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Target mimicry provides a new mechanism for regulation of microRNA activity

Abstract

MicroRNAs (miRNA) regulate key aspects of development and physiology in animals and plants. These regulatory RNAs act as guides of effector complexes to recognize specific mRNA sequences based on sequence complementarity, resulting in translational repression or site-specific cleavage1,2. In plants, most miRNA targets are cleaved and show almost perfect complementarity with the miRNAs around the cleavage site3,4,5,6,7,8. Here, we examined the non–protein coding gene IPS1 (INDUCED BY PHOSPHATE STARVATION1) from Arabidopsis thaliana. IPS1 contains a motif with sequence complementarity to the phosphate (Pi) starvation–induced miRNA miR-399, but the pairing is interrupted by a mismatched loop at the expected miRNA cleavage site. We show that IPS1 RNA is not cleaved but instead sequesters miR-399. Thus, IPS1 overexpression results in increased accumulation of the miR-399 target PHO2 mRNA and, concomitantly, in reduced shoot Pi content5,6,7,8. Engineering of IPS1 to be cleavable abolishes its inhibitory activity on miR-399. We coin the term 'target mimicry' to define this mechanism of inhibition of miRNA activity. Target mimicry can be generalized beyond the control of Pi homeostasis, as demonstrated using artificial target mimics.

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Figure 1: IPS1 RNA resistance to miR-399–guided cleavage.
Figure 2: Effect of IPS1 on miR-399–dependent PHO2 accumulation requires base-pairing between IPS1 and miR-399.
Figure 3: Efficient IPS1 inhibition of miR-399 requires IPS1 RNA resistance to miR-399–guided cleavage.
Figure 4: Artificial target mimics.

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Acknowledgements

We thank C. Aragoncillo, C. Castresana, M. Crespi, C. Martin, S. Prat, R. Solano and H. Vaucheret for reading the manuscript and C. Mark for editorial assistance. Technical assistance by M.J. Benito is acknowledged. We also thank the European Arabidopsis Stock Centre (NASC) stock centre for providing plant material. A.V. is supported by a fellowship from CSIC–Fondo Social Europeo (FSE), and I.M. and I.R.-S. by fellowships from the Spanish Ministry of Science and Education (MEC). Primary support for this work was provided by grants from the European Union, Comunidad de Madrid and MEC. Additional support came from the Marie Curie Research Training Network SY-STEM (M.T.) and the Max Planck Society (D.W.).

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J.M.F.-Z. was responsible for experiments in Figure 1 and Supplementary Figures 1, 2, 3, 4 and 5 and prepared constructs and performed RNA blot analysis corresponding to Figures 2, 3. A.V. performed transient expression assays in Figures 2, 3 and Supplementary Figure 5. M.I.P. and I.M. prepared constructs and performed Pi measurements shown in Supplementary Figure 3. M.T. and I.R.-S. performed the artificial mimicry experiments shown in Figure 4. D.W. supervised the artificial mimicry experiments in Figure 4. J.A.G. supervised the transient expression assays in Figures 2,3 and Supplementary Figure 5 and contributed to discussion of other experiments. J.P.-A. supervised this study and wrote the manuscript. A.L. contributed to supervision and general discussion. J.A.G., D.W., J.M.F.-Z. and all other authors commented on the manuscript.

Corresponding author

Correspondence to Javier Paz-Ares.

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The authors declare no competing financial interests.

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Supplementary Table 1, Supplementary Figures 1–5, Supplementary Methods (PDF 941 kb)

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Franco-Zorrilla, J., Valli, A., Todesco, M. et al. Target mimicry provides a new mechanism for regulation of microRNA activity. Nat Genet 39, 1033–1037 (2007). https://doi.org/10.1038/ng2079

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