title: SKI2 mediates degradation of RISC 5'-cleavage fragments and prevents secondary siRNA production from miRNA targets in Arabidopsis.
abstract: Small regulatory RNAs are fundamental in eukaryotic and prokaryotic gene regulation. In plants, an important element of post-transcriptional control is effected by 20-24 nt microRNAs (miRNAs) and short interfering RNAs (siRNAs) bound to the ARGONAUTE1 (AGO1) protein in an RNA induced silencing complex (RISC). AGO1 may cleave target mRNAs with small RNA complementarity, but the fate of the resulting cleavage fragments remains incompletely understood. Here, we show that SKI2, SKI3 and SKI8, subunits of a cytoplasmic cofactor of the RNA exosome, are required for degradation of RISC 5', but not 3'-cleavage fragments in Arabidopsis. In the absence of SKI2 activity, many miRNA targets produce siRNAs via the RNA-dependent RNA polymerase 6 (RDR6) pathway. These siRNAs are low-abundant, and map close to the cleavage site. In most cases, siRNAs were produced 5' to the cleavage site, but several examples of 3'-spreading were also identified. These observations suggest that siRNAs do not simply derive from RDR6 action on stable 5'-cleavage fragments and hence that SKI2 has a direct role in limiting secondary siRNA production in addition to its function in mediating degradation of 5'-cleavage fragments.
authors.affiliation: Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200 Copenhagen N, Denmark.
authors.forename: Anja
authors.initials: A
authors.lastname: Branscheid
authors.affiliation: Swiss Federal Institute of Technology (ETH) Zürich, Department of Biology, LFW D17/D18, Universitätsstrasse 2, CH-8092 Zürich, Switzerland.
authors.forename: Antonin
authors.initials: A
authors.lastname: Marchais
authors.affiliation: Swiss Federal Institute of Technology (ETH) Zürich, Department of Biology, LFW D17/D18, Universitätsstrasse 2, CH-8092 Zürich, Switzerland.
authors.forename: Gregory
authors.initials: G
authors.lastname: Schott
authors.affiliation: Institut de Biologie Moléculaire des Plantes du CNRS, 12 Rue du Général Zimmer, F-67084 Strasbourg Cedex, France.
authors.forename: Heike
authors.initials: H
authors.lastname: Lange
authors.affiliation: Institut de Biologie Moléculaire des Plantes du CNRS, 12 Rue du Général Zimmer, F-67084 Strasbourg Cedex, France.
authors.forename: Dominique
authors.initials: D
authors.lastname: Gagliardi
authors.affiliation: Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10, DK-8000 Aarhus C, Denmark.
authors.forename: Stig Uggerhøj
authors.initials: SU
authors.lastname: Andersen
authors.affiliation: Swiss Federal Institute of Technology (ETH) Zürich, Department of Biology, LFW D17/D18, Universitätsstrasse 2, CH-8092 Zürich, Switzerland.
authors.forename: Olivier
authors.initials: O
authors.lastname: Voinnet
authors.affiliation: Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200 Copenhagen N, Denmark pbrodersen@bio.ku.dk.
authors.forename: Peter
authors.initials: P
authors.lastname: Brodersen
mesh_terms: D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000502:physiology*; D035683:MicroRNAs / Q000378:metabolism*; D009154:Mutation; D020365:RNA Helicases / Q000235:genetics / Q000502:physiology*; D034741:RNA, Small Interfering / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics; D034743:RNA-Induced Silencing Complex / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D035683:MicroRNAs; D034741:RNA, Small Interfering; D034743:RNA-Induced Silencing Complex; C000608437:SKI3 protein, Arabidopsis; C513197:SKI8 protein, Arabidopsis; C514575:RDR6 protein, Arabidopsis; D012324:RNA-Dependent RNA Polymerase; C510422:RAX3 protein, Arabidopsis; D020365:RNA Helicases