A Calcium-Induced Strategy for Enhancing the Biosynthesis of the Characteristic Aroma in Shiitake Mushroom ( <i>Lentinula edodes</i> )

<i>Lentinula edodes</i> (shiitake mushroom) is renowned for its distinctive aroma, chiefly attributed to lenthionine (1,2,3,5,6-pentathiepane), a cyclic organosulfur compound. Exogenous calcium ion (Ca²⁺) application is known to stimulate secondary metabolite biosynthesis in plants and fungi. This study first tested the effects of calcium chloride (CaCl₂) on <i>L. edodes</i> through single-factor experiments, focusing on concentration and duration. It then optimized these conditions using response surface methodology (RSM), determining 8.18 mmol/L CaCl₂ applied for 16.02 days as the optimal parameters. This treatment elevated intracellular Ca²⁺ levels and upregulated the expression of key enzymes in lenthionine biosynthesis─γ-glutamyl transpeptidase (LEGGT) and cysteine sulfoxide lyase (LECSL)─resulting in enhanced lenthionine production. This study demonstrates that a low-cost Ca²⁺ inducer can upregulate the expression of key genes involved in lenthionine biosynthesis, thereby enhancing the catalytic activity of lenthionine-related enzymes and ultimately increasing lenthionine content for high-quality <i>L. edodes</i> production.