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

Abstract White‐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.