Role of calcineurin-responsive transcription factor CRZ1 in ganoderic acid biosynthesis by Ganoderma lucidum

Roles of functional genes in secondary metabolism of mushroom-forming basidiomycetes are rarely clarified due to the difficulty of gene knock-out in them. In Ganoderma lucidum, an antitumor ganoderic acid (GA) producing basidiomycete, two homologs of a Cys2-His2 (C2H2)-type zinc finger transcription factor (CRZ1) - GlCRZ1 and GlCRZ2 were found by bioinformatic analysis. Using a recently developed CRlSPR/Cas9 genome editing platform, each gene of the two CRZ1 orthologs (glcrz1 and glcrz2) was disrupted to reveal their function in calcium signaling of GA biosynthesis. Compared to the wild type strain (WT), in which Ca2+ addition enhanced the GA production, disruption of glcrz1 did not hamper the cell growth but abolished the calcium signaling effect on stimulating GA biosynthesis. For the glcrz2 knocked-out strain, both the mycelial growth and GA synthesis were severely attenuated and its GA production declined even with Ca2+ treatment. These results indicated that the calcium signaling via GlCRZ1 and GlCRZ2 were critical in the calcineurin-mediated regulation of GA biosynthesis by G. lucidum. The work may be also helpful to understand the function of CRZ1 in secondary metabolism regulation in other basidiomycetes.