title: The Vasa Homolog RDE-12 engages target mRNA and multiple argonaute proteins to promote RNAi in C. elegans.
abstract: Argonaute (AGO) proteins are key nuclease effectors of RNAi. Although purified AGOs can mediate a single round of target RNA cleavage in vitro, accessory factors are required for small interfering RNA (siRNA) loading and to achieve multiple-target turnover. To identify AGO cofactors, we immunoprecipitated the C. elegans AGO WAGO-1, which engages amplified small RNAs during RNAi. These studies identified a robust association between WAGO-1 and a conserved Vasa ATPase-related protein RDE-12. rde-12 mutants are deficient in RNAi, including viral suppression, and fail to produce amplified secondary siRNAs and certain endogenous siRNAs (endo-siRNAs). RDE-12 colocalizes with WAGO-1 in germline P granules and in cytoplasmic and perinuclear foci in somatic cells. These findings and our genetic studies suggest that RDE-12 is first recruited to target mRNA by upstream AGOs (RDE-1 and ERGO-1), where it promotes small RNA amplification and/or WAGO-1 loading. Downstream of these events, RDE-12 forms an RNase-resistant (target mRNA-independent) complex with WAGO-1 and may thus have additional functions in target mRNA surveillance and silencing.
authors.affiliation: Program in Molecular Medicine, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; RNA Therapeutics Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.
authors.forename: Masaki
authors.initials: M
authors.lastname: Shirayama
authors.affiliation: Program in Molecular Medicine, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; RNA Therapeutics Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.
authors.forename: William
authors.initials: W
authors.lastname: Stanney
authors.affiliation: Program in Molecular Medicine, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; RNA Therapeutics Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.
authors.forename: Weifeng
authors.initials: W
authors.lastname: Gu
authors.affiliation: Program in Molecular Medicine, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; RNA Therapeutics Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.
authors.forename: Meetu
authors.initials: M
authors.lastname: Seth
authors.affiliation: Program in Molecular Medicine, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; RNA Therapeutics Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA; Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA. Electronic address: craig.mello@umassmed.edu.
authors.forename: Craig C
authors.initials: CC
authors.lastname: Mello
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060565:Argonaute Proteins / Q000378:metabolism*; D015152:Blotting, Northern; D017173:Caenorhabditis elegans / Q000502:physiology*; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism*; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D015151:Immunoblotting; D047468:Immunoprecipitation; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D034622:RNA Interference / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D060565:Argonaute Proteins; D029742:Caenorhabditis elegans Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D053487:DEAD-box RNA Helicases; C000600925:RDE-12 protein, C elegans