Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.
Crystal Structure of Argonaute and Its Implications for RISC Slicer Activity
Ji-Joon Song,1,2Stephanie K. Smith,2Gregory J. Hannon,1Leemor Joshua-Tor1,2*
Argonaute proteins and small interfering RNAs (siRNAs) are theknown signature components of the RNA interference effectorcomplex RNA-induced silencing complex (RISC). However, the identityof "Slicer," the enzyme that cleaves the messenger RNA (mRNA)as directed by the siRNA, has not been resolved. Here, we reportthe crystal structure of the Argonaute protein from Pyrococcusfuriosus at 2.25 angstrom resolution. The structure revealsa crescent-shaped base made up of the amino-terminal, middle,and PIWI domains. The Piwi Argonaute Zwille (PAZ) domain isheld above the base by a "stalk"-like region. The PIWI domain(named for the protein piwi) is similar to ribonuclease H, witha conserved active site aspartate-aspartate-glutamate motif,strongly implicating Argonaute as "Slicer." The architectureof the molecule and the placement of the PAZ and PIWI domainsdefine a groove for substrate binding and suggest a mechanismfor siRNA-guided mRNA cleavage.
1 Watson School of Biological Sciences, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA. 2 Keck Structural Biology Laboratory, Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.
* To whom correspondence should be addressed. E-mail: leemor{at}cshl.edu
The editors suggest the following Related Resources on Science sites:
In Science Magazine
RESEARCH ARTICLES
Jidong Liu, Michelle A. Carmell, Fabiola V. Rivas, Carolyn G. Marsden, J. Michael Thomson, Ji-Joon Song, Scott M. Hammond, Leemor Joshua-Tor, and Gregory J. Hannon (3 September 2004) Science305 (5689), 1437.
[DOI: 10.1126/science.1102513] |Abstract »|Full Text »|PDF »|Supporting Online Material »
PERSPECTIVES
Erik J. Sontheimer and Richard W. Carthew (3 September 2004) Science305 (5689), 1409.
[DOI: 10.1126/science.1103076] |Summary »|Full Text »|PDF »
p100 increases AT1R expression through interaction with AT1R 3'-UTR.
K. Paukku, N. Kalkkinen, O. Silvennoinen, K. K. Kontula, and J. Y. A. Lehtonen (2008)
Nucleic Acids Res.
36, 4474-4487
|Abstract »|Full Text »|PDF »
An Argonaute Transports siRNAs from the Cytoplasm to the Nucleus.
S. Guang, A. F. Bochner, D. M. Pavelec, K. B. Burkhart, S. Harding, J. Lachowiec, and S. Kennedy (2008)
Science
321, 537-541
|Abstract »|Full Text »|PDF »
A multifunctional human Argonaute2-specific monoclonal antibody.
S. Rudel, A. Flatley, L. Weinmann, E. Kremmer, and G. Meister (2008)
RNA
14, 1244-1253
|Abstract »|Full Text »|PDF »
The PIWI proteins SMEDWI-2 and SMEDWI-3 are required for stem cell function and piRNA expression in planarians.
D. Palakodeti, M. Smielewska, Y.-C. Lu, G. W. Yeo, and B. R. Graveley (2008)
RNA
14, 1174-1186
|Abstract »|Full Text »|PDF »
Fundamental differences in the equilibrium considerations for siRNA and antisense oligodeoxynucleotide design.
Artificial trans-Acting siRNAs Confer Consistent and Effective Gene Silencing.
M. de la Luz Gutierrez-Nava, M. J. Aukerman, H. Sakai, S. V. Tingey, and R. W. Williams (2008)
Plant Physiology
147, 543-551
|Abstract »|Full Text »|PDF »
Widespread Translational Inhibition by Plant miRNAs and siRNAs.
P. Brodersen, L. Sakvarelidze-Achard, M. Bruun-Rasmussen, P. Dunoyer, Y. Y. Yamamoto, L. Sieburth, and O. Voinnet (2008)
Science
320, 1185-1190
|Abstract »|Full Text »|PDF »
Diversification of the Core RNA Interference Machinery in Chlamydomonas reinhardtii and the Role of DCL1 in Transposon Silencing.
J. A. Casas-Mollano, J. Rohr, E.-J. Kim, E. Balassa, K. van Dijk, and H. Cerutti (2008)
Genetics
179, 69-81
|Abstract »|Full Text »|PDF »
Small RNA guides for de novo DNA methylation in mammalian germ cells.
Chp1-Tas3 Interaction Is Required To Recruit RITS to Fission Yeast Centromeres and for Maintenance of Centromeric Heterochromatin.
J. L. DeBeauchamp, A. Moses, V. J. P. Noffsinger, D. L. Ulrich, G. Job, A. M. Kosinski, and J. F. Partridge (2008)
Mol. Cell. Biol.
28, 2154-2166
|Abstract »|Full Text »|PDF »
Functional dissection of siRNA sequence by systematic DNA substitution: modified siRNA with a DNA seed arm is a powerful tool for mammalian gene silencing with significantly reduced off-target effect.
K. Ui-Tei, Y. Naito, S. Zenno, K. Nishi, K. Yamato, F. Takahashi, A. Juni, and K. Saigo (2008)
Nucleic Acids Res.
36, 2136-2151
|Abstract »|Full Text »|PDF »
aubergine Gene Overexpression in Somatic Tissues of auberginesting Mutants Interferes With the RNAi Pathway of a yellow Hairpin dsRNA in Drosophila melanogaster.
V. Specchia, C. Benna, G. M. Mazzotta, A. Piccin, M. A. Zordan, R. Costa, and M. P. Bozzetti (2008)
Genetics
178, 1271-1282
|Abstract »|Full Text »|PDF »
Strand-specific 5'-O-methylation of siRNA duplexes controls guide strand selection and targeting specificity.
P. Y. Chen, L. Weinmann, D. Gaidatzis, Y. Pei, M. Zavolan, T. Tuschl, and G. Meister (2008)
RNA
14, 263-274
|Abstract »|Full Text »|PDF »
RNA Silencing Gene Truncation in the Filamentous Fungus Aspergillus nidulans.
T. M. Hammond, J. W. Bok, M. D. Andrewski, Y. Reyes-Dominguez, C. Scazzocchio, and N. P. Keller (2008)
Eukaryot. Cell
7, 339-349
|Abstract »|Full Text »|PDF »
Dicer-1, but not Loquacious, is critical for assembly of miRNA-induced silencing complexes.
X. Liu, J. K. Park, F. Jiang, Y. Liu, D. McKearin, and Q. Liu (2007)
RNA
13, 2324-2329
|Abstract »|Full Text »|PDF »
DNA Branch Nuclease Activity of Vaccinia A22 Resolvase.
M. J. Culyba, N. Minkah, Y. Hwang, O.-M. J. Benhamou, and F. D. Bushman (2007)
J. Biol. Chem.
282, 34644-34652
|Abstract »|Full Text »|PDF »
ARGONAUTE4 Is Required for Resistance to Pseudomonas syringae in Arabidopsis.
Crystal structure, stability and in vitro RNAi activity of oligoribonucleotides containing the ribo-difluorotoluyl nucleotide: insights into substrate requirements by the human RISC Ago2 enzyme.
F. Li, P. S. Pallan, M. A. Maier, K. G. Rajeev, S. L. Mathieu, C. Kreutz, Y. Fan, J. Sanghvi, R. Micura, E. Rozners, et al. (2007)
Nucleic Acids Res.
35, 6424-6438
|Abstract »|Full Text »|PDF »
Reduced levels of Ago2 expression result in increased siRNA competition in mammalian cells.
T. A. Vickers, W. F. Lima, J. G. Nichols, and S. T. Crooke (2007)
Nucleic Acids Res.
35, 6598-6610
|Abstract »|Full Text »|PDF »
Jidong Liu: Probing P-bodies for the secrets of silencing.
A Slicer-independent role for Argonaute 2 in hematopoiesis and the microRNA pathway.
D. O'Carroll, I. Mecklenbrauker, P. P. Das, A. Santana, U. Koenig, A. J. Enright, E. A. Miska, and A. Tarakhovsky (2007)
Genes & Dev.
21, 1999-2004
|Abstract »|Full Text »|PDF »
Combinatorial delivery of small interfering RNAs reduces RNAi efficacy by selective incorporation into RISC.
D. Castanotto, K. Sakurai, R. Lingeman, H. Li, L. Shively, L. Aagaard, H. Soifer, A. Gatignol, A. Riggs, and J. J. Rossi (2007)
Nucleic Acids Res.
35, 5154-5164
|Abstract »|Full Text »|PDF »
A Double-Stranded-RNA Response Program Important for RNA Interference Efficiency.
S. Choudhary, H.-C. Lee, M. Maiti, Q. He, P. Cheng, Q. Liu, and Y. Liu (2007)
Mol. Cell. Biol.
27, 3995-4005
|Abstract »|Full Text »|PDF »
Structure of Aquifex aeolicus Argonaute Highlights Conformational Flexibility of the PAZ Domain as a Potential Regulator of RNA-induced Silencing Complex Function.
U. J. Rashid, D. Paterok, A. Koglin, H. Gohlke, J. Piehler, and J. C.-H. Chen (2007)
J. Biol. Chem.
282, 13824-13832
|Abstract »|Full Text »|PDF »
RNA interference based gene therapy for neurological disease.
Antiviral Protein APOBEC3G Localizes to Ribonucleoprotein Complexes Found in P Bodies and Stress Granules.
S. Gallois-Montbrun, B. Kramer, C. M. Swanson, H. Byers, S. Lynham, M. Ward, and M. H. Malim (2007)
J. Virol.
81, 2165-2178
|Abstract »|Full Text »|PDF »
QIP, a putative exonuclease, interacts with the Neurospora Argonaute protein and facilitates conversion of duplex siRNA into single strands.
X. Zhang, Y.-R. Yuan, Y. Pei, S.-S. Lin, T. Tuschl, D. J. Patel, and N.-H. Chua (2006)
Genes & Dev.
20, 3255-3268
|Abstract »|Full Text »|PDF »
Effects of Dicer and Argonaute down-regulation on mRNA levels in human HEK293 cells.
D. Schmitter, J. Filkowski, A. Sewer, R. S. Pillai, E. J. Oakeley, M. Zavolan, P. Svoboda, and W. Filipowicz (2006)
Nucleic Acids Res.
34, 4801-4815
|Abstract »|Full Text »|PDF »
Competition for RISC binding predicts in vitro potency of siRNA.
E. Koller, S. Propp, H. Murray, W. Lima, B. Bhat, T. P. Prakash, C. R. Allerson, E. E. Swayze, E. G. Marcusson, and N. M. Dean (2006)
Nucleic Acids Res.
34, 4467-4476
|Abstract »|Full Text »|PDF »
Argonaute slicing is required for heterochromatic silencing and spreading..
D. V. Irvine, M. Zaratiegui, N. H. Tolia, D. B. Goto, D. H. Chitwood, M. W. Vaughn, L. Joshua-Tor, and R. A. Martienssen (2006)
Science
313, 1134-1137
|Abstract »|Full Text »|PDF »
A genomewide screen for components of the RNAi pathway in Drosophila cultured cells.
S. Dorner, L. Lum, M. Kim, R. Paro, P. A. Beachy, and R. Green (2006)
PNAS
103, 11880-11885
|Abstract »|Full Text »|PDF »
Small RNAs just got bigger: Piwi-interacting RNAs (piRNAs) in mammalian testes.
MicroRNA-Targeted and Small Interfering RNA-Mediated mRNA Degradation Is Regulated by Argonaute, Dicer, and RNA-Dependent RNA Polymerase in Arabidopsis.
M. Ronemus, M. W. Vaughn, and R. A. Martienssen (2006)
PLANT CELL
18, 1559-1574
|Abstract »|Full Text »|PDF »
Functional replacement of Trypanosoma brucei Argonaute by the human slicer Argonaute2.
Genome-Wide Analysis of mRNAs Regulated by Drosha and Argonaute Proteins in Drosophila melanogaster.
J. Rehwinkel, P. Natalin, A. Stark, J. Brennecke, S. M. Cohen, and E. Izaurralde (2006)
Mol. Cell. Biol.
26, 2965-2975
|Abstract »|Full Text »|PDF »
Control of translation and mRNA degradation by miRNAs and siRNAs..
M. A. Valencia-Sanchez, J. Liu, G. J. Hannon, and R. Parker (2006)
Genes & Dev.
20, 515-524
|Abstract »|Full Text »|PDF »
Role of the silkworm argonaute2 homolog gene in double-strand break repair of extrachromosomal DNA.
H. Tsukioka, M. Takahashi, H. Mon, K. Okano, K. Mita, T. Shimada, J. M. Lee, Y. Kawaguchi, K. Koga, and T. Kusakabe (2006)
Nucleic Acids Res.
34, 1092-1101
|Abstract »|Full Text »|PDF »
HelF, a putative RNA helicase acts as a nuclear suppressor of RNAi but not antisense mediated gene silencing.
B. Popova, M. Kuhlmann, A. Hinas, F. Soderbom, and W. Nellen (2006)
Nucleic Acids Res.
34, 773-784
|Abstract »|Full Text »|PDF »
Stem-cell protein Piwil2 is widely expressed in tumors and inhibits apoptosis through activation of Stat3/Bcl-XL pathway.
J. H. Lee, D. Schutte, G. Wulf, L. Fuzesi, H.-J. Radzun, S. Schweyer, W. Engel, and K. Nayernia (2006)
Hum. Mol. Genet.
15, 201-211
|Abstract »|Full Text »|PDF »
Small interfering RNAs containing full 2'-O-methylribonucleotide-modified sense strands display Argonaute2/eIF2C2-dependent activity.
Structural and Functional Characterization of Noncoding Repetitive RNAs Transcribed in Stressed Human Cells.
R. Valgardsdottir, I. Chiodi, M. Giordano, F. Cobianchi, S. Riva, and G. Biamonti (2005)
Mol. Biol. Cell
16, 2597-2604
|Abstract »|Full Text »|PDF »
Seawi--a sea urchin piwi/argonaute family member is a component of MT-RNP complexes.
A. J. RODRIGUEZ, S. A. SEIPEL, D. R. HAMILL, D. P. ROMANCINO, M. DI CARLO, K. A. SUPRENANT, and E. M. BONDER (2005)
RNA
11, 646-656
|Abstract »|Full Text »|PDF »
Cytoplasmic and Nuclear Retained DMPK mRNAs Are Targets for RNA Interference in Myotonic Dystrophy Cells.
M.-A. Langlois, C. Boniface, G. Wang, J. Alluin, P. M. Salvaterra, J. Puymirat, J. J. Rossi, and N. S. Lee (2005)
J. Biol. Chem.
280, 16949-16954
|Abstract »|Full Text »|PDF »
RDE-2 interacts with MUT-7 to mediate RNA interference in Caenorhabditis elegans.
B. B. J. Tops, H. Tabara, T. Sijen, F. Simmer, C. C. Mello, R. H. A. Plasterk, and R. F. Ketting (2005)
Nucleic Acids Res.
33, 347-355
|Abstract »|Full Text »|PDF »
Translation of stable hepadnaviral mRNA cleavage fragments induced by the action of phosphorothioate-modified antisense oligodeoxynucleotides.
P. Hasselblatt, B. Hockenjos, C. Thoma, H. E. Blum, and W.-B. Offensperger (2005)
Nucleic Acids Res.
33, 114-125
|Abstract »|Full Text »|PDF »
From The Cover: RNA-dependent RNA polymerase is an essential component of a self-enforcing loop coupling heterochromatin assembly to siRNA production.
T. Sugiyama, H. Cam, A. Verdel, D. Moazed, and S. I. S. Grewal (2005)
PNAS
102, 152-157
|Abstract »|Full Text »|PDF »