Abstract
Targeted gene silencing by RNA interference (RNAi) requires loading of a short guide RNA (small interfering RNA (siRNA) or microRNA (miRNA)) onto an Argonaute protein to form the functional center of an RNA-induced silencing complex (RISC). In humans, Argonaute2 (AGO2) assembles with the guide RNA-generating enzyme Dicer and the RNA-binding protein TRBP to form a RISC-loading complex (RLC), which is necessary for efficient transfer of nascent siRNAs and miRNAs from Dicer to AGO2. Here, using single-particle EM analysis, we show that human Dicer has an L-shaped structure. The RLC Dicer's N-terminal DExH/D domain, located in a short 'base branch', interacts with TRBP, whereas its C-terminal catalytic domains in the main body are proximal to AGO2. A model generated by docking the available atomic structures of Dicer and Argonaute homologs into the RLC reconstruction suggests a mechanism for siRNA transfer from Dicer to AGO2.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Argonaute Proteins
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Eukaryotic Initiation Factor-2 / genetics
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Eukaryotic Initiation Factor-2 / metabolism
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Eukaryotic Initiation Factor-2 / ultrastructure
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Humans
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MicroRNAs / genetics
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MicroRNAs / metabolism
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Microscopy, Electron
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Models, Biological
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Protein Binding / genetics
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RNA, Small Interfering / genetics
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RNA, Small Interfering / metabolism
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RNA-Binding Proteins / genetics
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RNA-Binding Proteins / metabolism
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RNA-Binding Proteins / ultrastructure
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RNA-Induced Silencing Complex / chemistry*
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RNA-Induced Silencing Complex / metabolism*
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RNA-Induced Silencing Complex / ultrastructure
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Ribonuclease III / genetics
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Ribonuclease III / ultrastructure
Substances
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AGO2 protein, human
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Argonaute Proteins
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Eukaryotic Initiation Factor-2
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MicroRNAs
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RNA, Small Interfering
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RNA-Binding Proteins
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RNA-Induced Silencing Complex
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trans-activation responsive RNA-binding protein
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Ribonuclease III