Short RNAs mediate gene silencing, a process associated with virus resistance, developmental control and heterochromatin formation in eukaryotes1,2,3,4,5. RNA silencing is initiated through Dicer-mediated processing of double-stranded RNA into small interfering RNA (siRNA)6,7. The siRNA guide strand associates with the Argonaute protein in silencing effector complexes, recognizes complementary sequences and targets them for silencing8,9,10,11. The PAZ domain is an RNA-binding module found in Argonaute and some Dicer proteins and its structure has been determined in the free state12,13,14. Here, we report the 2.6 Å crystal structure of the PAZ domain from human Argonaute eIF2c1 bound to both ends of a 9-mer siRNA-like duplex. In a sequence-independent manner, PAZ anchors the 2-nucleotide 3′ overhang of the siRNA-like duplex within a highly conserved binding pocket, and secures the duplex by binding the 7-nucleotide phosphodiester backbone of the overhang-containing strand and capping the 5′-terminal residue of the complementary strand. On the basis of the structure and on binding assays, we propose that PAZ might serve as an siRNA-end-binding module for siRNA transfer in the RNA silencing pathway, and as an anchoring site for the 3′ end of guide RNA within silencing effector complexes.
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Denli, A. M. & Hannon, G. J. RNAi: an ever-growing puzzle. Trends Biochem. Sci. 28, 196–201 (2003)
Bartel, D. P. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116, 281–297 (2004)
Voinnet, O. RNA silencing as a plant immune system against viruses. Trends Genet. 17, 449–459 (2001)
Volpe, T. A. et al. Regulation of heterochromatic silencing and histone H3 lysine-9 methylation by RNAi. Science 297, 1833–1837 (2002)
Hall, I. M. et al. Establishment and maintenance of a heterochromatin domain. Science 297, 2232–2237 (2002)
Bernstein, E., Caudy, A. A., Hammond, S. M. & Hannon, G. J. Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature 409, 363–366 (2001)
Elbashir, S. M., Lendeckel, W. & Tuschl, T. RNA interference is mediated by 21- and 22-nucleotide RNAs. Genes Dev. 15, 188–200 (2001)
Hammond, S. M., Bernstein, E., Beach, D. & Hannon, G. J. An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature 404, 293–296 (2000)
Hammond, S. M., Boettcher, S., Caudy, A. A., Kobayashi, R. & Hannon, G. J. Argonaute2, a link between genetic and biochemical analyses of RNAi. Science 293, 1146–1150 (2001)
Nykanen, A., Haley, B. & Zamore, P. D. ATP requirements and small interfering RNA structure in the RNA interference pathway. Cell 107, 309–321 (2001)
Martinez, J., Patkaniowska, A., Urlaub, H., Luhrmann, R. & Tuschl, T. Single-stranded antisense siRNAs guide target RNA cleavage in RNAi. Cell 110, 563–574 (2002)
Yan, K. S. et al. Structure and conserved RNA binding of the PAZ domain. Nature 426, 468–474 (2003)
Lingel, A., Simon, B., Izaurralde, E. & Sattler, M. Structure and nucleic-acid binding of the Drosophila Argonaute 2 PAZ domain. Nature 426, 465–469 (2003)
Song, J. J. et al. The crystal structure of the Argonaute2 PAZ domain reveals an RNA binding motif in RNAi effector complexes. Nature Struct. Biol. 10, 1026–1032 (2003)
Elbashir, S. M., Martinez, J., Patkaniowska, A., Lendeckel, W. & Tuschl, T. Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate. EMBO J. 20, 6877–6888 (2001)
Fire, A. et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391, 806–811 (1998)
Chiu, Y. L. & Rana, T. M. siRNA function in RNAi: a chemical modification analysis. RNA 9, 1034–1048 (2003)
Theobald, D. L., Mitton-Fry, R. M. & Wuttke, D. S. Nucleic acid recognition by OB-fold proteins. Annu. Rev. Biophys. Biomol. Struct. 32, 115–133 (2003)
Liu, Q. et al. R2D2, a bridge between the initiation and effector steps of the Drosophila RNAi pathway. Science 301, 1921–1925 (2003)
Hohjoh, H. RNA interference (RNAi) induction with various types of synthetic oligonucleotide duplexes in cultured human cells. FEBS Lett. 521, 195–199 (2002)
Harborth, J. et al. Sequence, chemical, and structural variation of small interfering RNAs and short hairpin RNAs and the effect on mammalian gene silencing. Antisense Nucleic Acid Drug Dev. 13, 83–105 (2003)
Chiu, Y. L. & Rana, T. M. RNAi in human cells: basic structural and functional features of small interfering RNA. Mol. Cell 10, 549–561 (2002)
Holen, T., Amarzguioui, M., Wiiger, M. T., Babaie, E. & Prydz, H. Positional effects of short interfering RNAs targeting the human coagulation trigger Tissue Factor. Nucleic Acids Res. 30, 1757–1766 (2002)
Amarzguioui, M., Holen, T., Babaie, E. & Prydz, H. Tolerance for mutations and chemical modifications in a siRNA. Nucleic Acids Res. 31, 589–595 (2003)
Czauderna, F. et al. Structural variations and stabilising modifications of synthetic siRNAs in mammalian cells. Nucleic Acids Res. 31, 2705–2716 (2003)
Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997)
Brunger, A. T. et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998)
Jones, T. & Kjeldgaard, M. Electron-density map interpretation. Methods Enzymol. 227, 174–208 (1997)
Nicholls, A., Sharp, K. A. & Honig, B. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins 11, 281–296 (1991)
Katsamba, P. S., Park, S. & Laird-Offringa, I. A. Kinetic studies of RNA-protein interactions using surface plasmon resonance. Methods 26, 95–104 (2002)
We thank K. Saigo for providing us with the eIF2C1 complementary DNA clone. This research was supported by the NIH. We thank Y. Cheng and personnel at the Advanced Photon Source (APS) beamlines 19BM and 14IDB for help in collecting the X-ray diffraction data. Use of the APS beamline was supported by the US Department of Energy, Basic Energy Sciences, Office of Science.
The authors declare that they have no competing financial interests.
Crystallographic statistics (DOC 30 kb)
Elution profiles of PAZ and PAZ-RNA complex. (JPG 26 kb)
Stereo view of structural alignment of PAZ domains (JPG 62 kb)
Sequence alignment of PAZ domains (JPG 172 kb)
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Ma, JB., Ye, K. & Patel, D. Structural basis for overhang-specific small interfering RNA recognition by the PAZ domain. Nature 429, 318–322 (2004). https://doi.org/10.1038/nature02519
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