title: Cytoplasmic Drosha activity generated by alternative splicing.
abstract: RNase III enzyme Drosha interacts with DGCR8 to form the Microprocessor, initiating canonical microRNA (miRNA) maturation in the nucleus. Here, we re-evaluated where Drosha functions in cells using Drosha and/or DGCR8 knock out (KO) cells and cleavage reporters. Interestingly, a truncated Drosha mutant located exclusively in the cytoplasm cleaved pri-miRNA effectively in a DGCR8-dependent manner. In addition, we demonstrated that in vitro generated pri-miRNAs when transfected into cells could be processed to mature miRNAs in the cytoplasm. These results indicate the existence of cytoplasmic Drosha (c-Drosha) activity. Although a subset of endogenous pri-miRNAs become enriched in the cytoplasm of Drosha KO cells, it remains unclear whether pri-miRNA processing is the main function of c-Drosha. We identified two novel in-frame Drosha isoforms generated by alternative splicing in both HEK293T and HeLa cells. One isoform loses the putative nuclear localization signal, generating c-Drosha. Further analysis indicated that the c-Drosha isoform is abundant in multiple cell lines, dramatically variable among different human tissues and upregulated in multiple tumors, suggesting that c-Drosha plays a unique role in gene regulation. Our results reveal a new layer of regulation on the miRNA pathway and provide novel insights into the ever-evolving functions of Drosha.
authors.affiliation: Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Lisheng
authors.initials: L
authors.lastname: Dai
authors.affiliation: Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Kevin
authors.initials: K
authors.lastname: Chen
authors.affiliation: Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Brenda
authors.initials: B
authors.lastname: Youngren
authors.affiliation: Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Julia
authors.initials: J
authors.lastname: Kulina
authors.affiliation: Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: Acong
authors.initials: A
authors.lastname: Yang
authors.affiliation: Department of Electrical and Computer Engineering & TEES-AgriLife Center for Bioinformatics and Genomic Systems Engineering, Texas A&M University, College Station, TX 77843, USA.
authors.forename: Zhengyu
authors.initials: Z
authors.lastname: Guo
authors.affiliation: Department of Electrical and Computer Engineering & TEES-AgriLife Center for Bioinformatics and Genomic Systems Engineering, Texas A&M University, College Station, TX 77843, USA.
authors.forename: Jin
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Electrical and Computer Engineering & TEES-AgriLife Center for Bioinformatics and Genomic Systems Engineering, Texas A&M University, College Station, TX 77843, USA.
authors.forename: Peng
authors.initials: P
authors.lastname: Yu
authors.affiliation: Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA shuo.gu@nih.gov.
authors.forename: Shuo
authors.initials: S
authors.lastname: Gu
mesh_terms: D017398:Alternative Splicing; D003593:Cytoplasm / Q000378:metabolism; D004789:Enzyme Activation; D055786:Gene Knockout Techniques; D057809:HEK293 Cells; D006801:Humans; D006868:Hydrolysis; D007527:Isoenzymes; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D043244:Ribonuclease III / Q000235:genetics* / Q000378:metabolism*
chemical_list: D007527:Isoenzymes; D035683:MicroRNAs; D012333:RNA, Messenger; C517918:DROSHA protein, human; D043244:Ribonuclease III