Metal ions affect cell physiology and metabolism significantly, but the role of Mn2+ in the secondary metabolism of mushrooms is yet unclear. In static liquid cultivation of Ganoderma lucidum for producing antitumor ganoderic acids (GAs), the Mn2+ addition was performed. Addition of 10 mM Mn2+ at the start of the static liquid cultivation resulted in 2.2-fold improvement of total GAs production. The expression levels of GA biosynthetic and Ca2+ sensors' genes were up-regulated with Mn2+ induction while down-regulated by adding cyclosporin A (calcineurin inhibitor), suggesting that higher GA production might result from calcineurin signal regulation. Intracellular Ca2+ imaging and calcineurin inhibitor study revealed that addition of Mn2+ led to Ca2+ influx from medium to the cells to trigger calcineurin signals. Mn2+ addition was therefore an efficient induction strategy for improving GAs production, whose regulation mechanism was via calcineurin signaling transduction. (C) 2014 Wiley Periodicals, Inc.