Enhanced cellulase production in Trichoderma reesei RUT C30 via constitution of minimal transcriptional activators

Cellulase can convert lignocellulosic feedstocks into fermentable sugars, which can be used for the industrial production of biofuels and chemicals. The high cost of cellulase production remains a challenge for lignocellulose breakdown. Trichoderma reesei RUT C30 serves as a well-known industrial workhorse for cellulase production. Therefore, the enhancement of cellulase production by T. reesei RUT C30 is of great importance.

Two sets of novel minimal transcriptional activators (DBD_ace2-VP16 and DBD_cre1-VP16) were designed and expressed in T. reesei RUT C30. Expression of DBD_ace2-VP16 and DBD_cre1-VP16 improved cellulase production under induction (avicel or lactose) and repression (glucose) conditions, respectively. The strain T_MTA66 under avicel and T_MTA139 under glucose with the highest cellulase activities outperformed other transformants and the parental strain under the corresponding conditions. For T_MTA66 strains, the highest FPase activity was approximately 1.3-fold greater than that of the parental strain RUT C30 at 120 h of cultivation in a shake flask using avicel as the sole carbon source. The FPase activity (U/mg biomass) in T_MTA139 strains was approximately 26.5-fold higher than that of the parental strain RUT C30 at 72 h of cultivation in a shake flask using glucose as the sole carbon source. Furthermore, the crude enzymes produced in the 7-L fermenter from T_MTA66 and T_MTA139 supplemented with commercial β-glucosidase hydrolyzed pretreated corn stover effectively.

These results show that replacing natural transcription factors with minimal transcriptional activators is a powerful strategy to enhance cellulase production in T. reesei. Our current study also offers an alternative genetic engineering strategy for the enhanced production of industrial products by other fungi.