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Arginine methylation of Piwi proteins catalysed by dPRMT5 is required for Ago3 and Aub stability

Nature Cell Biology volume 11, pages 652–658 (2009)Cite this article

Abstract

Piwi family proteins are essential for germline development and bind piwi-interacting RNAs (piRNAs)1,2,3. The grandchildless gene aub of Drosophila melanogaster encodes the piRNA-binding protein Aubergine (Aub), which is essential for formation of primordial germ cells (PGCs)4. Here we report that Piwi family proteins of mouse, Xenopus laevis and Drosophila contain symmetrical dimethylarginines (sDMAs). We found that Piwi proteins are expressed in Xenopus oocytes and we identified numerous Xenopus piRNAs. We report that the Drosophila homologue of protein methyltransferase 5 (dPRMT5, csul/dart5), which is also the product of a grandchildless gene5,6, is required for arginine methylation of Drosophila Piwi, Ago3 and Aub proteins in vivo. Loss of dPRMT5 activity led to a reduction in the levels of piRNAs, Ago3 and Aub proteins, and accumulation of retrotransposons in the Drosophila ovary. Our studies explain the relationship between aub and dPRMT5 (csul/dart5) genes by demonstrating that dPRMT5 is the enzyme that methylates Aub. Our findings underscore the significance of sDMA modification of Piwi proteins in the germline and suggest an interacting pathway of genes that are required for piRNA function and PGC specification.

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Figure 1: Miwi and Mili proteins contain sDMAs.
Figure 2: Xenopus Piwi proteins with bound piRNAs are immunoprecipitated by Y12 and contain sDMAs.
Figure 3: Drosophila PRMT5 (csul, dart5) is required for arginine methylation of Aub, Piwi and Ago3 proteins in ovaries.
Figure 4: Reduction of Piwi proteins and piRNAs with accumulation of HeT-A retrotransposons and marked reduction of the Aub protein that localizes in the pole plasm, in the absence of dPRMT5 (csul) activity.
Figure 5: Proposed classification for selected Drosophila grandchildless genes.

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References

  1. Kim, V. N. Small RNAs just got bigger: Piwi-interacting RNAs (piRNAs) in mammalian testes. Genes Dev. 20, 1993–1997 (2006).

    Article  CAS  Google Scholar 

  2. O'Donnell K, A. & Boeke, J. D. Mighty Piwis Defend the Germline against Genome Intruders. Cell 129, 37–44 (2007).

    Article  CAS  Google Scholar 

  3. Hartig, J. V., Tomari, Y. & Forstemann, K. piRNAs--the ancient hunters of genome invaders. Genes Dev. 21, 1707–1713 (2007).

    Article  CAS  Google Scholar 

  4. Harris, A. N. & Macdonald, P. M. Aubergine encodes a Drosophila polar granule component required for pole cell formation and related to eIF2C. Development 128, 2823–32 (2001).

    CAS  PubMed  Google Scholar 

  5. Anne, J., Ollo, R., Ephrussi, A. & Mechler, B. M. Arginine methyltransferase Capsuleen is essential for methylation of spliceosomal Sm proteins and germ cell formation in Drosophila. Development 134, 137–146 (2007).

    Article  CAS  Google Scholar 

  6. Gonsalvez, G. B., Rajendra, T. K., Tian, L. & Matera, A. G. The Sm-protein methyltransferase, dart5, is essential for germ-cell specification and maintenance. Curr. Biol. 16, 1077–1089 (2006).

    Article  CAS  Google Scholar 

  7. Cox, D. N. et al. A novel class of evolutionarily conserved genes defined by piwi are essential for stem cell self-renewal. Genes Dev. 12, 3715–3727 (1998).

    Article  CAS  Google Scholar 

  8. Gunawardane, L. S. et al. A slicer-mediated mechanism for repeat-associated siRNA 5′ end formation in Drosophila. Science 315, 1587–1590 (2007).

    Article  CAS  Google Scholar 

  9. Brennecke, J. et al. Discrete small RNA-generating loci as master regulators of transposon activity in Drosophila. Cell 128, 1089–1103 (2007).

    Article  CAS  Google Scholar 

  10. Girard, A. & Hannon, G. J. Conserved themes in small-RNA-mediated transposon control. Trends Cell Biol. 18, 136–148 (2008).

    Article  CAS  Google Scholar 

  11. Aravin, A. A. et al. The small RNA profile during Drosophila melanogaster development. Dev. Cell 5, 337–350 (2003).

    Article  CAS  Google Scholar 

  12. Sarot, E., Payen-Groschene, G., Bucheton, A. & Pelisson, A. Evidence for a piwi-dependent RNA silencing of the gypsy endogenous retrovirus by the Drosophila melanogaster flamenco gene. Genetics 166, 1313–1321 (2004).

    Article  CAS  Google Scholar 

  13. Siomi, M. C., Saito, K. & Siomi, H. How selfish retrotransposons are silenced in Drosophila germline and somatic cells. FEBS Lett. 582, 2473–2478 (2008).

    Article  CAS  Google Scholar 

  14. Aravin, A. A., Sachidanandam, R., Girard, A., Fejes-Toth, K. & Hannon, G. J. Developmentally regulated piRNA clusters implicate MILI in transposon control. Science 316, 744–747 (2007).

    Article  CAS  Google Scholar 

  15. Houwing, S. et al. A Role for Piwi and piRNAs in germ cell maintenance and transposon silencing in zebrafish. Cell 129, 69–82 (2007).

    Article  CAS  Google Scholar 

  16. Krause, C. D. et al. Protein arginine methyltransferases: evolution and assessment of their pharmacological and therapeutic potential. Pharmacol. Ther. 113, 50–87 (2007).

    Article  CAS  Google Scholar 

  17. Bedford, M. T. & Richard, S. Arginine methylation an emerging regulator of protein function. Mol. Cell 18, 263–272 (2005).

    Article  CAS  Google Scholar 

  18. Friesen, W. J. et al. The methylosome, a 20S complex containing JBP1 and pICln, produces dimethylarginine-modified Sm proteins. Mol. Cell Biol. 21, 8289–300 (2001).

    Article  CAS  Google Scholar 

  19. Meister, G. et al. Methylation of Sm proteins by a complex containing PRMT5 and the putative U snRNP assembly factor pICln. Curr. Biol. 11, 1990–1994 (2001).

    Article  CAS  Google Scholar 

  20. Lerner, E. A., Lerner, M. R., Janeway, C. A., Jr & Steitz, J. A. Monoclonal antibodies to nucleic acid-containing cellular constituents: probes for molecular biology and autoimmune disease. Proc. Natl Acad. Sci. USA 78, 2737–2741 (1981).

    Article  CAS  Google Scholar 

  21. Brahms, H. et al. The C-terminal RG dipeptide repeats of the spliceosomal Sm proteins D1 and D3 contain symmetrical dimethylarginines, which form a major B-cell epitope for anti-Sm autoantibodies. J. Biol. Chem. 275, 17122–17129 (2000).

    Article  CAS  Google Scholar 

  22. Boisvert, F. M., Cote, J., Boulanger, M. C. & Richard, S. A proteomic analysis of arginine-methylated protein complexes. Mol. Cell Proteomics 2, 1319–1330 (2003).

    Article  CAS  Google Scholar 

  23. Bowes, J. B. et al. Xenbase: a Xenopus biology and genomics resource. Nucleic Acids Res. 36, 761–767 (2008).

    Article  Google Scholar 

  24. Kirino, Y. & Mourelatos, Z. Mouse Piwi-interacting RNAs are 2′-O-methylated at their 3′ termini. Nature Struct. Mol. Biol. 14, 347–348 (2007).

    Article  CAS  Google Scholar 

  25. Ohara, T. et al. The 3′ termini of mouse Piwi-interacting RNAs are 2′-O-methylated. Nature Struct. Mol. Biol. 14, 349–350 (2007).

    Article  CAS  Google Scholar 

  26. Saito, K. et al. Pimet, the Drosophila homolog of HEN1, mediates 2′-O-methylation of Piwi- interacting RNAs at their 3′ ends. Genes Dev. 21, 1603–1608 (2007).

    Article  CAS  Google Scholar 

  27. Horwich, M. D. et al. The Drosophila RNA methyltransferase, DmHen1, modifies germline piRNAs and single-stranded siRNAs in RISC. Curr. Biol. 17, 1265–1272 (2007).

    Article  CAS  Google Scholar 

  28. Gonsalvez, G. B. et al. Two distinct arginine methyltransferases are required for biogenesis of Sm-class ribonucleoproteins. J. Cell Biol. 178, 733–740 (2007).

    Article  CAS  Google Scholar 

  29. Gonsalvez, G. B., Praveen, K., Hicks, A. J., Tian, L. & Matera, A. G. Sm protein methylation is dispensable for snRNP assembly in Drosophila melanogaster. RNA 14, 878–887 (2008).

    Article  CAS  Google Scholar 

  30. Vagin, V. V. et al. A distinct small RNA pathway silences selfish genetic elements in the germline. Science 313, 320–324 (2006).

    Article  CAS  Google Scholar 

  31. Cox, D. N., Chao, A. & Lin, H. piwi encodes a nucleoplasmic factor whose activity modulates the number and division rate of germline stem cells. Development 127, 503–514 (2000).

    CAS  PubMed  Google Scholar 

  32. Strome, S. & Lehmann, R. Germ versus soma decisions: lessons from flies and worms. Science 316, 392–393 (2007).

    Article  CAS  Google Scholar 

  33. Anne, J. & Mechler, B. M. Valois, a component of the nuage and pole plasm, is involved in assembly of these structures, and binds to Tudor and the methyltransferase Capsuleen. Development 132, 2167–2177 (2005).

    Article  CAS  Google Scholar 

  34. Boswell, R. E. & Mahowald, A. P. tudor, a gene required for assembly of the germ plasm in Drosophila melanogaster. Cell 43, 97–104 (1985).

    Article  CAS  Google Scholar 

  35. Arkov, A. L., Wang, J. Y., Ramos, A. & Lehmann, R. The role of Tudor domains in germline development and polar granule architecture. Development 133, 4053–4062 (2006).

    Article  CAS  Google Scholar 

  36. Selenko, P. et al. SMN tudor domain structure and its interaction with the Sm proteins. Nature Struct. Biol. 8, 27–31 (2001).

    Article  CAS  Google Scholar 

  37. Cote, J. & Richard, S. Tudor domains bind symmetrical dimethylated arginines. J. Biol. Chem. 280, 28476–28483 (2005).

    Article  CAS  Google Scholar 

  38. Chuma, S. et al. Tdrd1/Mtr-1, a tudor-related gene, is essential for male germ-cell differentiation and nuage/germinal granule formation in mice. Proc. Natl Acad. Sci. USA 103, 15894–15899 (2006).

    Article  CAS  Google Scholar 

  39. Savitsky, M., Kwon, D., Georgiev, P., Kalmykova, A. & Gvozdev, V. Telomere elongation is under the control of the RNAi-based mechanism in the Drosophila germline. Genes Dev. 20, 345–354 (2006).

    Article  CAS  Google Scholar 

  40. Lim, A. K. & Kai, T. Unique germ-line organelle, nuage, functions to repress selfish genetic elements in Drosophila melanogaster. Proc. Natl Acad. Sci. USA 104, 6714–6719 (2007).

    Article  CAS  Google Scholar 

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Acknowledgements

We are grateful to M.C. Siomi, H. Siomi, K. Saito, G. Dreyfuss for antibodies; to J. Anne for csul flies; to S. Kuramochi-Miyagawa and T. Nakano for Miwi and Mili cDNA constructs; to Rebecca Beerman for help with fly methodology; to Y. Kawamura for immunofluorescence protocols and to members of the Mourelatos lab for discussions. Mass spectrometry was performed at the Proteomics Core Facility (University of Pennsylvania) and at the Proteomics Resource of the Keck Foundation (Yale University). Illumina sequencing was performed at the Functional Genomics Core of the University of Pennsylvania. Protein production was at the Protein Expression Core of the Wistar Institute. We apologize to colleagues whose studies were not cited because of space limitations. This work was supported by a Human Frontier Science Program Long Term Fellowship to Y.K., and NIH grants to T.A.J. (NS046573), P.S.K. (GM76621) and Z.M.(GM0720777, NS056070, UL1RR024134). Z.M. also received ITMAT-PENN and URF-PENN.

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Authors and Affiliations

  1. Department of Pathology and Laboratory Medicine, Division of Neuropathology, University of Pennsylvania School of Medicine, Philadelphia, 19104, Pennsylvania, USA

    Yohei Kirino, Namwoo Kim, Mariàngels de Planell-Saguer, Eugene Khandros & Zissimos Mourelatos

  2. Department of Genetics, University of Pennsylvania School of Medicine, Philadelphia, 19104, Pennsylvania, USA

    Stephanie Chiorean & Thomas A. Jongens

  3. Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, 19104, Pennsylvania, USA

    Peter S. Klein

  4. Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, 19104, Pennsylvania, USA

    Peter S. Klein

  5. Bioinformatics and Pattern Discovery Group, IBM TJ Watson Research Center, Yorktown Heights, 10598, NY, USA

    Isidore Rigoutsos

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Contributions

Y.K. and Z.M. conceived and designed the experiments; Y.K., N.K., M.P-S, E.K., S.C., P.S.K. and I.R. performed the experiments and analysis and generated all the figures; T.A.J. guided the Drosophila methodology and experiments; I.R. performed the bioinformatics analysis; T.A.J. and I.R. provided substantial input into the writing of the manuscript; Y.K. and Z.M. wrote the manuscript.

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Correspondence to Zissimos Mourelatos.

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Kirino, Y., Kim, N., de Planell-Saguer, M. et al. Arginine methylation of Piwi proteins catalysed by dPRMT5 is required for Ago3 and Aub stability. Nat Cell Biol 11, 652–658 (2009). https://doi.org/10.1038/ncb1872

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