Abstract
Fungi are the most important and efficient plant biomass degrading microorganisms and use different strategies to degrade plant biomass, reflecting their ecological niches. Only a deep understanding of these strategies, especially during growth on crude plant biomass and not only on simple mono- or polysaccharides, will lead to further improvements
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of biorefinery processes that use them as a substrate. The aim of this PhD thesis is to dissect the plant biomass degrading strategies of specialist (Podospora anserina), semi-specialist (Trichoderma reesei) and generalist (Aspergillus niger) fungi. Chapter 2 described the characterization of the missing regulator responding to L-arabinose and D-galactose (two of the main monosaccharides in plant biomass) in T. reesei: ARA1, which is a functional but not sequence ortholog of the arabinanolytic regulator AraR from the generalist Aspergillus niger, demonstrating a clear case of parallel evolution. Chapter 3 described a simple method to overcome senescence in P. anserina, which was used in Chapter 4 for a time-course transcriptome during growth on two industrial feedstocks, soybean hulls (SBH) (dicot, richer in pectin) and corn stover (CS) (monocot, richer in hemicellulose). Overall, SBH resulted in a larger diversity of expressed genes, confirming previous proteomics studies. Our results provide an in depth view of the transcriptomic adaptation of P. anserina to substrate composition, but also pointed out strategies to improve saccharification of plant biomass at the industrial level, such as novel enzymes or harvesting time for enzyme production. The same feedstocks (CS and SBH) were used in Chapter 5 to investigate the time course transcriptomic response of T. reesei. Two regulatory (Δxyr1 and Δara1) and one catabolic (Δxki1) mutant were used together with the wild type to deeply investigate its degrading strategy. CS induced a broader range and higher expression of CAZyme encoding genes in T. reesei, while SBH induced more pectinolytic and mannanolytic genes. XYR1 was the major TF regulating CS utilization, while ARA1 had a stronger effect on SBH utilization, matching with the substrate composition. 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 Δara1 at later time points. Many of these genes were XYR1 regulated, suggesting that inducer(s) for this regulator accumulated over time on both substrates. 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. Finally in Chapter 6 I discussed the initial question that leads this thesis: what makes a fungus a generalist, a semi-specialist or a specialist? This answer is only at its initial stage and more studies are required, but using the data from Chapters 2, 4 and 5 and previously studies in A. niger, it appears that CAZyme gene content in fungal genomes can only partially indicate the fungal strategy, while their regulation may be the key factor, supporting our initial hypothesis (see Chapter 1).
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