title: Dicer-TRBP complex formation ensures accurate mammalian microRNA biogenesis.
abstract: RNA-mediated gene silencing in human cells requires the accurate generation of ∼22 nt microRNAs (miRNAs) from double-stranded RNA substrates by the endonuclease Dicer. Although the phylogenetically conserved RNA-binding proteins TRBP and PACT are known to contribute to this process, their mode of Dicer binding and their genome-wide effects on miRNA processing have not been determined. We solved the crystal structure of the human Dicer-TRBP interface, revealing the structural basis of the interaction. Interface residues conserved between TRBP and PACT show that the proteins bind to Dicer in a similar manner and by mutual exclusion. Based on the structure, a catalytically active Dicer that cannot bind TRBP or PACT was designed and introduced into Dicer-deficient mammalian cells, revealing selective defects in guide strand selection. These results demonstrate the role of Dicer-associated RNA binding proteins in maintenance of gene silencing fidelity.
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
authors.forename: Ross C
authors.initials: RC
authors.lastname: Wilson
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
authors.forename: Akshay
authors.initials: A
authors.lastname: Tambe
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
authors.forename: Mary Anne
authors.initials: MA
authors.lastname: Kidwell
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
authors.forename: Cameron L
authors.initials: CL
authors.lastname: Noland
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
authors.forename: Catherine P
authors.initials: CP
authors.lastname: Schneider
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, CA 94720, USA; Department of Chemistry, University of California, Berkeley, CA 94720, USA; Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA. Electronic address: doudna@berkeley.edu.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Doudna
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000378:metabolism; D020134:Catalytic Domain; D002478:Cells, Cultured; D018360:Crystallography, X-Ray; D053487:DEAD-box RNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020868:Gene Silencing; D006801:Humans; D051379:Mice; D035683:MicroRNAs / Q000378:metabolism*; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D043244:Ribonuclease III / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D000099253:Trans-Activation Responsive RNA-Binding Protein
chemical_list: D060565:Argonaute Proteins; D053487:DEAD-box RNA Helicases; D035683:MicroRNAs; D016601:RNA-Binding Proteins; D043244:Ribonuclease III; D000099253:Trans-Activation Responsive RNA-Binding Protein; C104647:PRKRA protein, human; C408087:DICER1 protein, human; C509781:Dicer1 protein, mouse