could not relieve the dependence of cellulase expression on cellulose as an inducer, suggesting that <i>clrB</i> is controlled post-transcriptionally. In this study, using a reporter gene system in yeast, we identified the C-terminal region of ClrB/CLR-2 as a transcriptional activation domain. Expression of <i>clrB^ID</i> , encoding a ClrB derivative in which the DNA-binding and transcriptional activation domains are fused together to remove the middle region, led to cellulase production in the absence of cellulose in <i>P. oxalicum</i> Strikingly, the <i>clrB^ID</i> -expressing strain produced cellulase on carbon sources that normally repress cellulase expression, including glucose and glycerol. Results from deletion of the carbon catabolite repressor gene <i>creA</i> in the <i>clrB^ID</i> -expressing strain suggested that the effect of <i>clrB^ID</i> is independent of CreA's repressive function. A similar modification of <i>clrB</i> in <i>Aspergillus niger</i> resulted in the production of a mannanase in glucose medium. Taken together, these results indicate that ClrB suppression under noninducing conditions involves its middle region, suggesting a potential strategy to engineer fungal strains for improved cellulase production on commonly used carbon sources.