title: Argonaute proteins regulate HIV-1 multiply spliced RNA and viral production in a Dicer independent manner.
abstract: Argonaute (Ago) proteins associate with microRNAs (miRNAs) to form the core of the RNA-induced silencing complex (RISC) that mediates post-transcriptional gene silencing of target mRNAs. As key players in anti-viral defense, Ago proteins are thought to have the ability to interact with human immunodeficiency virus type 1 (HIV-1) RNA. However, the role of this interaction in regulating HIV-1 replication has been debated. Here, we used high throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) to explore the interaction between Ago2 and HIV-1 RNA in infected cells. By only considering reads of 50 nucleotides length in our analysis, we identified more than 30 distinct binding sites for Ago2 along the viral RNA genome. Using reporter assays, we found four binding sites, located near splice donor sites, capable of repressing Luciferase gene expression in an Ago-dependent manner. Furthermore, inhibition of Ago1 and Ago2 levels in cells expressing HIV-1 led to an increase of viral multiply spliced transcripts and to a strong reduction in the extracellular CAp24 level. Depletion of Dicer did not affect these activities. Our results highlight a new role of Ago proteins in the control of multiply spliced HIV-1 transcript levels and viral production, independently of the miRNA pathway.
authors.affiliation: INSERM, U1016, Institut Cochin, Paris 75014, France.|CNRS, UMR8104, Paris 75014, France.|Université Paris Descartes, Sorbonne Paris Cité, Paris 75006, France.
authors.forename: Agathe
authors.initials: A
authors.lastname: Eckenfelder
authors.affiliation: INSERM, U1016, Institut Cochin, Paris 75014, France.|CNRS, UMR8104, Paris 75014, France.|Université Paris Descartes, Sorbonne Paris Cité, Paris 75006, France.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Ségéral
authors.affiliation: CNRS, UPR 1142, Institut de Génétique Humaine, Montpellier 34396, France.
authors.forename: Natalia
authors.initials: N
authors.lastname: Pinzón
authors.affiliation: CNAM, Laboratoire Génomique, Bioinformatique et Applications (EA 4627), Paris 75003, France.
authors.forename: Damien
authors.initials: D
authors.lastname: Ulveling
authors.affiliation: INSERM, U1016, Institut Cochin, Paris 75014, France.|CNRS, UMR8104, Paris 75014, France.|Université Paris Descartes, Sorbonne Paris Cité, Paris 75006, France.
authors.forename: Céline
authors.initials: C
authors.lastname: Amadori
authors.affiliation: INSERM, U1154, CNRS, UMR7196, Museum National d'Histoire Naturelle, Paris 75231, France.
authors.forename: Marine
authors.initials: M
authors.lastname: Charpentier
authors.affiliation: Plateforme MGX, Institut de Génomique Fonctionnelle, CNRS, UMR5203, INSERM, U661, Montpellier 34094, France.
authors.forename: Sabine
authors.initials: S
authors.lastname: Nidelet
authors.affiliation: INSERM, U1154, CNRS, UMR7196, Museum National d'Histoire Naturelle, Paris 75231, France.
authors.forename: Jean-Paul
authors.initials: JP
authors.lastname: Concordet
authors.affiliation: CNAM, Laboratoire Génomique, Bioinformatique et Applications (EA 4627), Paris 75003, France.
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Zagury
authors.affiliation: Architecture et Réactivité de l'ARN, Université de Strasbourg, CNRS, IBMC, Strasbourg 67084, France.
authors.forename: Jean-Christophe
authors.initials: JC
authors.lastname: Paillart
authors.affiliation: INSERM, U1016, Institut Cochin, Paris 75014, France.|CNRS, UMR8104, Paris 75014, France.|Université Paris Descartes, Sorbonne Paris Cité, Paris 75006, France.
authors.forename: Clarisse
authors.initials: C
authors.lastname: Berlioz-Torrent
authors.affiliation: CNRS, UPR 1142, Institut de Génétique Humaine, Montpellier 34396, France.
authors.forename: Hervé
authors.initials: H
authors.lastname: Seitz
authors.affiliation: INSERM, U1016, Institut Cochin, Paris 75014, France.|CNRS, UMR8104, Paris 75014, France.|Université Paris Descartes, Sorbonne Paris Cité, Paris 75006, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Emiliani
authors.affiliation: INSERM, U1016, Institut Cochin, Paris 75014, France.|CNRS, UMR8104, Paris 75014, France.|Université Paris Descartes, Sorbonne Paris Cité, Paris 75006, France.
authors.forename: Sarah
authors.initials: S
authors.lastname: Gallois-Montbrun
mesh_terms: D017398:Alternative Splicing; D060565:Argonaute Proteins / Q000378:metabolism*; D001665:Binding Sites; D053487:DEAD-box RNA Helicases / Q000378:metabolism; D039642:Eukaryotic Initiation Factors / Q000378:metabolism; D016679:Genome, Viral; D057809:HEK293 Cells; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology; D006367:HeLa Cells; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D047468:Immunoprecipitation; D019169:Jurkat Cells; D012322:RNA Precursors / Q000378:metabolism; D022821:RNA Splice Sites; D012333:RNA, Messenger / Q000378:metabolism*; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism*; D043244:Ribonuclease III / Q000378:metabolism; D017423:Sequence Analysis, RNA; D014771:Virion / Q000502:physiology
chemical_list: C115848:AGO1 protein, human; C466422:AGO2 protein, human; D060565:Argonaute Proteins; D039642:Eukaryotic Initiation Factors; D012322:RNA Precursors; D022821:RNA Splice Sites; D012333:RNA, Messenger; D012367:RNA, Viral; C408087:DICER1 protein, human; D043244:Ribonuclease III; D053487:DEAD-box RNA Helicases