RNA interference (RNAi) regulates gene expression by the cleavage of messenger RNA, by mRNA degradation and by preventing protein synthesis. These effects are mediated by a ribonucleoprotein complex known as RISC (RNA-induced silencing complex)1. We have previously identified four Drosophila components (short interfering RNAs1, Argonaute 2 (ref. 2), VIG and FXR3) of a RISC enzyme that degrades specific mRNAs in response to a double-stranded-RNA trigger. Here we show that Tudor-SN (tudor staphylococcal nuclease)—a protein containing five staphylococcal/micrococcal nuclease domains and a tudor domain—is a component of the RISC enzyme in Caenorhabditis elegans, Drosophila and mammals. Although Tudor-SN contains non-canonical active-site sequences, we show that purified Tudor-SN exhibits nuclease activity similar to that of other staphylococcal nucleases. Notably, both purified Tudor-SN and RISC are inhibited by a specific competitive inhibitor of micrococcal nuclease. Tudor-SN is the first RISC subunit to be identified that contains a recognizable nuclease domain, and could therefore contribute to the RNA degradation observed in RNAi.
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The authors declare that they have no competing financial interests.
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We thank A. Mildvan, M. Tijsterman and T. Sijen for discussions. We thank T. Keenan for an anti-p100 antibody, T. Hobman for GERP (EIF2C2/hAgo2) antibody, H. Siomi for a FXR monoclonal antibody, and F. Slack for the lacZ–lin-41 reporter. A.A.C. is a George A. and Marjorie H. Anderson Fellow of the Watson School of Biological Sciences, and a Howard Hughes Medical Institute Predoctoral Fellow. A.M.D. is a David Koch Fellow of the Watson School of Biological Sciences. J.M.S. is supported by a postdoctoral fellowship from the US Army Prostate Cancer Research programme. G.J.H. is a Rita Allen Foundation Scholar and is supported by an Innovator Award from the US Army Breast Cancer Research programme. This work was also supported by a grant from the National Institutes of Health (G.J.H.) and by a VENI fellowship from the Netherlands Organization for Scientific Research (RFK).
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