The lignin‐degrading abilities of Gelatoporia subvermispora gat1 and pex1 mutants generated via CRISPR/Cas9

Abstract

AbstractWhite‐rot fungi efficiently degrade wood lignin; however, the mechanisms involved remain largely unknown. Recently, a forward genetics approach to identify several genes in Pleurotus ostreatus (Agaricales) in which mutations cause defects in wood lignin degradation was used. For example, pex1 encodes a peroxisome biogenesis factor and gat1 encodes a putative Agaricomycetes‐specific DNA‐binding transcription factor. In this study, we examined the effects of single‐gene mutations in pex1 or gat1 on wood lignin degradation in another white‐rot fungus, Gelatoporia (Ceriporiopsis) subvermispora (Polyporales), to investigate conserved and derived degradation mechanisms in white‐rot fungi. G. subvermispora pex1 and gat1 single‐gene mutant strains were generated from a monokaryotic wild‐type strain, FP‐90031‐Sp/1, using plasmid‐based CRISPR/Cas9. As in P. ostreatus, Gsgat1 mutants were nearly unable to degrade lignin sourced from beech wood sawdust medium (BWS), while Gspex1 mutants exhibited a delay in lignin degradation. We also found that the transcripts of lignin‐modifying enzyme‐encoding genes, mnp4, mnp5, mnp6, mnp7, and mnp11, which predominantly accumulate in FP‐90031‐Sp/1 cultured with BWS, were greatly downregulated in Gsgat1 mutants. Taken together, the results suggest that Gat1 may be a conserved regulator of the ligninolytic system of white‐rot fungi and that the contribution of peroxisomes to the ligninolytic system may differ among species.