Multiomics Analysis across the Life Cycle Identifies Zn₂Cys₆_61 as a Target for Enhancing Triterpenoid Production in...

Ganoderic acids (GAs) are high-value lanostane-type triterpenoids derived from Ganoderma lucidum (G. lucidum) with broad applications in functional foods and nutraceuticals, yet their low natural abundance limits industrial production. In this study, an integrated life-cycle multiomics analysis combining metabolomics, transcriptomics, and proteomics was conducted across six developmental stages in four G. lucidum strains to elucidate regulatory mechanisms governing GA biosynthesis. Weighted gene coexpression network analysis identified candidate cytochrome P450 enzymes and transcription factors associated with GA accumulation. A Zn₂Cys₆-type transcription factor, Zn₂Cys₆_61, was identified as a central regulator and functionally validated through overexpression and RNA interference. Genetic manipulation of Zn₂Cys₆_61 expression significantly altered GA levels, with overexpression markedly enhancing GA accumulation. Further analysis demonstrated that Zn₂Cys₆_61 directly binds to and activates the promoter of squalene synthase, a key enzyme in triterpenoid backbone biosynthesis. Together, these findings identify Zn₂Cys₆_61 as an effective engineering target and provide a transcription factor-based strategy for improving GA production in medicinal mushrooms.