Ergosterol is an important component of fungal cell membrane. Ergosterol biosynthesis involves sterol C-14 reductase, a key enzyme in ergosterol biosynthesis, which has been well studied in Saccharomyces cerevisiae. However, little studies about this important enzyme in Aspergillus oryzae. In this study, two sterol C-14 reductases named AoErg24A and AoErg24B were identified in A. oryzae using bioinformatics analysis. Through phylogenetic tree, expression pattern, subcellular localization, and yeast functional complementation analyses, we discovered that both AoErg24A and AoErg24B are conserved and localized to the endoplasmic reticulum (ER). Both enzymes can partially restore the temperature sensitivity phenotype of a S. cerevisiae erg24 weak mutant. Overexpression of AoErg24A in A. oryzae increased 1.6 times of ergosterol content, while overexpression of AoErg24B led to a slight decrease of ergosterol. Both genes affect the sporulation of A. oryzae. These results uncovered that the two genes function differently in ergosterol biosynthesis. Thus, this study further enhances our understanding of ergosterol biosynthesis in A. oryzae and lays a good foundation for A. oryzae to be used in industrial ergosterol production.

麦角甾醇是真菌细胞膜的重要成分。麦角甾醇生物合成涉及甾醇C-14还原酶，它是麦角甾醇生物合成中的关键酶，已在酿酒酵母中得到充分研究。然而，对于米曲霉中这一重要酶的研究却很少。在本研究中，利用生物信息学分析在米曲霉中鉴定出两种甾醇 C-14 还原酶，名为AoErg24A和AoErg24B 。通过系统发育树、表达模式、亚细胞定位和酵母功能互补分析，我们发现AoErg24A和AoErg24B都是保守的并定位于内质网（ER）。两种酶都可以部分恢复酿酒酵母 erg24弱突变体的温度敏感性表型。米曲霉中AoErg24A的过表达使麦角甾醇含量增加1.6倍，而AoErg24B的过表达导致麦角甾醇含量略有下降。这两个基因都会影响米曲霉的孢子形成。这些结果揭示了这两个基因在麦角甾醇生物合成中的功能不同。因此，本研究进一步加深了我们对米曲霉麦角甾醇生物合成的认识，为米曲霉应用于工业麦角甾醇生产奠定了良好的基础。