Mutation of the Xylanase regulator 1 causes a glucose blind hydrolase expressing phenotype in industrially used Trichoderma strains

Abstract

BackgroundTrichoderma reesei is an organism involved in degradation of (hemi)cellulosic biomass. Consequently, the corresponding enzymes are commonly used in different types of industries, and recently gained considerable importance for production of second-generation biofuel. Many industrial T. reesei strains currently in use are derived from strain Rut-C30, in which cellulase and hemicellulase expression is released from carbon catabolite repression. Nevertheless, inducing substances are still necessary for a satisfactory amount of protein formation.ResultsHere, we report on a T. reesei strain, which exhibits a very high level of xylanase expression regardless if inducing substances (e.g. D-xylose, xylobiose) are used. We found that a single point mutation in the gene encoding the Xylanase regulator 1 (Xyr1) is responsible for this strong deregulation of endo-xylanase expression and, moreover, a highly elevated basal level of cellulase expression. This point mutation is localized in a domain that is common in binuclear zinc cluster transcription factors. Only the use of sophorose as inducer still leads to a slight induction of cellulase expression. Under all tested conditions, the formation of cbh1 and cbh2 transcript level strictly follows the transcript levels of xyr1. The correlation of xyr1 transcript levels and cbh1/cbh2 transcript levels and also their inducibility via sophorose is not restricted to this strain, but occurs in all ancestor strains up to the wild-type QM6a.ConclusionsEngineering a key transcription factor of a target regulon seems to be a promising strategy in order to increase enzymes yields independent of the used substrate or inducer. The regulatory domain where the described mutation is located is certainly an interesting research target for all organisms that also depend so far on certain inducing conditions.

The Genomes of Three Uneven Siblings: Footprints of the Lifestyles of Three Trichoderma Species

“…This protein is a homolog to XlnR of A. niger (1010) and has been demonstrated to act as an activator of xylanase and cellulase genes (1008,1009,1011). A third Zn(II) 2 Cys 6 TF has been studied in T. reesei (1012).…”

Effect of different terminators on transcription regulatory factor ClrB and XlnR in Penicillium oxalicum 114-2

“…But most researchers will focus on the rep-lacement of the stronger promoter, overexpression, and directional transformation [11,12] . Few resea-rchers have studied the effects of terminators on cellulase production.…”

Organic acid production in Aspergillus niger and other filamentous fungi

“…Regarding the T. reesei genes, str1 expression was strongly induced by xylose, in a similar way to A. niger xltA, and also by sophorose, which is also an inducer of the T. reesei xylanolytic system (Derntl et al, 2013). In addition, it was clearly transcriptionally regulated by Xyr1, encoding the main regulator of the xylose metabolism in T. reesei (Stricker et al, 2006).…”

“…to the Michaelis-Menten model and used to estimate the appropriate kinetic parameters (K m and V max ) as previously described (Derntl et al, 2013) (Table 6.2) (Additional file 7).…”

“…6.5.6 Transcriptional analysis of A. niger and T. reesei genes RNA isolation from A. niger (Sloothaak et al, 2015) and T. reesei (Derntl et al, 2013) mycelium was done as described previously . Reverse transcription, quantitative PCRs and calculations were performed following the protocols and instruments described in Mach-Aigner et al, 2012(Mach-Aigner et al, 2012.…”