Skip to main content
NCBI home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1997 Oct;17(10):6076–6086. doi: 10.1128/mcb.17.10.6076

Interference with the expression of a novel human polycomb protein, hPc2, results in cellular transformation and apoptosis.

D P Satijn 1, D J Olson 1, J van der Vlag 1, K M Hamer 1, C Lambrechts 1, H Masselink 1, M J Gunster 1, R G Sewalt 1, R van Driel 1, A P Otte 1
PMCID: PMC232457  PMID: 9315667

Abstract

Polycomb (Pc) is involved in the stable and heritable repression of homeotic gene activity during Drosophila development. Here, we report the identification of a novel human Pc homolog, hPc2. This gene is more closely related to a Xenopus Pc homolog, XPc, than to a previously described human Pc homolog, CBX2 (hPc1). However, the hPc2 and CBX2/hPc1 proteins colocalize in interphase nuclei of human U-2 OS osteosarcoma cells, suggesting that the proteins are part of a common protein complex. To study the functions of the novel human Pc homolog, we generated a mutant protein, delta hPc2, which lacks an evolutionarily conserved C-terminal domain. This C-terminal domain is important for hPc2 function, since the delta hPc2 mutant protein which lacks the C-terminal domain is unable to repress gene activity. Expression of the delta hPc2 protein, but not of the wild-type hPc2 protein, results in cellular transformation of mammalian cell lines as judged by phenotypic changes, altered marker gene expression, and anchorage-independent growth. Specifically in delta hPc2-transformed cells, the expression of the c-myc proto-oncogene is strongly enhanced and serum deprivation results in apoptosis. In contrast, overexpression of the wild-type hPc2 protein results in decreased c-myc expression. Our data suggest that hPc2 is a repressor of proto-oncogene activity and that interference with hPc2 function can lead to derepression of proto-oncogene transcription and subsequently to cellular transformation.

Full Text

The Full Text of this article is available as a PDF (3.0 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alkema M. J., Bronk M., Verhoeven E., Otte A., van 't Veer L. J., Berns A., van Lohuizen M. Identification of Bmi1-interacting proteins as constituents of a multimeric mammalian polycomb complex. Genes Dev. 1997 Jan 15;11(2):226–240. doi: 10.1101/gad.11.2.226. [DOI] [PubMed] [Google Scholar]
  2. Bradley R. S., Brown A. M. A soluble form of Wnt-1 protein with mitogenic activity on mammary epithelial cells. Mol Cell Biol. 1995 Aug;15(8):4616–4622. doi: 10.1128/mcb.15.8.4616. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brown A. M., Wildin R. S., Prendergast T. J., Varmus H. E. A retrovirus vector expressing the putative mammary oncogene int-1 causes partial transformation of a mammary epithelial cell line. Cell. 1986 Sep 26;46(7):1001–1009. doi: 10.1016/0092-8674(86)90699-9. [DOI] [PubMed] [Google Scholar]
  4. Bunker C. A., Kingston R. E. Transcriptional repression by Drosophila and mammalian Polycomb group proteins in transfected mammalian cells. Mol Cell Biol. 1994 Mar;14(3):1721–1732. doi: 10.1128/mcb.14.3.1721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cohen K. J., Hanna J. S., Prescott J. E., Dang C. V. Transformation by the Bmi-1 oncoprotein correlates with its subnuclear localization but not its transcriptional suppression activity. Mol Cell Biol. 1996 Oct;16(10):5527–5535. doi: 10.1128/mcb.16.10.5527. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Coré N., Bel S., Gaunt S. J., Aurrand-Lions M., Pearce J., Fisher A., Djabali M. Altered cellular proliferation and mesoderm patterning in Polycomb-M33-deficient mice. Development. 1997 Feb;124(3):721–729. doi: 10.1242/dev.124.3.721. [DOI] [PubMed] [Google Scholar]
  7. Evan G. I., Lewis G. K., Ramsay G., Bishop J. M. Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol Cell Biol. 1985 Dec;5(12):3610–3616. doi: 10.1128/mcb.5.12.3610. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Evan G. I., Wyllie A. H., Gilbert C. S., Littlewood T. D., Land H., Brooks M., Waters C. M., Penn L. Z., Hancock D. C. Induction of apoptosis in fibroblasts by c-myc protein. Cell. 1992 Apr 3;69(1):119–128. doi: 10.1016/0092-8674(92)90123-t. [DOI] [PubMed] [Google Scholar]
  9. Evan G., Harrington E., Fanidi A., Land H., Amati B., Bennett M. Integrated control of cell proliferation and cell death by the c-myc oncogene. Philos Trans R Soc Lond B Biol Sci. 1994 Aug 30;345(1313):269–275. doi: 10.1098/rstb.1994.0105. [DOI] [PubMed] [Google Scholar]
  10. Fanidi A., Harrington E. A., Evan G. I. Cooperative interaction between c-myc and bcl-2 proto-oncogenes. Nature. 1992 Oct 8;359(6395):554–556. doi: 10.1038/359554a0. [DOI] [PubMed] [Google Scholar]
  11. Franke A., DeCamillis M., Zink D., Cheng N., Brock H. W., Paro R. Polycomb and polyhomeotic are constituents of a multimeric protein complex in chromatin of Drosophila melanogaster. EMBO J. 1992 Aug;11(8):2941–2950. doi: 10.1002/j.1460-2075.1992.tb05364.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Franke A., Messmer S., Paro R. Mapping functional domains of the polycomb protein of Drosophila melanogaster. Chromosome Res. 1995 Sep;3(6):351–360. doi: 10.1007/BF00710016. [DOI] [PubMed] [Google Scholar]
  13. Gavin B. J., McMahon A. P. Differential regulation of the Wnt gene family during pregnancy and lactation suggests a role in postnatal development of the mammary gland. Mol Cell Biol. 1992 May;12(5):2418–2423. doi: 10.1128/mcb.12.5.2418. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gecz J., Gaunt S. J., Passage E., Burton R. D., Cudrey C., Pearce J. J., Fontes M. Assignment of a Polycomb-like chromobox gene (CBX2) to human chromosome 17q25. Genomics. 1995 Mar 1;26(1):130–133. doi: 10.1016/0888-7543(95)80091-y. [DOI] [PubMed] [Google Scholar]
  15. Gunster M. J., Satijn D. P., Hamer K. M., den Blaauwen J. L., de Bruijn D., Alkema M. J., van Lohuizen M., van Driel R., Otte A. P. Identification and characterization of interactions between the vertebrate polycomb-group protein BMI1 and human homologs of polyhomeotic. Mol Cell Biol. 1997 Apr;17(4):2326–2335. doi: 10.1128/mcb.17.4.2326. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Isfort R. J., Cody D. B., Lovell G., Doersen C. J. Analysis of oncogenes, tumor suppressor genes, autocrine growth-factor production, and differentiation state of human osteosarcoma cell lines. Mol Carcinog. 1995 Nov;14(3):170–178. doi: 10.1002/mc.2940140306. [DOI] [PubMed] [Google Scholar]
  17. Kanno M., Hasegawa M., Ishida A., Isono K., Taniguchi M. mel-18, a Polycomb group-related mammalian gene, encodes a transcriptional negative regulator with tumor suppressive activity. EMBO J. 1995 Nov 15;14(22):5672–5678. doi: 10.1002/j.1460-2075.1995.tb00254.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kingston R. E., Bunker C. A., Imbalzano A. N. Repression and activation by multiprotein complexes that alter chromatin structure. Genes Dev. 1996 Apr 15;10(8):905–920. doi: 10.1101/gad.10.8.905. [DOI] [PubMed] [Google Scholar]
  19. Koopman G., Reutelingsperger C. P., Kuijten G. A., Keehnen R. M., Pals S. T., van Oers M. H. Annexin V for flow cytometric detection of phosphatidylserine expression on B cells undergoing apoptosis. Blood. 1994 Sep 1;84(5):1415–1420. [PubMed] [Google Scholar]
  20. Lewis E. B. A gene complex controlling segmentation in Drosophila. Nature. 1978 Dec 7;276(5688):565–570. doi: 10.1038/276565a0. [DOI] [PubMed] [Google Scholar]
  21. Martin S. J., Reutelingsperger C. P., McGahon A. J., Rader J. A., van Schie R. C., LaFace D. M., Green D. R. Early redistribution of plasma membrane phosphatidylserine is a general feature of apoptosis regardless of the initiating stimulus: inhibition by overexpression of Bcl-2 and Abl. J Exp Med. 1995 Nov 1;182(5):1545–1556. doi: 10.1084/jem.182.5.1545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Messmer S., Franke A., Paro R. Analysis of the functional role of the Polycomb chromo domain in Drosophila melanogaster. Genes Dev. 1992 Jul;6(7):1241–1254. doi: 10.1101/gad.6.7.1241. [DOI] [PubMed] [Google Scholar]
  23. Müller J., Gaunt S., Lawrence P. A. Function of the Polycomb protein is conserved in mice and flies. Development. 1995 Sep;121(9):2847–2852. doi: 10.1242/dev.121.9.2847. [DOI] [PubMed] [Google Scholar]
  24. Müller J. Transcriptional silencing by the Polycomb protein in Drosophila embryos. EMBO J. 1995 Mar 15;14(6):1209–1220. doi: 10.1002/j.1460-2075.1995.tb07104.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Olson D. J., Papkoff J. Regulated expression of Wnt family members during proliferation of C57mg mammary cells. Cell Growth Differ. 1994 Feb;5(2):197–206. [PubMed] [Google Scholar]
  26. Orlando V., Paro R. Mapping Polycomb-repressed domains in the bithorax complex using in vivo formaldehyde cross-linked chromatin. Cell. 1993 Dec 17;75(6):1187–1198. doi: 10.1016/0092-8674(93)90328-n. [DOI] [PubMed] [Google Scholar]
  27. Paro R., Hogness D. S. The Polycomb protein shares a homologous domain with a heterochromatin-associated protein of Drosophila. Proc Natl Acad Sci U S A. 1991 Jan 1;88(1):263–267. doi: 10.1073/pnas.88.1.263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Paro R. Imprinting a determined state into the chromatin of Drosophila. Trends Genet. 1990 Dec;6(12):416–421. doi: 10.1016/0168-9525(90)90303-n. [DOI] [PubMed] [Google Scholar]
  29. Pearce J. J., Singh P. B., Gaunt S. J. The mouse has a Polycomb-like chromobox gene. Development. 1992 Apr;114(4):921–929. doi: 10.1242/dev.114.4.921. [DOI] [PubMed] [Google Scholar]
  30. Pelegri F., Lehmann R. A role of polycomb group genes in the regulation of gap gene expression in Drosophila. Genetics. 1994 Apr;136(4):1341–1353. doi: 10.1093/genetics/136.4.1341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rastelli L., Chan C. S., Pirrotta V. Related chromosome binding sites for zeste, suppressors of zeste and Polycomb group proteins in Drosophila and their dependence on Enhancer of zeste function. EMBO J. 1993 Apr;12(4):1513–1522. doi: 10.1002/j.1460-2075.1993.tb05795.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Reijnen M. J., Hamer K. M., den Blaauwen J. L., Lambrechts C., Schoneveld I., van Driel R., Otte A. P. Polycomb and bmi-1 homologs are expressed in overlapping patterns in Xenopus embryos and are able to interact with each other. Mech Dev. 1995 Sep;53(1):35–46. doi: 10.1016/0925-4773(95)00422-x. [DOI] [PubMed] [Google Scholar]
  33. Samuels M. L., Weber M. J., Bishop J. M., McMahon M. Conditional transformation of cells and rapid activation of the mitogen-activated protein kinase cascade by an estradiol-dependent human raf-1 protein kinase. Mol Cell Biol. 1993 Oct;13(10):6241–6252. doi: 10.1128/mcb.13.10.6241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Satijn D. P., Gunster M. J., van der Vlag J., Hamer K. M., Schul W., Alkema M. J., Saurin A. J., Freemont P. S., van Driel R., Otte A. P. RING1 is associated with the polycomb group protein complex and acts as a transcriptional repressor. Mol Cell Biol. 1997 Jul;17(7):4105–4113. doi: 10.1128/mcb.17.7.4105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Schön A., Michiels L., Janowski M., Merregaert J., Erfle V. Expression of protooncogenes in murine osteosarcomas. Int J Cancer. 1986 Jul 15;38(1):67–74. doi: 10.1002/ijc.2910380112. [DOI] [PubMed] [Google Scholar]
  36. Simon J. Locking in stable states of gene expression: transcriptional control during Drosophila development. Curr Opin Cell Biol. 1995 Jun;7(3):376–385. doi: 10.1016/0955-0674(95)80093-x. [DOI] [PubMed] [Google Scholar]
  37. Singh P. B., Miller J. R., Pearce J., Kothary R., Burton R. D., Paro R., James T. C., Gaunt S. J. A sequence motif found in a Drosophila heterochromatin protein is conserved in animals and plants. Nucleic Acids Res. 1991 Feb 25;19(4):789–794. doi: 10.1093/nar/19.4.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Small M. B., Hay N., Schwab M., Bishop J. M. Neoplastic transformation by the human gene N-myc. Mol Cell Biol. 1987 May;7(5):1638–1645. doi: 10.1128/mcb.7.5.1638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Smith P. K., Krohn R. I., Hermanson G. T., Mallia A. K., Gartner F. H., Provenzano M. D., Fujimoto E. K., Goeke N. M., Olson B. J., Klenk D. C. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985 Oct;150(1):76–85. doi: 10.1016/0003-2697(85)90442-7. [DOI] [PubMed] [Google Scholar]
  40. Smouse D., Goodman C., Mahowald A., Perrimon N. polyhomeotic: a gene required for the embryonic development of axon pathways in the central nervous system of Drosophila. Genes Dev. 1988 Jul;2(7):830–842. doi: 10.1101/gad.2.7.830. [DOI] [PubMed] [Google Scholar]
  41. Smouse D., Perrimon N. Genetic dissection of a complex neurological mutant, polyhomeotic, in Drosophila. Dev Biol. 1990 May;139(1):169–185. doi: 10.1016/0012-1606(90)90286-r. [DOI] [PubMed] [Google Scholar]
  42. Stone J., de Lange T., Ramsay G., Jakobovits E., Bishop J. M., Varmus H., Lee W. Definition of regions in human c-myc that are involved in transformation and nuclear localization. Mol Cell Biol. 1987 May;7(5):1697–1709. doi: 10.1128/mcb.7.5.1697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Tagawa M., Sakamoto T., Shigemoto K., Matsubara H., Tamura Y., Ito T., Nakamura I., Okitsu A., Imai K., Taniguchi M. Expression of novel DNA-binding protein with zinc finger structure in various tumor cells. J Biol Chem. 1990 Nov 15;265(32):20021–20026. [PubMed] [Google Scholar]
  44. Wagner A. J., Kokontis J. M., Hay N. Myc-mediated apoptosis requires wild-type p53 in a manner independent of cell cycle arrest and the ability of p53 to induce p21waf1/cip1. Genes Dev. 1994 Dec 1;8(23):2817–2830. doi: 10.1101/gad.8.23.2817. [DOI] [PubMed] [Google Scholar]
  45. Zink B., Paro R. In vivo binding pattern of a trans-regulator of homoeotic genes in Drosophila melanogaster. Nature. 1989 Feb 2;337(6206):468–471. doi: 10.1038/337468a0. [DOI] [PubMed] [Google Scholar]
  46. van Lohuizen M., Verbeek S., Scheijen B., Wientjens E., van der Gulden H., Berns A. Identification of cooperating oncogenes in E mu-myc transgenic mice by provirus tagging. Cell. 1991 May 31;65(5):737–752. doi: 10.1016/0092-8674(91)90382-9. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES