Skip to main content

Advertisement

SpringerLink
Log in

Human DNA Methyltransferase 3a does not Associate with MicroRNAs in the Regulation of DNA Methylation

  • Published:
Journal of Cardiovascular Translational Research Aims and scope Submit manuscript

Abstract

Despite important roles in mammalian gene regulation, the critical targeting mechanism of DNA methylation for specific DNA sequences remains unclear. Recently, small non-coding RNAs were reported to be essential for DNA methylation in plants as well as in mice, suggesting that small non-coding RNAs might interact with DNA methyltransferases (DNMTs) to provide them with target sequence information. In the present study, we attempted to detect and isolate microRNAs by immunoprecipitation (IP) that might be associated with human DNMT3a. When analyzed by gel electrophoresis after radioisotope-labeling, DNMT3a IP revealed no detectable levels of microRNA. RNA from DNMT3a IP was also analyzed with microRNA microarray and with subsequent RT-PCR. The results showed no specific enrichment of candidate microRNAs in the DNMT3a IP. DNMT3a was not directly associated with microRNAs. But, other classes of small non-coding RNA could still be implicated with DNMTs in a specific tissue such as testis where such RNA species are highly abundant.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

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

    Article  CAS  PubMed  Google Scholar 

  2. Bird, A. (2007). Perceptions of epigenetics. Nature, 447, 396–398.

    Article  CAS  PubMed  Google Scholar 

  3. Chen, C., Ridzon, D. A., Broomer, A. J., et al. (2005). Real-time quantification of microRNAs by stem-loop RT-PCR. Nucleic Acids Research, 33, e179.

    Article  PubMed  Google Scholar 

  4. Edwards, C. A., & Ferguson-Smith, A. C. (2007). Mechanisms regulating imprinted genes in clusters. Current Opinion in Cell Biology, 19, 281–289.

    Article  CAS  PubMed  Google Scholar 

  5. Esteller, M. (2007). Epigenetic gene silencing in cancer: the DNA hypermethylome. Human Molecular Genetics, 16, R50–R59.

    Article  CAS  PubMed  Google Scholar 

  6. Futscher, B. W., Oshiro, M. M., Wozniak, R. J., et al. (2002). Role for DNA methylation in the control of cell type specific maspin expression. Nature Genetics, 31, 175–179.

    Article  CAS  PubMed  Google Scholar 

  7. Girard, A., Sachidanandam, R., Hannon, G. J., & Carmell, M. A. (2006). A germline-specific class of small RNAs binds mammalian Piwi proteins. Nature, 442, 199–202.

    PubMed  Google Scholar 

  8. Heard, E., Clerc, P., & Avner, P. (1997). X-chromosome inactivation in mammals. Annual Review of Genetics, 31, 571–610.

    Article  CAS  PubMed  Google Scholar 

  9. Hutvágner, G., & Zamore, P. D. (2002). A microRNA in a multiple-turnover RNAi enzyme complex. Science, 297, 2056–2060.

    Article  PubMed  Google Scholar 

  10. Hwang, H. W., Wentzel, E. A., & Mendell, J. T. (2007). A hexanucleotide element directs microRNA nuclear import. Science, 315, 97–100.

    Article  CAS  PubMed  Google Scholar 

  11. Jeffery, L., & Nakielny, S. (2004). Components of the DNA methylation system of chromatin control are RNA-binding proteins. Journal of Biological Chemistry, 279, 49479–49487.

    Article  CAS  PubMed  Google Scholar 

  12. Kawasaki, H., & Taira, K. (2006). Retraction: induction of DNA methylation and gene silencing by short interfering RNAs in human cells. Nature, 441, 1176.

    Article  CAS  Google Scholar 

  13. Kim, G. D., Ni, J., Kelesoglu, N., Roberts, R. J., & Pradhan, S. (2002). Co-operation and communication between the human maintenance and de novo DNA (cytosine-5) methyltransferases. EMBO Journal, 21, 4183–4195.

    Article  CAS  PubMed  Google Scholar 

  14. Klose, R. J., & Bird, A. P. (2006). Genomic DNA methylation: the mark and its mediators. Trends in Biochemical Sciences, 31, 89–97.

    Article  CAS  PubMed  Google Scholar 

  15. Kuramochi-Miyagawa, S., Watanabe, T., Gotoh, K., et al. (2008). DNA methylation of retrotransposon genes is regulated by Piwi family members MILI and MIWI2 in murine fetal testes. Genes and Development, 22, 908–917.

    Article  CAS  PubMed  Google Scholar 

  16. Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 227, 680–685.

    Article  CAS  PubMed  Google Scholar 

  17. Lagos-Quintana, M., Rauhut, R., Lendeckel, W., & Tuschl, T. (2001). Identification of novel genes coding for small expressed RNAs. Science, 294, 853–858.

    Article  CAS  PubMed  Google Scholar 

  18. Lagos-Quintana, M., Rauhut, R., Meyer, J., Borkhardt, A., & Tuschl, T. (2003). New microRNAs from mouse and human. RNA, 9, 175–179.

    Article  CAS  PubMed  Google Scholar 

  19. Lee, R. C., Feinbaum, R. L., & Ambros, V. (1993). The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell, 75, 843–854.

    Article  CAS  PubMed  Google Scholar 

  20. Lee, Y., Ahn, C., Han, J., Choi, H., Kim, J., Yim, J., et al. (2003). The nuclear RNase III Drosha initiates microRNA processing. Nature, 425, 415–419.

    Article  CAS  PubMed  Google Scholar 

  21. Li, E., Bestor, T. H., & Jaenisch, R. (1992). Targeted mutation of the DNA methyltransferase gene results in embryonic lethality. Cell, 69, 915–926.

    Article  CAS  PubMed  Google Scholar 

  22. Lund, E., Guttinger, S., Calado, A., Dahlberg, J. E., & Kutay, U. (2004). Nuclear export of microRNA precursors. Science, 303, 95–98.

    Article  CAS  PubMed  Google Scholar 

  23. Margot, J. B., Cardoso, M. C., & Leonhardt, H. (2001). Mammalian DNA methyltransferases show different subnuclear distributions. Journal of Cellular Biochemistry, 83, 373–379.

    Article  CAS  PubMed  Google Scholar 

  24. Margot, J. B., Ehrenhofer-Murray, A. E., & Leonhardt, H. (2003). Interactions within the mammalian DNA methyltransferase family. BMC Molecular Biology, 4, 7.

    Article  PubMed  Google Scholar 

  25. Matzke, M., Kanno, T., Huettel, B., Daxinger, L., & Matzke, A. J. (2007). Targets of RNA-directed DNA methylation. Current Opinion in Plant Biology, 10, 512–519.

    Article  CAS  PubMed  Google Scholar 

  26. Morris, K. V., Chan, S. W., Jacobsen, S. E., & Looney, D. J. (2004). Small interfering RNA-induced transcriptional gene silencing in human cells. Science, 305, 1289–1292.

    Article  CAS  PubMed  Google Scholar 

  27. Mourelatos, Z., Dostie, J., Paushkin, S., Sharma, A., Charroux, B., Abel, L., et al. (2002). miRNPs: a novel class of ribonucleoproteins containing numerous microRNAs. Genes and Development, 16, 720–728.

    Article  CAS  PubMed  Google Scholar 

  28. Murchison, E. P., Partridge, J. F., Tam, O. H., Cheloufi, S., & Hannon, G. J. (2005). Characterization of Dicer-deficient murine embryonic stem cells. Proceedings of the National Academy of Sciences of the United States of America, 102, 12135–12140.

    Article  CAS  PubMed  Google Scholar 

  29. Okano, M., Bell, D. W., Haber, D. A., & Li, E. (1999). DNA methyltransferases Dnmt3a and Dnmt3b are essential for de novo methylation and mammalian development. Cell, 99, 247–257.

    Article  CAS  PubMed  Google Scholar 

  30. Park, C. W., Chen, Z., Kren, B. T., & Steer, C. J. (2004). Double-stranded siRNA targeted to the huntingtin gene does not induce DNA methylation. Biochemical and Biophysical Research Communications, 323, 275–280.

    Article  CAS  PubMed  Google Scholar 

  31. Sakai, T., Toguchida, J., Ohtani, N., Yandell, D. W., Rapaport, J. M., & Dryja, T. P. (1991). Allele-specific hypermethylation of the retinoblastoma tumor-suppressor gene. American Journal of Human Genetics, 48, 880–888.

    CAS  PubMed  Google Scholar 

  32. Suetake, I., Shinozaki, F., Miyagawa, J., Takeshima, H., & Tajima, S. (2004). DNMT3L stimulates the DNA methylation activity of Dnmt3a and Dnmt3b through a direct interaction. Journal of Biological Chemistry, 279, 27816–27823.

    Article  CAS  PubMed  Google Scholar 

  33. Thomson, J. M., Parker, J., Perou, C. M., & Hammond, S. M. (2004). A custom microarray platform for analysis of microRNA gene expression. Nature Methods, 1, 47–53.

    Article  CAS  PubMed  Google Scholar 

  34. Ting, A. H., Schuebel, K. E., Herman, J. G., & Baylin, S. B. (2005). Short double-stranded RNA induces transcriptional gene silencing in human cancer cells in the absence of DNA methylation. Nature Genetics, 37, 906–910.

    Article  CAS  PubMed  Google Scholar 

  35. Yekta, S., Shih, I. H., & Bartel, D. P. (2004). MicroRNA-directed cleavage of HOXB8 mRNA. Science, 304, 594–596.

    Article  CAS  PubMed  Google Scholar 

  36. Zeng, Y., & Cullen, B. R. (2003). Sequence requirements for micro RNA processing and function in human cells. RNA, 9, 112–123.

    Article  CAS  PubMed  Google Scholar 

  37. Zhu, H., Geiman, T. M., Xi, S., Jiang, Q., Schmidtmann, A., Chen, T., et al. (2006). Lsh is involved in de novo methylation of DNA. EMBO Journal, 25, 335–345.

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Medicine, University of Minnesota Medical School, Minneapolis, MN, 55455, USA

    Chang Won Park & Clifford J. Steer

  2. Department of Genetics, Cell Biology, and Development, University of Minnesota Medical School, Minneapolis, MN, 55455, USA

    Clifford J. Steer

  3. Department of Pharmacology, University of Minnesota Medical School, Minneapolis, MN, 55455, USA

    Yan Zeng

Authors
  1. Chang Won Park
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yan Zeng
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Clifford J. Steer
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Clifford J. Steer.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Park, C.W., Zeng, Y. & Steer, C.J. Human DNA Methyltransferase 3a does not Associate with MicroRNAs in the Regulation of DNA Methylation. J. of Cardiovasc. Trans. Res. 3, 290–295 (2010). https://doi.org/10.1007/s12265-010-9167-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12265-010-9167-9

Keywords

Search

Navigation