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Loss of the Mili-interacting Tudor domain–containing protein-1 activates transposons and alters the Mili-associated small RNA profile

Nature Structural & Molecular Biology volume 16, pages 639–646 (2009)Cite this article

Abstract

Piwi proteins and their associated Piwi-interacting RNAs (piRNAs) are implicated in transposon silencing in the mouse germ line. There is currently little information on additional proteins in the murine Piwi complex and how they might regulate the entry of transcripts that accumulate as piRNAs in the Piwi ribonucleoprotein (piRNP). We isolated Mili-containing complexes from adult mouse testes and identified Tudor domain–containing protein-1 (Tdrd1) as a factor specifically associated with the Mili piRNP throughout spermatogenesis. Complex formation is promoted by the recognition of symmetrically dimethylated arginines at the N terminus of Mili by the tudor domains of Tdrd1. Similar to a Mili mutant, mice lacking Tdrd1 show derepression of L1 transposons accompanied by a loss of DNA methylation at their regulatory elements and delocalization of Miwi2 from the nucleus to the cytoplasm. Finally, we show that Mili piRNPs devoid of Tdrd1 accept the entry of abundant cellular transcripts into the piRNA pathway and accumulate piRNAs with a profile that is drastically different from that of the wild type. Our data suggest that Tdrd1 ensures the entry of correct transcripts into the normal piRNA pool.

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Figure 1: Tdrd1 is a Mili-specific interacting factor.
Figure 2: Tudor domains of Tdrd1 interact with the Mili N terminus, which carries symmetrical dimethyl modifications on RG dipeptides.
Figure 3: Tdrd1 associates with the Mili small RNAs throughout spermatogenesis.
Figure 4: Activation of L1 transposons in Tdrd1 mutant testes is coupled to loss of DNA methylation of the 5′ regulatory region of the L1 element.
Figure 5: Activation of L1 and mislocalization of Miwi2 in the fetal germ cells of Tdrd1 mutants.
Figure 6: Loss of Tdrd1 leads to overrepresentation of exonic reads in Mili.

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Acknowledgements

We thank A. Aravin and G. Hannon (Cold Spring Harbour Laboratory), J. Martinez (Institute of Molecular Biotechnology, Vienna), S. Martin (University of Colorado School of Medicine), A. Bortvin (Carnegie Institution of Washington) and D. Schuemperli and M. Ruepp (University of Bern) for reagents. We thank the European Molecular Biology Laboratory (EMBL) Mononclonal Antibody Facility, Protein Expression and Gene Core facilities for antibody production and Solexa sequencing. We are grateful to D. O'Carroll, A. Verdel and W. Filipowicz and members of the Pillai group for excellent discussions and reading of the manuscript. Research in the Pillai group is supported by EMBL and a grant from Region Rhone Alp (CIBLE 2008).

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Authors and Affiliations

  1. European Molecular Biology Laboratory, Grenoble Outstation, France

    Michael Reuter & Ramesh S Pillai

  2. Institute for Frontier Medical Sciences, Kyoto University, Japan

    Shinichiro Chuma & Takashi Tanaka

  3. European Molecular Biology Laboratory, Heidelberg, Germany

    Thomas Franz

  4. Institute of Molecular Pathology, Vienna, Austria

    Alexander Stark

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  1. Michael Reuter
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Contributions

M.R. performed most of the experiments described in this study; S.C. provided immunostaining data and Tdrd1 mutant mice; T.T. purified germ cells by FACS; T.F. provided MS analysis; A.S. performed Solexa sequence analysis; R.S.P. designed the research and wrote the paper.

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Correspondence to Ramesh S Pillai.

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Reuter, M., Chuma, S., Tanaka, T. et al. Loss of the Mili-interacting Tudor domain–containing protein-1 activates transposons and alters the Mili-associated small RNA profile. Nat Struct Mol Biol 16, 639–646 (2009). https://doi.org/10.1038/nsmb.1615

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