title: Structure of C3PO and mechanism of human RISC activation.
abstract: Assembly of the RNA-induced silencing complex (RISC) consists of loading duplex (guide-passenger) siRNA onto Argonaute (Ago2) and removing the passenger strand. Ago2 contributes critically to RISC activation by nicking the passenger strand. Here we reconstituted duplex siRNA-initiated RISC activity using recombinant human Ago2 (hAgo2) and C3PO, indicating that C3PO has a critical role in hAgo2-RISC activation. Consistently, genetic depletion of C3PO compromised RNA silencing in mammalian cells. We determined the crystal structure of hC3PO, which reveals an asymmetric octamer barrel consisting of six translin and two TRAX subunits. This asymmetric assembly is critical for the function of C3PO as an endonuclease that cleaves RNA at the interior surface. The current work supports a Dicer-independent mechanism for human RISC activation, in which Ago2 directly binds duplex siRNA and nicks the passenger strand, and then C3PO activates RISC by degrading the Ago2-nicked passenger strand.
authors.affiliation: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas, USA.
authors.forename: Xuecheng
authors.initials: X
authors.lastname: Ye
authors.forename: Nian
authors.initials: N
authors.lastname: Huang
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.forename: Zain
authors.initials: Z
authors.lastname: Paroo
authors.forename: Carlos
authors.initials: C
authors.lastname: Huerta
authors.forename: Peng
authors.initials: P
authors.lastname: Li
authors.forename: She
authors.initials: S
authors.lastname: Chen
authors.forename: Qinghua
authors.initials: Q
authors.lastname: Liu
authors.forename: Hong
authors.initials: H
authors.lastname: Zhang
mesh_terms: D060565:Argonaute Proteins; D002268:Carboxypeptidases / Q000096:biosynthesis*; D018360:Crystallography, X-Ray; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004722:Endoribonucleases / Q000737:chemistry / Q000378:metabolism*; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D034741:RNA, Small Interfering / Q000378:metabolism*
chemical_list: C466422:AGO2 protein, human; D060565:Argonaute Proteins; D004268:DNA-Binding Proteins; D015852:Eukaryotic Initiation Factor-2; D034741:RNA, Small Interfering; C095563:TSN protein, human; C498305:TSNAX protein, human; D004722:Endoribonucleases; D002268:Carboxypeptidases; C436327:SCPEP1 protein, human