The Impact of DNA Methylation in Trichoderma reesei on Cellulase Production and Strain Degeneration

The spontaneous loss of cellulase productivity of industrial T. reesei strains during production results in significant economic losses. This phenomenon was suggested to be epigenetically regulated, but the previous findings did not explain which epigenetic mechanisms occur and how they promote strain degeneration. Until now, the epigenetic landscape of T. reesei has been poorly understood. This study investigated whether DNA methylation and cellulase production are connected, and, if so, what that connection is and how it relates to strain degeneration. In order to determine what the impact of DNA methylation is on strain degeneration, we induced hypomethylation with hydralazine HCL, which showed a reduced non-productive phenotype and partially restored cellulase productivity. As a second test, we conducted a global DNA cytosine methylation assay, which showed T. reesei DNA methylation levels of between 0.2 and 1.3% 5-mC. Importantly, non-productive strains exhibited stronger methylation than productive counterparts, and global methylation patterns varied depending on the carbon source. As a final test, we carried out deletion experiments targeting the putative DNA methyltransferases Dim2 and Rid1, which initially reduced the occurrence of a non-producing subpopulation, but subsequent sub-cultivation eliminated cellulase productivity. This study shows that DNA methylation impacts T. reesei cellulase productivity, an understanding that can help us develop targeted strategies to reduce strain degeneration and improve cellulase production in industrial applications.