Identification of an active RNAi pathway in .

RNA interference (RNAi) is a fundamental regulatory pathway with a wide range of functions, including regulation of gene expression and maintenance of genome stability. Although RNAi is widespread in the fungal kingdom, well-known species, such as the model yeast , have lost the RNAi pathway. Until now evidence has been lacking for a fully functional RNAi pathway in , a human fungal pathogen considered critically important by the World Health Organization. Here, we demonstrated that the widely used reference strain (SC5314) contains an inactivating missense mutation in the gene encoding for the central RNAi component Argonaute. In contrast, most other isolates contain a canonical Argonaute protein predicted to be functional and RNAi-active. Indeed, using high-throughput small and long RNA sequencing combined with seamless CRISPR/Cas9-based gene editing, we demonstrate that an active RNAi machinery represses expression of subtelomeric gene families. Thus, an intact and functional RNAi pathway exists in , highlighting the importance of using multiple reference strains when studying this dangerous pathogen.