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Miwi catalysis is required for piRNA amplification-independent LINE1 transposon silencing

Nature volume 480, pages 264–267 (2011)Cite this article

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Abstract

Repetitive-element-derived Piwi-interacting RNAs (piRNAs)1,2 act together with Piwi proteins Mili (also known as Piwil2) and Miwi2 (also known as Piwil4) in a genome defence mechanism that initiates transposon silencing via DNA methylation in the mouse male embryonic germ line. This silencing depends on the participation of the Piwi proteins in a slicer-dependent piRNA amplification pathway and is essential for male fertility3,4. A third Piwi family member, Miwi (also known as Piwil1), is expressed in specific postnatal germ cells and associates with a unique set of piRNAs of unknown function5,6,7. Here we show that Miwi is a small RNA-guided RNase (slicer) that requires extensive complementarity for target cleavage in vitro. Disruption of its catalytic activity in mice by a single point mutation causes male infertility, and mutant germ cells show increased accumulation of LINE1 retrotransposon transcripts. We provide evidence for Miwi slicer activity directly cleaving transposon messenger RNAs, offering an explanation for the continued maintenance of repeat-derived piRNAs long after transposon silencing is established in germline stem cells. Furthermore, our study supports a slicer-dependent silencing mechanism that functions without piRNA amplification. Thus, Piwi proteins seem to act in a two-pronged mammalian transposon silencing strategy: one promotes transcriptional repression in the embryo, the other reinforces silencing at the post-transcriptional level after birth.

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Figure 1: Miwi is a slicer requiring extensive complementarity for target cleavage.
Figure 2: Miwi catalytic activity is essential for spermatogenesis.
Figure 3: Primary piRNA biogenesis is globally unaffected in the Miwi ADH mutants and mRNAs are not targeted by piRNAs.
Figure 4: Miwi piRNAs target L1 transposons.

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Gene Expression Omnibus

Data deposits

Deep sequencing and microarray data related to this study are deposited with the Gene Expression Omnibus under accession GSE32183.

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Acknowledgements

We thank S. Cusack for initial support for the project and discussions. We thank D. O’Carroll and S. Khochbin for help with mouse experiments; J. Xiol and N. Nakatsuji for helpful discussions; D. O'Carroll, S. Martin, S. Kistler and R. Balhorn for antibodies. We thank the Mouse Biology Unit, European Molecular Biology Laboratory (EMBL) Monterotondo and EMBL facilities (Gene, Proteomics, Protein Expression and Purification). This work was supported by a Deutsche Forschungsgemeinschaft (DFG) postdoctoral fellowship to M.R. and grants to R.S.P. from Agence National de la Recherche (ANR) (piRmachines) and the European Union (European Research Council, ERC Starting Grant, pisilence). The R.S.P. laboratory is supported by the EMBL.

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Authors and Affiliations

  1. European Molecular Biology Laboratory, 6 Rue Jules Horowitz, BP 181, 38042 Grenoble, France,

    Michael Reuter, Philipp Berninger & Ramesh S. Pillai

  2. CNRS-UJF-EMBL International Unit (UMI 3265) for Virus Host Cell Interactions (UVHCI), 38042 Grenoble, France,

    Michael Reuter, Philipp Berninger & Ramesh S. Pillai

  3. Institute for Frontier Medical Sciences, Kyoto University, Kyoto 606-8507, Japan,

    Shinichiro Chuma & Mihoko Hosokawa

  4. Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University, Kyoto 606-8501, Japan,

    Shinichiro Chuma & Mihoko Hosokawa

  5. Department of Genetics and Genomic Sciences, Mount Sinai School of Medicine, New York, 10029, New York, USA

    Hardik Shah & Ravi Sachidanandam

  6. European Molecular Biology Laboratory, EMBL Meyerhof Str. 1, 69117 Heidelberg, Germany,

    Charlotta Funaya & Claude Antony

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Contributions

M.R. performed all biochemical and deep sequencing experiments. S.C., M.R. and M.H. performed imaging. C.F. and C.A. performed electron microscopy analysis. P.B., R.S., R.S.P. and H.S performed bioinformatics analysis. R.S.P., M.R., S.C. and R.S. prepared the manuscript. M.R. and R.S.P. designed research.

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Correspondence to Ramesh S. Pillai.

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Reuter, M., Berninger, P., Chuma, S. et al. Miwi catalysis is required for piRNA amplification-independent LINE1 transposon silencing. Nature 480, 264–267 (2011). https://doi.org/10.1038/nature10672

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