Cytoplasmic and nuclear DROSHA in human villous trophoblasts

In villous trophoblasts, DROSHA is a key ribonuclease III enzyme that processes pri-microRNAs (pri-miRNAs) into pre-miRNAs at the placenta-specific, chromosome 19 miRNA cluster (C19MC) locus. However, little is known of its other functions. We performed formaldehyde crosslinking, immunoprecipitation, and sequencing (fCLIP-seq) analysis of terminal chorionic villi to identify DROSHA-binding RNAs in villous trophoblasts. In villous trophoblasts, DROSHA predominantly generated placenta-specific C19MC pre-miRNAs, including antiviral C19MC pre-miRNAs. The fCLIP-seq analysis also identified non-miRNA transcripts with hairpin structures potentially capable of binding to DROSHA (e.g., SNORD100 and VTRNA1-1). Moreover, in vivo immunohistochemical analysis revealed DROSHA in the cytoplasm of villous trophoblasts. DROSHA was abundant in the cytoplasm of villous trophoblasts, particularly in the apical region of syncytiotrophoblast, in the full-term placenta. Furthermore, in BeWo trophoblasts infected with Sindbis virus (SINV), DROSHA translocated to the cytoplasm and recognized the genomic RNA of SINV. Therefore, in trophoblasts, DROSHA not only regulates RNA metabolism, including the biogenesis of placenta-specific miRNAs, but also recognizes viral RNAs. After SINV infection, BeWo DROSHA-binding VTRNA1-1 was significantly upregulated, and cellular VTRNA1-1 was significantly downregulated, suggesting that DROSHA soaks up VTRNA1-1 in response to viral infection. These results suggest that the DROSHA-mediated recognition of RNAs defends against viral infection in villous trophoblasts. Our data provide insight into the antiviral functions of DROSHA in villous trophoblasts of the human placenta.