title: Short-interfering-RNA-mediated gene silencing in mammalian cells requires Dicer and eIF2C translation initiation factors.
abstract: RNA interference (RNAi) is the process of long, double-stranded (ds), RNA-dependent posttranscriptional gene silencing (PTGS). In lower eukaryotes, dsRNA introduced into the cytoplasm is cleaved by the RNaseIII-like enzyme, Dicer, to 21-23 nt RNA (short interfering [si] RNA), which may serve as guide for target mRNA degradation. In mammals, long-dsRNA-dependent PTGS is applicable only to a limited number of cell types, whereas siRNA synthesized in vitro is capable of effectively inducing gene silencing in a wide variety of cells. Although biochemical and genetic analyses in lower eukaryotes showed that Dicer and some PIWI family member proteins are essential for long-dsRNA-dependent PTGS, little is known about the molecular mechanisms underlying siRNA-based PTGS. Here, we show that Dicer and eIF2C translation initiation factors belonging to the PIWI family (eIF2C1-4) play an essential role in mammalian siRNA-mediated PTGS, most probably through synergistic interactions. Immunoprecipitation experiments suggest that, in human and mouse cells, complex formation occurs between Dicer and eIF2C1 or 2 and that the PIWI domain of eIF2C is essential for the formation of this complex.
authors.affiliation: Department of Biophysics, Graduate School of Science, University of Tokyo, 7-3-1 Hongo, 113-0033, Bunkyo-ku, Tokyo, Japan.
authors.forename: Noboru
authors.initials: N
authors.lastname: Doi
authors.forename: Shuhei
authors.initials: S
authors.lastname: Zenno
authors.forename: Ryu
authors.initials: R
authors.lastname: Ueda
authors.forename: Hiroko
authors.initials: H
authors.lastname: Ohki-Hamazaki
authors.forename: Kumiko
authors.initials: K
authors.lastname: Ui-Tei
authors.forename: Kaoru
authors.initials: K
authors.lastname: Saigo
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D060565:Argonaute Proteins; D004722:Endoribonucleases / Q000235:genetics / Q000378:metabolism*; D039642:Eukaryotic Initiation Factors; D020868:Gene Silencing / Q000502:physiology*; D016259:Genes, myc; D006367:HeLa Cells; D006801:Humans; D046911:Macromolecular Substances; D008322:Mammals / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D010448:Peptide Initiation Factors / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D014176:Protein Biosynthesis / Q000502:physiology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000502:physiology*; D043244:Ribonuclease III
chemical_list: C115848:AGO1 protein, human; D060565:Argonaute Proteins; D039642:Eukaryotic Initiation Factors; D046911:Macromolecular Substances; D010448:Peptide Initiation Factors; D034741:RNA, Small Interfering; D004722:Endoribonucleases; D043244:Ribonuclease III