Filamentous fungi, such asNeurospora crassa, are very efficient in deconstructing plant biomass by the secretion of an arsenal of plant cell wall-degrading enzymes, by remodeling metabolism to accommodate production of secreted enzymes, and by enabling transport and intracellular utilization of plant biomass components. Although a number of enzymes and transcriptional regulators involved in plant biomass utilization have been identified, how filamentous fungi sense and integrate nutritional information encoded in the plant cell wall into a regulatory hierarchy for optimal utilization of complex carbon sources is not understood. Here, we performed transcriptional profiling ofN. crassaon 40 different carbon sources, including plant biomass, to provide data on how fungi sense simple to complex carbohydrates. From these data, we identified regulatory factors inN. crassaand characterized one (PDR-2) associated with pectin utilization and one with pectin/hemicellulose utilization (ARA-1). Using in vitro DNA affinity purification sequencing (DAP-seq), we identified direct targets of transcription factors involved in regulating genes encoding plant cell wall-degrading enzymes. In particular, our data clarified the role of the transcription factor VIB-1 in the regulation of genes encoding plant cell wall-degrading enzymes and nutrient scavenging and revealed a major role of the carbon catabolite repressor CRE-1 in regulating the expression of major facilitator transporter genes. These data contribute to a more complete understanding of cross talk between transcription factors and their target genes, which are involved in regulating nutrient sensing and plant biomass utilization on a global level.

Identifying nutrients available in the environment and utilizing them in the most efficient manner is a challenge common to all organisms. The model filamentous fungus Neurospora crassa is capable of utilizing a variety of carbohydrates, from simple sugars to the complex carbohydrates found in plant cell walls. The zinc binuclear cluster transcription factor CLR-1 is necessary for utilization of cellulose, a major, recalcitrant component of the plant cell wall; however, expression of clr-1 in the absence of an inducer is not sufficient to induce cellulase gene expression. We performed a screen for unidentified actors in the cellulose-response pathway and identified a gene encoding a hypothetical protein (clr-3) that is required for repression of CLR-1 activity in the absence of an inducer. Using clr-3 mutants, we implicated the hyperosmotic-response pathway in the tunable regulation of glycosyl hydrolase production in response to changes in osmolarity. The role of the hyperosmotic-response pathway in nutrient sensing may indicate that cells use osmolarity as a proxy for the presence of free sugar in their environment. These signaling pathways form a nutrient-sensing network that allows N. crassa cells to tightly regulate gene expression in response to environmental conditions. MAP kinase cascade | carbon metabolism | hyperosmotic response | carbon catabolite repression | plant biomass degradation

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