title: Single processing center models for human Dicer and bacterial RNase III.
abstract: Dicer is a multidomain ribonuclease that processes double-stranded RNAs (dsRNAs) to 21 nt small interfering RNAs (siRNAs) during RNA interference, and excises microRNAs from precursor hairpins. Dicer contains two domains related to the bacterial dsRNA-specific endonuclease, RNase III, which is known to function as a homodimer. Based on an X-ray structure of the Aquifex aeolicus RNase III, models of the enzyme interaction with dsRNA, and its cleavage at two composite catalytic centers, have been proposed. We have generated mutations in human Dicer and Escherichia coli RNase III residues implicated in the catalysis, and studied their effect on RNA processing. Our results indicate that both enzymes have only one processing center, containing two RNA cleavage sites and generating products with 2 nt 3' overhangs. Based on these and other data, we propose that Dicer functions through intramolecular dimerization of its two RNase III domains, assisted by the flanking RNA binding domains, PAZ and dsRBD.
authors.affiliation: Friedrich Miescher Institute for Biomedical Research, PO Box 2543, 4002 Basel, Switzerland.
authors.forename: Haidi
authors.initials: H
authors.lastname: Zhang
authors.forename: Fabrice A
authors.initials: FA
authors.lastname: Kolb
authors.forename: Lukasz
authors.initials: L
authors.lastname: Jaskiewicz
authors.forename: Eric
authors.initials: E
authors.lastname: Westhof
authors.forename: Witold
authors.initials: W
authors.lastname: Filipowicz
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D017124:Conserved Sequence; D053487:DEAD-box RNA Helicases; D019281:Dimerization; D004722:Endoribonucleases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D006801:Humans; D008345:Manganese / Q000378:metabolism; D035683:MicroRNAs / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020365:RNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012323:RNA Processing, Post-Transcriptional; D012330:RNA, Double-Stranded / Q000737:chemistry / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D043244:Ribonuclease III / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
chemical_list: D035683:MicroRNAs; D012330:RNA, Double-Stranded; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D008345:Manganese; D004722:Endoribonucleases; C408087:DICER1 protein, human; D043244:Ribonuclease III; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases