Abstract
BackgroundThe degradation of cellulose and hemicellulose molecules into simpler sugars such as glucose is part of the second generation biofuel production process. Hydrolysis of lignocellulosic substrates is usually performed by enzymes produced and secreted by the fungus Trichoderma reesei. Studies identifying transcription factors involved in the regulation of cellulase production have been conducted but no overview of the whole regulation network is available. A transcriptomic approach with mixtures of glucose and lactose, used as a substrate for cellulase induction, was used to help us decipher missing parts in the network of T. reesei Rut-C30.ResultsExperimental results on the Rut-C30 hyperproducing strain confirmed the impact of sugar mixtures on the enzymatic cocktail composition. The transcriptomic study shows a temporal regulation of the main transcription factors and a lactose concentration impact on the transcriptional profile. A gene regulatory network built using BRANE Cut software reveals three sub-networks related to i) a positive correlation between lactose concentration and cellulase production, ii) a particular dependence of the lactose onto the β-glucosidase regulation and iii) a negative regulation of the development process and growth.ConclusionsThis work is the first investigating a transcriptomic study regarding the effects of pure and mixed carbon sources in a fed-batch mode. Our study expose a co-orchestration of xyr1, clr2 and ace3 for cellulase and hemicellulase induction and production, a fine regulation of the β-glucosidase and a decrease of growth in favor of cellulase production. These conclusions provide us with potential targets for further genetic engineering leading to better cellulase-producing strains in industry-like conditions.
Highlights
The degradation of cellulose and hemicellulose molecules into simpler sugars such as glucose is part of the second generation biofuel production process
Cellulase production is increased with lactose proportion but β-glucosidase activity is higher in glucose-lactose mixture In order to study its transcriptomic behavior on various carbon sources, T. reesei Rut-C30 was cultivated in fedbatch mode in a miniaturized experimental device called “fed-flask” [53], allowing us to obtain up to 6 biological replicates with minimal equipment
Glucose feed resulted in almost no protein production but in biomass growth (4.2 g L−1 biomass produced during fed-batch, see Additional file 1) while glucose/lactose mixtures resulted in intermediate profiles, with 0.6 g L−1 protein produced on 10 % lactose (G90-L10), and 1.4 g L−1 protein produced on 25 % lactose (G75-L25)
Summary
The degradation of cellulose and hemicellulose molecules into simpler sugars such as glucose is part of the second generation biofuel production process. Hydrolysis of lignocellulosic substrates is usually performed by enzymes produced and secreted by the fungus Trichoderma reesei. A transcriptomic approach with mixtures of glucose and lactose, used as a substrate for cellulase induction, was used to help us decipher missing parts in the network of T. reesei Rut-C30. One of the key challenges for industrial bio-ethanol production is to improve the competitiveness of plant biomass hydrolysis into fermentable sugars, using cellulosic enzymes. The filamentous fungus Trichoderma reesei, because of its high secretion capacity and cellulase production capability, is the most used microorganism for the industrial production of cellulolytic enzymes. Among the variety of mutant strains, Rut-C30 is known as the reference hyper-producer [6, 7], and its cellulase production is 15-20 times that of QM6a [8]. While most specificities (mutations, deletions, etc.) of the genetic background of Rut-C30 are seemingly unrelated to the production of cellulases [13], their impact should not be totally neglected and assesed according to a dedicated experimental design
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