Research Article: Investigation of lignocellulolytic enzymes during different growth phases of Ganoderma lucidum strain G0119 using genomic, transcriptomic and secretomic analyses

Date Published: May 31, 2018

Publisher: Public Library of Science

Author(s): Shuai Zhou, Jingsong Zhang, Fuying Ma, Chuanhong Tang, Qingjiu Tang, Xiaoyu Zhang, Jean-Guy Berrin.

http://doi.org/10.1371/journal.pone.0198404

Abstract

Ganoderma lucidum is a medicinal mushroom that is well known for its ability to enhance human health, and products made from this fungus have been highly profitable. The substrate-degrading ability of G. lucidum could be related to its growth. CAZy proteins were more abundant in its genome than in the other white rot fungi models. Among these CAZy proteins, changes in lignocellulolytic enzymes during growth have not been well studied. Using genomic, transcriptomic and secretomic analyses, this study focuses on the lignocellulolytic enzymes of G. lucidum strain G0119 to determine which of these degradative enzymes contribute to its growth. From the genome sequencing data, genes belonging to CAZy protein families, especially genes involved in lignocellulose degradation, were investigated. The gene expression, protein abundance and enzymatic activity of lignocellulolytic enzymes in mycelia over a growth cycle were analysed. The overall expression cellulase was higher than that of hemicellulase and lignin-modifying enzymes, particularly during the development of fruiting bodies. The cellulase and hemicellulase abundances and activities increased after the fruiting bodies matured, when basidiospores were produced in massive quantities till the end of the growth cycle. Additionally, the protein abundances of the lignin-modifying enzymes and the expression of their corresponding genes, including laccases and lignin-degrading heme peroxidases, were highest when the mycelia fully spread in the compost bag. Type I cellobiohydrolase was observed to be the most abundant extracellular lignocellulolytic enzyme produced by the G. lucidum strain G0119. The AA2 family haem peroxidases were the dominant lignin-modifying enzyme expressed during the mycelial growth phase, and several laccases might play roles during the formation of the primordium. This study provides insight into the changes in the lignocellulose degradation ability of G. lucidum during its growth and will facilitate the discovery of new approaches to accelerate the growth of G. lucidum in culture.

Partial Text

Ganoderma lucidum (Leyss. ex Fr.) Karst, which is also known as “Lingzhi”, is a lamella-less basidiomycetous fungus that belongs to family Polyporaceae [1]. G. lucidum has been used as a traditional herbal medicine for the treatment of various diseases, including hepatopathy, nephritis, neurasthenia and asthma [2]. As a species of white-rot fungus, G. lucidum decays plant biomass via secreted enzymes. Similar to other mushrooms with economic value, G. lucidum is artificially cultivated in logs or compost. Typically, the compost used to cultivate mushrooms contains approximately 60–70% lignocellulose by dry weight [3]. The lignocellulose in the compost is degraded and transformed into the fungal biomass [4, 5] and accounts for 85% of the fruiting body and 45% of the mycelial dry weights [6]. Thus, the substrate-degrading ability of G. lucidum could be a factor related to its growth.

In this study, we sequenced the genome of the G. lucidum strain G0119. Multiple strains of Ganoderma spp. have been sequenced at present. For instance, the genome size of strain CGMCC5.0026 (NCBI accession number: PRJNA71455) was reported to be 43.4 Mb in size, and containing 16,113 genes [11], of which 451 were CAZy genes. The reported genome size of strain Ganoderma sp. 10,597 SS1 in the JGI database is 39.52 Mb, and containing 12,910 genes [33]. Phylogenetic analysis of these two strains and G. lucidum G0119 indicated that strain G0119 was more closely related to strain CGMCC5.0026 than 10597 SS1, but that some gene loss has during its evolution [17]. Strain G0119 contained more CAZy protein genes than strain CGMCC5.0026, particularly with respect to the CE10 and CBM50 protein families, which primarily included esterases, chitinases and peptide enzymes. The number of CAZy genes identified in strain 10597 SS1 was much lower than that of strain G0119, possibly due to the use of the Sanger sequencing method instead of the next-generation sequencing approach used in this study [33]. Compared with the genomes of other wood-degrading fungal models, such as Phanerochaete chrysosporium, Laccaria bicolor and Schizophyllum commune [34–36], G. lucidum strain G0119 contained more GH and CE family genes, particularly those of the GH18, CE10 and CE16 families, which primarily consisted of chitinases, N-acetylglucosaminidases, arylesterases, and acetylesterases.

The sequencing data (NCBI: PRJNA406843), the G. lucidum G0119 genome sequence (NCBI: SRR6026942) and the raw Illumina sequencing data during the 5 growth phases, performed in duplicates (NCBI: SRR6026942, SRR6027258, SRR6027261, SRR6027352, SRR6037451, SRR6037587, SRR6037726, SRR6040102, SRR6040213 and SRR6043945), were deposited at NCBI.

 

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http://doi.org/10.1371/journal.pone.0198404

 

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