Project description:Trichoderma reesei is used to produce saccharifying enzyme cocktails for biofuels. There is limited understanding of the transcription factors (TFs) that regulate the release and catabolism of L-arabinose and D-galactose, as the main TF XYR1 is only partially involved. The T. reesei ortholog of ARA1 from Pyricularia oryzae that regulates L-arabinose release and catabolism, was deleted and characterized by growth profiling and transcriptomics along with a xyr1 mutant and xyr1/ara1 double mutant. Our results show that, in addition to the L-arabinose-related role, T. reesei ARA1 is essential for D-galactose release and catabolism, while XYR1 is not involved in this process. Overall design: In this study, we aim to identify the target genes of the arabinanolytic transcription factor ARA1 in the Ascomycetes Trichoderma reesei while growing on the monosaccharides L-arabinose and D-galactose through transcriptome analysis. We compare the transcriptomes of the reference strain QM9414 with three deletion strains: hemicellulytic regulator XYR1, ARA1 and the double deletion XYR1ARA1 after 2h of growth in the different carbon sources.
Project description:Background:Trichoderma reesei is one of the major producers of enzymes for the conversion of plant biomass to sustainable fuels and chemicals. Crude plant biomass can induce the production of CAZymes in T. reesei, but there is limited understanding of how the transcriptional response to crude plant biomass is regulated. In addition, it is unknown whether induction on untreated recalcitrant crude plant biomass (with a large diversity of inducers) can be sustained for longer. We investigated the transcriptomic response of T. reesei to the two industrial feedstocks, corn stover (CS) and soybean hulls (SBH), over time (4 h, 24 h and 48 h), and its regulatory basis using transcription factor deletion mutants (?xyr1 and ?ara1). We also investigated whether deletion of a xylulokinase gene (?xki1) from the pentose catabolic pathway that converts potential inducers could lead to increased CAZyme gene expression. Results:By analyzing the transcriptomic responses using clustering as well as differential and cumulative expression of plant biomass degrading CAZymes, we found that corn stover induced a broader range and higher expression of CAZymes in T. reesei, while SBH induced more pectinolytic and mannanolytic transcripts. XYR1 was the major TF regulating CS utilization, likely due to the significant amount of d-xylose in this substrate. In contrast, ARA1 had a stronger effect on SBH utilization, which correlates with a higher abundance of l-arabinose in SBH that activates ARA1. Blocking pentose catabolism by deletion of xki1 led to higher expression of CAZyme encoding genes on both substrates at later time points. Surprisingly, this was also observed for ?ara1 at later time points. Many of these genes were XYR1 regulated, suggesting that inducers for this regulator accumulated over time on both substrates. Conclusion:Our data demonstrates the complexity of the regulatory system related to plant biomass degradation in T. reesei and the effect the feedstock composition has on this. Furthermore, this dataset provides leads to improve the efficiency of a T. reesei enzyme cocktail, such as by the choice of substrate or by deleting xki1 to obtain higher production of plant biomass degrading CAZymes.