Abstract
Drosophila melanogaster has two Dicer proteins with specialized functions. Dicer-1 liberates miRNA-miRNA* duplexes from precursor miRNAs (pre-miRNAs), whereas Dicer-2 processes long double-stranded RNAs into small interfering RNA duplexes. It was recently demonstrated that Dicer-2 is rendered highly specific for long double-stranded RNA substrates by inorganic phosphate and a partner protein R2D2. However, it remains unclear how Dicer-1 exclusively recognize pre-miRNAs. Here we show that fly Dicer-1 recognizes the single-stranded terminal loop structure of pre-miRNAs through its N-terminal helicase domain, checks the loop size and measures the distance between the 3′ overhang and the terminal loop. This unique mechanism allows fly Dicer-1 to strictly inspect the authenticity of pre-miRNA structures.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$189.00 per year
only $15.75 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Bartel, D.P. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116, 281–297 (2004).
Kim, V.N., Han, J. & Siomi, M.C. Biogenesis of small RNAs in animals. Nat. Rev. Mol. Cell Biol. 10, 126–139 (2009).
Förstemann, K. et al. Normal microRNA maturation and germ-line stem cell maintenance requires Loquacious, a double-stranded RNA-binding domain protein. PLoS Biol. 3, e236 (2005).
Saito, K., Ishizuka, A., Siomi, H. & Siomi, M.C. Processing of pre-microRNAs by the Dicer-1–Loquacious complex in Drosophila cells. PLoS Biol. 3, e235 (2005).
Jiang, F. et al. Dicer-1 and R3D1-L catalyze microRNA maturation in Drosophila. Genes Dev. 19, 1674–1679 (2005).
Chendrimada, T.P. et al. TRBP recruits the Dicer complex to Ago2 for microRNA processing and gene silencing. Nature 436, 740–744 (2005).
Haase, A.D. et al. TRBP, a regulator of cellular PKR and HIV-1 virus expression, interacts with Dicer and functions in RNA silencing. EMBO Rep. 6, 961–967 (2005).
Lee, Y. et al. The role of PACT in the RNA silencing pathway. EMBO J. 25, 522–532 (2006).
Kok, K.H., Ng, M.H., Ching, Y.P. & Jin, D.Y. Human TRBP and PACT directly interact with each other and associate with dicer to facilitate the production of small interfering RNA. J. Biol. Chem. 282, 17649–17657 (2007).
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).
Okamura, K., Ishizuka, A., Siomi, H. & Siomi, M.C. Distinct roles for Argonaute proteins in small RNA-directed RNA cleavage pathways. Genes Dev. 18, 1655–1666 (2004).
Meister, G. et al. Human Argonaute2 mediates RNA cleavage targeted by miRNAs and siRNAs. Mol. Cell 15, 185–197 (2004).
Liu, J. et al. Argonaute2 is the catalytic engine of mammalian RNAi. Science 305, 1437–1441 (2004).
Iwasaki, S. et al. Hsc70/Hsp90 chaperone machinery mediates ATP-dependent RISC loading of small RNA duplexes. Mol. Cell 39, 292–299 (2010).
Iki, T. et al. In vitro assembly of plant RNA-induced silencing complexes facilitated by molecular chaperone HSP90. Mol. Cell 39, 282–291 (2010).
Miyoshi, T., Takeuchi, A., Siomi, H. & Siomi, M.C. A direct role for Hsp90 in pre-RISC formation in Drosophila. Nat. Struct. Mol. Biol. 17, 1024–1026 (2010).
Kawamata, T., Seitz, H. & Tomari, Y. Structural determinants of miRNAs for RISC loading and slicer-independent unwinding. Nat. Struct. Mol. Biol. 16, 953–960 (2009).
Yoda, M. et al. ATP-dependent human RISC assembly pathways. Nat. Struct. Mol. Biol. 17, 17–23 (2010).
Liu, Q. et al. R2D2, a bridge between the initiation and effector steps of the Drosophila RNAi pathway. Science 301, 1921–1925 (2003).
Pham, J.W., Pellino, J.L., Lee, Y.S., Carthew, R.W. & Sontheimer, E.J.A. Dicer-2-dependent 80S complex cleaves targeted mRNAs during RNAi in Drosophila. Cell 117, 83–94 (2004).
Tomari, Y. et al. RISC assembly defects in the Drosophila RNAi mutant armitage. Cell 116, 831–841 (2004).
Matranga, C., Tomari, Y., Shin, C., Bartel, D.P. & Zamore, P.D. Passenger-strand cleavage facilitates assembly of siRNA into Ago2-containing RNAi enzyme complexes. Cell 123, 607–620 (2005).
Rand, T.A., Petersen, S., Du, F. & Wang, X. Argonaute2 cleaves the anti-guide strand of siRNA during RISC activation. Cell 123, 621–629 (2005).
Leuschner, P.J., Ameres, S.L., Kueng, S. & Martinez, J. Cleavage of the siRNA passenger strand during RISC assembly in human cells. EMBO Rep. 7, 314–320 (2006).
Miyoshi, K., Tsukumo, H., Nagami, T., Siomi, H. & Siomi, M.C. Slicer function of Drosophila Argonautes and its involvement in RISC formation. Genes Dev. 19, 2837–2848 (2005).
Fire, A. et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391, 806–811 (1998).
Zhang, H., Kolb, F.A., Brondani, V., Billy, E. & Filipowicz, W. Human Dicer preferentially cleaves dsRNAs at their termini without a requirement for ATP. EMBO J. 21, 5875–5885 (2002).
Provost, P. et al. Ribonuclease activity and RNA binding of recombinant human Dicer. EMBO J. 21, 5864–5874 (2002).
Lee, Y.S. et al. Distinct roles for Drosophila Dicer-1 and Dicer-2 in the siRNA/miRNA silencing pathways. Cell 117, 69–81 (2004).
Cenik, E.S. et al. Phosphate and R2D2 Restrict the Substrate Specificity of Dicer-2, an ATP-Driven Ribonuclease. Mol. Cell 42, 172–184 (2011).
Ye, X., Paroo, Z. & Liu, Q. Functional anatomy of the Drosophila microRNA-generating enzyme. J. Biol. Chem. 282, 28373–28378 (2007).
Ma, J.B., Ye, K. & Patel, D.J. Structural basis for overhang-specific small interfering RNA recognition by the PAZ domain. Nature 429, 318–322 (2004).
Macrae, I.J. et al. Structural basis for double-stranded RNA processing by Dicer. Science 311, 195–198 (2006).
Fairman-Williams, M.E., Guenther, U.P. & Jankowsky, E. SF1 and SF2 helicases: family matters. Curr. Opin. Struct. Biol. 20, 313–324 (2010).
Jankowsky, E. RNA helicases at work: binding and rearranging. Trends Biochem. Sci. 36, 19–29 (2011).
Linder, P. Dead-box proteins: a family affair—active and passive players in RNP-remodeling. Nucleic Acids Res. 34, 4168–4180 (2006).
Hilbert, M., Karow, A.R. & Klostermeier, D. The mechanism of ATP-dependent RNA unwinding by DEAD box proteins. Biol. Chem. 390, 1237–1250 (2009).
Welker, N.C. et al. Dicer's helicase domain discriminates dsRNA termini to promote an altered reaction mode. Mol. Cell 41, 589–599 (2011).
Wang, H.W. et al. Structural insights into RNA processing by the human RISC-loading complex. Nat. Struct. Mol. Biol. 16, 1148–1153 (2009).
Lau, P.W., Potter, C.S., Carragher, B. & MacRae, I.J. Structure of the human Dicer-TRBP complex by electron microscopy. Structure 17, 1326–1332 (2009).
Nykänen, A., Haley, B. & Zamore, P.D. ATP requirements and small interfering RNA structure in the RNA interference pathway. Cell 107, 309–321 (2001).
Welker, N.C. et al. Dicer's helicase domain is required for accumulation of some, but not all, C. elegans endogenous siRNAs. RNA 16, 893–903 (2010).
Mateos, J.L., Bologna, N.G., Chorostecki, U. & Palatnik, J.F. Identification of microRNA processing determinants by random mutagenesis of Arabidopsis MIR172a precursor. Curr. Biol. 20, 49–54 (2010).
Song, L., Axtell, M.J. & Fedoroff, N.V. RNA secondary structural determinants of miRNA precursor processing in Arabidopsis. Curr. Biol. 20, 37–41 (2010).
Werner, S., Wollmann, H., Schneeberger, K. & Weigel, D. Structure determinants for accurate processing of miR172a in Arabidopsis thaliana. Curr. Biol. 20, 42–48 (2010).
Bologna, N.G., Mateos, J.L., Bresso, E.G. & Palatnik, J.F. A loop-to-base processing mechanism underlies the biogenesis of plant microRNAs miR319 and miR159. EMBO J. 28, 3646–3656 (2009).
Dong, Z., Han, M.H. & Fedoroff, N. The RNA-binding proteins HYL1 and SE promote accurate in vitro processing of pri-miRNA by DCL1. Proc. Natl. Acad. Sci. USA 105, 9970–9975 (2008).
Ma, E., MacRae, I.J., Kirsch, J.F. & Doudna, J.A. Autoinhibition of human dicer by its internal helicase domain. J. Mol. Biol. 380, 237–243 (2008).
Soifer, H.S. et al. A role for the Dicer helicase domain in the processing of thermodynamically unstable hairpin RNAs. Nucleic Acids Res. 36, 6511–6522 (2008).
Myong, S. et al. Cytosolic viral sensor RIG-I is a 5′-triphosphate-dependent translocase on double-stranded RNA. Science 323, 1070–1074 (2009).
Tomari, Y., Du, T. & Zamore, P.D. Sorting of Drosophila small silencing RNAs. Cell 130, 299–308 (2007).
Förstemann, K., Horwich, M.D., Wee, L., Tomari, Y. & Zamore, P.D. Drosophila microRNAs are sorted into functionally distinct Argonaute complexes after production by Dicer-1. Cell 130, 287–297 (2007).
Iwasaki, S., Kawamata, T. & Tomari, Y. Drosophila Argonaute1 and Argonaute2 use distinct mechanisms for translational repression. Mol. Cell 34, 58–67 (2009).
Acknowledgements
We are grateful to Q. Liu (University of Texas Southwestern Medical Center) for the pFastBac-His-Dcr-1 wild type and E1908A E2139A plasmids, to M. Siomi and H. Siomi (Keio University) for the plasmid for expressing Loqs-PB, and to S. Katsuma for his advice on protein expression. We thank P. Bas Kwak for assistance with the amino acid sequence alignment, and we thank K. Förstemann and members of the Tomari laboratory for discussions, suggestions and critical comments on the manuscript. This work was supported in part by a Grant-in-Aid for Scientific Research on Innovative Areas ('Functional machinery for noncoding RNAs') and a a Grant-in-Aid for Young Scientists from the Japan Ministry of Education, Culture, Sports, Science and Technology to Y.T.; and a Carrier Development Award from The International Human Frontier Science Program Organization to Y.T. N.I. is a recipient of a Research Fellowship from the Japan Society for the Promotion of Science.
Author information
Authors and Affiliations
Contributions
A.T. conducted biochemical experiments with the assistance of T.K. and N.I., H.S. conducted bioinformatic analyses, Y.T. supervised the study; A.T. and Y.T. wrote the manuscript, and all authors discussed the results and approved the manuscript.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Supplementary information
Supplementary Text and Figures
Supplementary Figures 1–7 and Supplementary Table 1 (PDF 1651 kb)
Rights and permissions
About this article
Cite this article
Tsutsumi, A., Kawamata, T., Izumi, N. et al. Recognition of the pre-miRNA structure by Drosophila Dicer-1. Nat Struct Mol Biol 18, 1153–1158 (2011). https://doi.org/10.1038/nsmb.2125
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nsmb.2125
This article is cited by
-
Regulation of microRNA expression by the adaptor protein GRB2
Scientific Reports (2023)
-
Sequence determinant of small RNA production by DICER
Nature (2023)
-
Secondary structure RNA elements control the cleavage activity of DICER
Nature Communications (2022)
-
DNA 5-hydroxymethylcytosine in pediatric central nervous system tumors may impact tumor classification and is a positive prognostic marker
Clinical Epigenetics (2021)
-
Visualizing a protonated RNA state that modulates microRNA-21 maturation
Nature Chemical Biology (2021)