N,N-dimethylformamide induces cellulase production in the filamentous fungus Trichoderma reesei

The filamentous fungus Trichoderma reesei produces cellulase enzymes that are widely studied for lignocellulose bioconversion to biofuel. N,N-dimethylformamide (DMF) is a versatile organic solvent used in large quantities in industries.

In this study, we serendipitously found that biologically relevant concentrations of extracellular DMF-induced cellulase production in the T. reesei hyper-cellulolytic mutant Rut-C30 and wild-type strain QM6a. Next, by transcriptome analysis, we determined that plc-e encoding phospholipase C was activated by DMF and revealed that cytosolic Ca²⁺ plays a vital role in the response of T. reesei to DMF. Using EGTA (a putative extracellular Ca²⁺ chelator) and LaCl₃ (a plasma membrane Ca²⁺ channel blocker), we demonstrated that DMF induced a cytosolic Ca²⁺ burst via extracellular Ca²⁺ and Ca²⁺ channels in T. reesei, and that the cytosolic Ca²⁺ burst induced by DMF-mediated overexpression of cellulase through calcium signaling. Deletion of crz1 confirmed that calcium signaling plays a dominant role in DMF-induced cellulase production. Additionally, 0.5-2% DMF increases the permeability of T. reesei mycelia for cellulase release. Simultaneous supplementation with 1% DMF and 10 mM Mn²⁺ to T. reesei Rut-C30 increased cellulase activity approximately fourfold compared to that without treatment and was also more than that observed in response to either treatment alone.

Our results reveal that DMF-induced cellulase production via calcium signaling and permeabilization. Our results also provide insight into the role of calcium signaling in enzyme production for enhanced cellulase production and the development of novel inducers of cellulase.