Deciphering the Regulation of Biomass Degradation By Anaerobic Fungi

To strengthen existing renewable technologies, more efficient methods to extract sugars from crude plant biomass (lignocellulose) are required. Biomass breakdown represents a significant barrier towards the production of fuels and chemicals. However, some microbes in extreme environments have evolved to degrade lignocellulose efficiently. Anaerobic gut fungi are the primary colonizers of plant material in the guts of herbivores, like sheep and goats. Encoded within their genomes is an incredible variety of biomass degrading enzymes including cellulases, hemicellulases, esterases, and polysaccharide deacetylases. However, little is known about the overall suite of biomass degrading enzymes produced, and even less is known about their regulation.

Sequencing technologies have improved and become more accessible offering a useful method to examine regulation of biomass degrading genes and gene regulators in gut fungi. We have used RNA sequencing technologies to elucidate regulation patterns in anaerobic fungi in response to carbon source of varying complexity. Two species of fungi, Anaeromyces robustus and Neocallimastix californiae, were grown on glucose, cellobiose, cellulose, and lignocellulose. Expression counts of transcripts were compared to the base case of glucose, where expression of biomass degrading enzymes was lowest represented the smallest percentage of total transcriptome expression. In both cases gene set enrichment analysis demonstrated enrichment of cellulases and hemicellulases on more complex substrates. However, in the case of A. robustus enrichment appears to be primarily triggered by cellobiose, but for N. californiae enrichment occurs more gradually, which some GH classes enriched during growth on cellobiose and cellulose while others were enriched only on biomass. These results suggest that different species of gut fungi maintain varying global regulation networks. For A. robustus, while there may be multiple integrated regulatory mechanisms, the regulation of biomass degrading machinery may be primarily controlled by simple break out molecules, like cellobiose.

Further, we are integrating these results with recently obtained genomic information to identify common regulons and promoters that govern transcriptional regulation by substrates. By examining the upstream regions of genes with similar predicted function as well as similar regulation patterns, we may identify promoters used to control the expression of these enzymes. Collectively, this information will provide important insight into gut fungal metabolism, and present new strategies to amplify their breakdown abilities.