Abstract:RNA interference (RNAi) is a conserved eukaryotic mechanism that uses small non-coding RNAs to mediate transcriptional/post-transcriptional gene silencing. Despite the deficiency of RNAi mechanism in some fungi, evidence accumulates that fungal RNAi not only maintains genome integrity but also plays a pivotal role in the regulation of fungal growth and development, heterochromatin formation, centromere evolution, and development of fungal drug resistance and pathogenicity. This review summarizes the biological functions of RNAi in fungi, which is expected to lay a basis for further research on the mechanism of fungal RNAi.