Cellulolytic enzyme genes are significantly upregulated by most of the mutations that cause defects in the ligninolytic activity in the white-rot fungus Pleurotus ostreatus

Takehito Nakazawa 1 Rina Kodera 1 Ryota Morimoto 1 Atsuki Takenaka 1 Shoko Tsuji 2 Hiroshi Nishimura 2 Takashi Watanabe 2 Masahiro Sakamoto 1 Yoichi Honda 1
1Graduate School of Agriculture, Kyoto University, Kyoto, Japan
2Research Institute for Sustainable Humanosphere, Kyoto University, Kyoto, Japan

White-rot fungi play an important role in the global carbon cycle because wood lignin is almost exclusively biodegraded by them in nature. The oyster mushroom Pleurotus ostreatus is frequently used for biochemical and genomic studies of lignin biodegradation. Recently, we identified three genes, wtr1, chd1 and pex1, in which mutations cause defects in the ligninolytic activity, RBBR decolorization and wood lignin degradation, in P. ostreatus. Here, we show results of comparative RNA-seq analysis to identify genes of which expression are significantly affected by aforementioned mutations when they are grown on sawdust media. It was shown that many genes probably involved in lignin degradation were significantly downregulated in the single-gene disruptant of the three genes. We also found that many genes encoding cellulolytic enzymes, putative four endo-β-1, 4 glucanases belonging to GH6 and GH7, cellobiose dehydrogenase, many copper-dependent lytic polysaccharide monooxigenases belonging to AA9, and an exo- or endo- glucanase belonging to GH131, are significantly upregulated (70-22 folds in RPKM values) by chd1 or pex1 disruption, but not by wtr1 disruption. It was shown that extracellular enzymes obtained from mutant UVJ3-3, the chd1-1 mutant, grown on sawdust medium saccharified Avicel much more efficiently than the wild-type strain PC9, which is consistent with the result of RNA-seq. We recently isolated new mutants (total five) defective in the ligninolytic activity, and genes responsible for the phenotype of some mutants were identified. qRT-PCR analysis revealed that aforementioned cellulolytic enzyme genes are also significantly upregulated in four out of five mutants, while genes probably involved in ligninolysis are downregulated in all of the five mutants. These suggest gene regulation mechanisms switching between ligninolytic mode and cellulolytic one in P. ostreatus although it remains unclear whether the change in gene expression is the cause or result of the defects in ligninolysis.