Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Epigenetic silencers and Notch collaborate to promote malignant tumours by Rb silencing

Abstract

Cancer is both a genetic and an epigenetic disease. Inactivation of tumour-suppressor genes by epigenetic changes is frequently observed in human cancers, particularly as a result of the modifications of histones and DNA methylation. It is therefore important to understand how these damaging changes might come about. By studying tumorigenesis in the Drosophila eye, here we identify two Polycomb group epigenetic silencers, Pipsqueak and Lola, that participate in this process. When coupled with overexpression of Delta, deregulation of the expression of Pipsqueak and Lola induces the formation of metastatic tumours. This phenotype depends on the histone-modifying enzymes Rpd3 (a histone deacetylase), Su(var)3-9 and E(z), as well as on the chromodomain protein Polycomb. Expression of the gene Retinoblastoma-family protein (Rbf ) is downregulated in these tumours and, indeed, this downregulation is associated with DNA hypermethylation. Together, these results establish a mechanism that links the Notch–Delta pathway, epigenetic silencing pathways and cell-cycle control in the process of tumorigenesis.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Isolation of tumour-initiating genes in Drosophila melanogaster.
Figure 2: eyeful collaborates with Delta to cause malignant tumours.
Figure 3: eyeful is a complex mutation affecting two BTB family genes, lola and psq.
Figure 4: Aberrant histone modifications associated with the tumours.
Figure 5: Silencing of Rbf is associated with promoter DNA hypermethylation.

References

  1. Logan, C. Y. & Nusse, R. The Wnt signalling pathway in development and disease. Annu. Rev. Cell. Dev. Biol. 20, 781–810 (2004)

    Article  CAS  Google Scholar 

  2. Hooper, J. E. & Scott, M. P. Communicating with Hedgehogs. Nature Rev. Mol. Cell Biol. 6, 306–317 (2005)

    Article  CAS  Google Scholar 

  3. Artavanis-Tsakonas, S., Rand, M. D. & Lake, R. J. Notch signalling: cell fate control and signal integration in development. Science 284, 770–776 (1999)

    Article  ADS  CAS  Google Scholar 

  4. Hanahan, D. & Weinberg, R. A. The hallmarks of cancer. Cell 100, 57–70 (2000)

    Article  CAS  Google Scholar 

  5. Taipale, J. & Beachy, P. A. The Hedgehog and Wnt signalling pathways in cancer. Nature 411, 349–354 (2001)

    Article  ADS  CAS  Google Scholar 

  6. Allenspach, E. J., Maillard, I., Aster, J. C. & Pear, W. S. Notch signalling in cancer. Cancer Biol. Ther. 1, 466–476 (2002)

    Article  Google Scholar 

  7. Lund, A. H. & van Lohuizen, M. Epigenetics and cancer. Genes Dev. 18, 2315–2335 (2004)

    Article  CAS  Google Scholar 

  8. Valk-Lingbeek, M. E., Bruggeman, S. W. & van Lohuizen, M. Stem cells and cancer; the polycomb connection. Cell 118, 409–418 (2004)

    Article  CAS  Google Scholar 

  9. Ringrose, L. & Paro, R. Epigenetic regulation of cellular memory by the Polycomb and Trithorax group proteins. Annu. Rev. Genet. 38, 413–443 (2004)

    Article  CAS  Google Scholar 

  10. Cao, R. & Zhang, Y. The functions of E(Z)/EZH2-mediated methylation of lysine 27 in histone H3. Curr. Opin. Genet. Dev. 14, 155–164 (2004)

    Article  CAS  Google Scholar 

  11. Nguyen, C. T. et al. Histone H3-lysine 9 methylation is associated with aberrant gene silencing in cancer cells and is rapidly reversed by 5-aza-2′-deoxycytidine. Cancer Res. 62, 6456–6461 (2002)

    CAS  PubMed  Google Scholar 

  12. Lachner, M., O'Sullivan, R. J. & Jenuwein, T. An epigenetic road map for histone lysine methylation. J. Cell Sci. 116, 2117–2124 (2003)

    Article  CAS  Google Scholar 

  13. Feinberg, A. P. & Tycko, B. The history of cancer epigenetics. Nature Rev. Cancer 4, 143–153 (2004)

    Article  CAS  Google Scholar 

  14. Weinberg, R. A. The retinoblastoma protein and cell cycle control. Cell 81, 323–330 (1995)

    Article  CAS  Google Scholar 

  15. Woodhouse, E. C. & Liotta, L. A. Drosophila invasive tumors: a model for understanding metastasis. Cell Cycle 3, 38–40 (2004)

    Article  CAS  Google Scholar 

  16. Dominguez, M. & Casares, F. Organ specification-growth control connection: new insights from the Drosophila eye-antennal disc. Dev. Dyn. 232, 673–684 (2005)

    Article  CAS  Google Scholar 

  17. Toba, G. et al. The gene search system. A method for efficient detection and rapid molecular identification of genes in Drosophila melanogaster. Genetics 151, 725–737 (1999)

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Dominguez, M. & de Celis, J. F. A dorsal/ventral boundary established by Notch controls growth and polarity in the Drosophila eye. Nature 396, 276–278 (1998)

    Article  ADS  CAS  Google Scholar 

  19. Giniger, E., Tietje, K., Jan, L. Y. & Jan, Y. N. lola encodes a putative transcription factor required for axon growth and guidance in Drosophila. Development 120, 1385–1398 (1994)

    CAS  PubMed  Google Scholar 

  20. Madden, K., Crowner, D. & Giniger, E. LOLA has the properties of a master regulator of axon-target interaction for SNb motor axons of Drosophila. Dev. Biol. 213, 301–313 (1999)

    Article  CAS  Google Scholar 

  21. Goeke, S. et al. Alternative splicing of lola generates 19 transcription factors controlling axon guidance in Drosophila. Nature Neurosci 6, 917–924 (2003)

    Article  ADS  CAS  Google Scholar 

  22. Ohsako, T., Horiuchi, T., Matsuo, T., Komaya, S. & Aigaki, T. Drosophila lola encodes a family of BTB-transcription regulators with highly variable C-terminal domains containing zinc finger motifs. Gene 311, 59–69 (2003)

    Article  CAS  Google Scholar 

  23. Horowitz, H. & Berg, C. A. The Drosophila pipsqueak gene encodes a nuclear BTB-domain-containing protein required early in oogenesis. Development 122, 1859–1871 (1996)

    CAS  PubMed  Google Scholar 

  24. Weber, U., Siegel, V. & Mlodzik, M. pipsqueak encodes a novel nuclear protein required downstream of seven-up for the development of photoreceptors R3 and R4. EMBO J. 14, 6247–6257 (1995)

    Article  CAS  Google Scholar 

  25. Siegmund, T. & Lehmann, M. The Drosophila Pipsqueak protein defines a new family of helix-turn-helix DNA-binding proteins. Dev. Genes Evol. 212, 152–157 (2002)

    Article  CAS  Google Scholar 

  26. Siegel, V., Jongens, T. A., Jan, L. Y. & Jan, Y. N. pipsqueak, an early acting member of the posterior group of genes, affects vasa level and germ cell-somatic cell interaction in the developing egg chamber. Development 119, 1187–1202 (1993)

    CAS  PubMed  Google Scholar 

  27. Lehmann, M., Siegmund, T., Lintermann, K. G. & Korge, G. The pipsqueak protein of Drosophila melanogaster binds to GAGA sequences through a novel DNA-binding domain. J. Biol. Chem. 273, 28504–28509 (1998)

    Article  CAS  Google Scholar 

  28. Schwendemann, A. & Lehmann, M. Pipsqueak and GAGA factor act in concert as partners at homeotic and many other loci. Proc. Natl Acad. Sci. USA 99, 12883–12888 (2002)

    Article  ADS  CAS  Google Scholar 

  29. Rorth, P. et al. Systematic gain-of-function genetics in Drosophila. Development 125, 1049–1057 (1998)

    CAS  PubMed  Google Scholar 

  30. Barna, M. et al. Plzf mediates transcriptional repression of HoxD gene expression through chromatin remodeling. Dev. Cell 3, 499–510 (2002)

    Article  CAS  Google Scholar 

  31. Melnick, A. et al. Critical residues within the BTB domain of PLZF and Bcl-6 modulate interaction with corepressors. Mol. Cell Biol. 22, 1804–1818 (2002)

    Article  CAS  Google Scholar 

  32. Schotta, G. et al. A silencing pathway to induce H3-K9 and H4-K20 trimethylation at constitutive heterochromatin. Genes Dev. 18, 1251–1262 (2004)

    Article  CAS  Google Scholar 

  33. Irvine, K. D. Fringe, Notch, and making developmental boundaries. Curr. Opin. Genet. Dev. 9, 434–441 (1999)

    Article  CAS  Google Scholar 

  34. Dominguez, M., Ferres-Marco, D., Gutierrez-Avino, F. J., Speicher, S. A. & Beneyto, M. Growth and specification of the eye are controlled independently by Eyegone and Eyeless in Drosophila melanogaster. Nature Genet. 36, 31–39 (2004)

    Article  CAS  Google Scholar 

  35. Chao, J. L., Tsai, Y. C., Chiu, S. J. & Sun, Y. H. Localized Notch signal acts through eyg and upd to promote global growth in Drosophila eye. Development 131, 3839–3847 (2004)

    Article  CAS  Google Scholar 

  36. Bray, S., Musisi, H. & Bienz, M. Bre1 is required for Notch signalling and histone modification. Dev. Cell 8, 279–286 (2005)

    Article  CAS  Google Scholar 

  37. Muller, J. et al. Histone methyltransferase activity of a Drosophila Polycomb group repressor complex. Cell 111, 197–208 (2002)

    Article  CAS  Google Scholar 

  38. Tie, F., Furuyama, T., Prasad-Sinha, J., Jane, E. & Harte, P. J. The Drosophila Polycomb Group proteins ESC and E(Z) are present in a complex containing the histone-binding protein p55 and the histone deacetylase RPD3. Development 128, 275–286 (2001)

    CAS  PubMed  Google Scholar 

  39. Czermin, B. et al. Drosophila enhancer of Zeste/ESC complexes have a histone H3 methyltransferase activity that marks chromosomal Polycomb sites. Cell 111, 185–196 (2002)

    Article  CAS  Google Scholar 

  40. Janody, F. et al. A mosaic genetic screen reveals distinct roles for trithorax and polycomb group genes in Drosophila eye development. Genetics 166, 187–200 (2004)

    Article  CAS  Google Scholar 

  41. Harbour, J. W. & Dean, D. C. The Rb/E2F pathway: expanding roles and emerging paradigms. Genes Dev. 14, 2393–2409 (2000)

    Article  CAS  Google Scholar 

  42. Gowher, H., Leismann, O. & Jeltsch, A. DNA of Drosophila melanogaster contains 5-methylcytosine. EMBO J. 19, 6918–6923 (2000)

    Article  CAS  Google Scholar 

  43. Jabbari, K. & Bernardi, G. Cytosine methylation and CpG, TpG (CpA) and TpA frequencies. Gene 333, 143–149 (2004)

    Article  CAS  Google Scholar 

  44. Salzberg, A., Fisher, O., Siman-Tov, R. & Ankri, S. Identification of methylated sequences in genomic DNA of adult Drosophila melanogaster. Biochem. Biophys. Res. Commun. 322, 465–469 (2004)

    Article  CAS  Google Scholar 

  45. Kunert, N., Marhold, J., Stanke, J., Stach, D. & Lyko, F. A Dnmt2-like protein mediates DNA methylation in Drosophila. Development 130, 5083–5090 (2003)

    Article  CAS  Google Scholar 

  46. Hung, M. S. et al. Drosophila proteins related to vertebrate DNA (5-cytosine) methyltransferases. Proc. Natl Acad. Sci. USA 96, 11940–11945 (1999)

    Article  ADS  CAS  Google Scholar 

  47. Narsa Reddy, M., Tang, L. Y., Lee, T. L. & James Shen, C. K. A candidate gene for Drosophila genome methylation. Oncogene 22, 6301–6303 (2003)

    Article  CAS  Google Scholar 

  48. Marhold, J., Kramer, K., Kremmer, E. & Lyko, F. The Drosophila MBD2/3 protein mediates interactions between the MI-2 chromatin complex and CpT/A-methylated DNA. Development 131, 6033–6039 (2004)

    Article  CAS  Google Scholar 

  49. Baonza, A. & Freeman, M. Control of cell proliferation in the Drosophila eye by notch signalling. Dev. Cell 8, 529–539 (2005)

    Article  CAS  Google Scholar 

  50. Firth, L. C. & Baker, N. E. Extracellular signals responsible for spatially regulated proliferation in the differentiating Drosophila eye. Dev. Cell 8, 541–551 (2005)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank J. Müller, E. Giniger, A. Schwendemann, G. Reuter, F. Lyko, W. Chia, and A. Baonza for reagents; the Bloomington Stock Centre and Exelixis for fly stocks; the Developmental Studies Hybridoma Bank for antibodies; E. Ballesta-Illan for technical assistance; F. J. Garcia-Cozar for sharing quantitative RT–PCR expertise; L. A. Garcia-Alonso, J. Galceran and F. Viana for critically reading the manuscript; and M. Sefton for improvements to the manuscript. I.G.G. is a fellow of the CSIC I3P Programme. This work was supported by grants from the ‘Ministerio de Educación y Ciencia’ of Spain and a European Molecular Biology Organization Young Investigator Award to M.D. Author Contributions D.F-M. conceived the experiment to isolate the mutants; D.F-M, I.G-G, D.M.V, FJ.G-A. and J.B. performed the experiments; and M.D. designed the experiments, carried out the data analysis and wrote the paper.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Maria Dominguez.

Ethics declarations

Competing interests

Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Table 1

EP and GS lines in the lola and psq region. (DOC 71 kb)

Supplementary Figure 1

Map of the EP and GS lines in the intron of lola gene, RT-PCR and in situ hybridisation analyses of the expression of lola and psq in the eyeful GS line. (PDF 190 kb)

Supplementary Figure 2

Images showing representative individuals of EMS-induced revertants of eyeful. (PDF 173 kb)

Supplementary Figure 3

Mutations in the BTB region of psq and lola associated with reversion of the eyeful phenotype and comparison of BTB domains of Drosophila proteins and human BCL-6 and PLZF. (PDF 405 kb)

Supplementary Figure 4

Psq and Lola behave as epigenetic silencers in vivo. (PDF 185 kb)

Supplementary Notes

This file contains Supplementary Methods and Supplementary Figure Legends. (DOC 161 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Ferres-Marco, D., Gutierrez-Garcia, I., Vallejo, D. et al. Epigenetic silencers and Notch collaborate to promote malignant tumours by Rb silencing. Nature 439, 430–436 (2006). https://doi.org/10.1038/nature04376

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature04376

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing