Short RNA duplexes guide sequence-dependent cleavage by human Dicer.
Dicer is a member of the double-stranded (ds) RNA-specific ribonuclease III (RNase III) family that is required for RNA processing and degradation. Like most members of the RNase III family, Dicer possesses a dsRNA binding domain and cleaves long RNA duplexes in vitro. In this study, Dicer substrate selectivity was examined using bipartite substrates. These experiments revealed that an RNA helix possessing a 2-nucleotide (nt) 3'-overhang may bind and direct sequence-specific Dicer-mediated cleavage in trans at a fixed distance from the 3'-end overhang. Chemical modifications of the substrate indicate that the presence of the ribose 2'-hydroxyl group is not required for Dicer binding, but some located near the scissile bonds are needed for RNA cleavage. This suggests a flexible mechanism for substrate selectivity that recognizes the overall shape of an RNA helix. Examination of the structure of natural pre-microRNAs (pre-miRNAs) suggests that they may form bipartite substrates with complementary mRNA sequences, and thus induce seed-independent Dicer cleavage. Indeed, in vitro, natural pre-miRNA directed sequence-specific Dicer-mediated cleavage in trans by supporting the formation of a substrate mimic.
Bergeron
Lucien
L
RNA group/Groupe ARN, Université de Sherbrooke, Sherbrooke, Québec J1H 5N4, Canada.
Perreault
Jean-Pierre
JP
Abou Elela
Sherif
S
D001482:Base Composition; D001483:Base Sequence / Q000502:physiology*; D001665:Binding Sites; D002478:Cells, Cultured; D053487:DEAD-box RNA Helicases / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D035683:MicroRNAs / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D012323:RNA Processing, Post-Transcriptional; D012330:RNA, Double-Stranded / Q000737:chemistry / Q000378:metabolism*; D043244:Ribonuclease III / Q000378:metabolism* / Q000502:physiology; D058727:RNA, Small Untranslated
D035683:MicroRNAs; D012330:RNA, Double-Stranded; C408087:DICER1 protein, human; D043244:Ribonuclease III; D053487:DEAD-box RNA Helicases; D058727:RNA, Small Untranslated