期刊
MOLECULAR CELL
卷 66, 期 2, 页码 258-+出版社
CELL PRESS
DOI: 10.1016/j.molcel.2017.03.013
关键词
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资金
- Institute for Basic Science from the Ministry of Science, ICT and Future Planning of Korea [IBS-R008-D1]
- BK21 Research Fellowships from the Ministry of Education of Korea
- Ministry of Science & ICT (MSIT), Republic of Korea [IBS-R008-D1-2017-A00] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
MicroRNA (miRNA) maturation is initiated by DROSHA, a double-stranded RNA (dsRNA)-specific RNase III enzyme. By cleaving primary miRNAs (pri-miRNAs) at specific positions, DROSHA serves as a main determinant of miRNA sequences and a highly selective gatekeeper for the canonical miRNA pathway. However, the sites of DROSHA-mediated processing have not been annotated, and it remains unclear to what extent DROSHA functions outside the miRNA pathway. Here, we establish a protocol termed formaldehyde crosslinking, immunoprecipitation, and sequencing (fCLIP-seq), which allows identification of DROSHA cleavage sites at single-nucleotide resolution. fCLIP identifies numerous processing sites, suggesting widespread end modifications during miRNA maturation. fCLIP also finds many pri-miRNAs that undergo alternative processing, yielding multiple miRNA isoforms. Moreover, we discovered dozens of DROSHA substrates on non-miRNA loci, which may serve as cis-elements for DROSHA-mediated gene regulation. We anticipate that fCLIP-seq could be a general tool for investigating interactions between dsRNA-binding proteins and structured RNAs.
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