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. 1995 Jun 1;129(5):1363–1378. doi: 10.1083/jcb.129.5.1363

bcl-2 overexpression inhibits cell death and promotes the morphogenesis, but not tumorigenesis of human mammary epithelial cells [published erratum appears in J Cell Biol 1995 Nov;131(4):following 1121]

PMCID: PMC2120474  PMID: 7775580

Abstract

Overexpression of the B cell leukemia/lymphoma-2 (bcl-2) gene has been shown to confer a survival advantage on cells by inhibiting apoptosis. In epithelia, the bcl-2 gene is also related to development and differentiation, and the protein is strongly expressed in the embryo in the epithelial cells of the developing mammary gland. To investigate directly the effect of bcl-2 on human epithelial cells, we used an amphotropic recombinant retrovirus to introduce the gene into nontumorigenic cell lines developed from luminal epithelial cells cultured from milk. Here we demonstrate that while bcl-2 overexpression does not directly induce the tumorigenic phenotype, it provides a survival advantage to the mammary epithelial cells by inhibiting cell death at confluence or under conditions of serum starvation, bcl-2 can also affect the phenotype of the original epithelial cells, and promote epithelial-mesenchymal conversion, accompanied by loss of the cell adhesion molecules E-cadherin and alpha 2 beta 1 integrin. The extent of the epithelial-mesenchymal conversion varies with small differences in the phenotype of the parental line and with the level of expression of Bcl-2 and in some cases cell lines emerge with a mixed phenotype. The increased survival of Bcl-2-expressing cells at confluence results in multilayering, and the development of three- dimensional structures. Where a mixed phenotype is observed these structures consist of an outer layer of polarized epithelial cells separated by a basement membrane-like layer from an inner mass of fibroblastoid cells. Branching morphogenesis of bcl-2 transfectants is also observed in collagen gels (in the absence of fibroblast growth factors). The results strongly indicate that by increasing their survival under restrictive growth conditions, and by modifying the epithelial phenotype, bcl-2 can influence the specific morphogenetic behavior of mammary epithelial cells.

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Selected References

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  1. Alnemri E. S., Fernandes T. F., Haldar S., Croce C. M., Litwack G. Involvement of BCL-2 in glucocorticoid-induced apoptosis of human pre-B-leukemias. Cancer Res. 1992 Jan 15;52(2):491–495. [PubMed] [Google Scholar]
  2. Baffy G., Miyashita T., Williamson J. R., Reed J. C. Apoptosis induced by withdrawal of interleukin-3 (IL-3) from an IL-3-dependent hematopoietic cell line is associated with repartitioning of intracellular calcium and is blocked by enforced Bcl-2 oncoprotein production. J Biol Chem. 1993 Mar 25;268(9):6511–6519. [PubMed] [Google Scholar]
  3. Bakhshi A., Jensen J. P., Goldman P., Wright J. J., McBride O. W., Epstein A. L., Korsmeyer S. J. Cloning the chromosomal breakpoint of t(14;18) human lymphomas: clustering around JH on chromosome 14 and near a transcriptional unit on 18. Cell. 1985 Jul;41(3):899–906. doi: 10.1016/s0092-8674(85)80070-2. [DOI] [PubMed] [Google Scholar]
  4. Bartek J., Bartkova J., Kyprianou N., Lalani E. N., Staskova Z., Shearer M., Chang S., Taylor-Papadimitriou J. Efficient immortalization of luminal epithelial cells from human mammary gland by introduction of simian virus 40 large tumor antigen with a recombinant retrovirus. Proc Natl Acad Sci U S A. 1991 May 1;88(9):3520–3524. doi: 10.1073/pnas.88.9.3520. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bartek J., Taylor-Papadimitriou J., Miller N., Millis R. Patterns of expression of keratin 19 as detected with monoclonal antibodies in human breast tissues and tumours. Int J Cancer. 1985 Sep 15;36(3):299–306. [PubMed] [Google Scholar]
  6. Behrens J., Mareel M. M., Van Roy F. M., Birchmeier W. Dissecting tumor cell invasion: epithelial cells acquire invasive properties after the loss of uvomorulin-mediated cell-cell adhesion. J Cell Biol. 1989 Jun;108(6):2435–2447. doi: 10.1083/jcb.108.6.2435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Berdichevsky F., Gilbert C., Shearer M., Taylor-Papadimitriou J. Collagen-induced rapid morphogenesis of human mammary epithelial cells: the role of the alpha 2 beta 1 integrin. J Cell Sci. 1992 Jul;102(Pt 3):437–446. doi: 10.1242/jcs.102.3.437. [DOI] [PubMed] [Google Scholar]
  8. Berdichevsky F., Taylor-Papadimitriou J. Morphological differentiation of hybrids of human mammary epithelial cell lines is dominant and correlates with the pattern of expression of intermediate filaments. Exp Cell Res. 1991 Jun;194(2):267–274. doi: 10.1016/0014-4827(91)90364-z. [DOI] [PubMed] [Google Scholar]
  9. Boyer B., Tucker G. C., Vallés A. M., Franke W. W., Thiery J. P. Rearrangements of desmosomal and cytoskeletal proteins during the transition from epithelial to fibroblastoid organization in cultured rat bladder carcinoma cells. J Cell Biol. 1989 Oct;109(4 Pt 1):1495–1509. doi: 10.1083/jcb.109.4.1495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Burchell J., Durbin H., Taylor-Papadimitriou J. Complexity of expression of antigenic determinants, recognized by monoclonal antibodies HMFG-1 and HMFG-2, in normal and malignant human mammary epithelial cells. J Immunol. 1983 Jul;131(1):508–513. [PubMed] [Google Scholar]
  11. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  12. Church G. M., Gilbert W. Genomic sequencing. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1991–1995. doi: 10.1073/pnas.81.7.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Cleary M. L., Smith S. D., Sklar J. Cloning and structural analysis of cDNAs for bcl-2 and a hybrid bcl-2/immunoglobulin transcript resulting from the t(14;18) translocation. Cell. 1986 Oct 10;47(1):19–28. doi: 10.1016/0092-8674(86)90362-4. [DOI] [PubMed] [Google Scholar]
  14. D'Souza B., Berdichevsky F., Kyprianou N., Taylor-Papadimitriou J. Collagen-induced morphogenesis and expression of the alpha 2-integrin subunit is inhibited in c-erbB2-transfected human mammary epithelial cells. Oncogene. 1993 Jul;8(7):1797–1806. [PubMed] [Google Scholar]
  15. D'souza B., Taylor-Papadimitriou J. Overexpression of ERBB2 in human mammary epithelial cells signals inhibition of transcription of the E-cadherin gene. Proc Natl Acad Sci U S A. 1994 Jul 19;91(15):7202–7206. doi: 10.1073/pnas.91.15.7202. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Deng G., Podack E. R. Suppression of apoptosis in a cytotoxic T-cell line by interleukin 2-mediated gene transcription and deregulated expression of the protooncogene bcl-2. Proc Natl Acad Sci U S A. 1993 Mar 15;90(6):2189–2193. doi: 10.1073/pnas.90.6.2189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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]
  18. Greenburg G., Hay E. D. Cytoskeleton and thyroglobulin expression change during transformation of thyroid epithelium to mesenchyme-like cells. Development. 1988 Mar;102(3):605–622. doi: 10.1242/dev.102.3.605. [DOI] [PubMed] [Google Scholar]
  19. Hockenbery D. M., Zutter M., Hickey W., Nahm M., Korsmeyer S. J. BCL2 protein is topographically restricted in tissues characterized by apoptotic cell death. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):6961–6965. doi: 10.1073/pnas.88.16.6961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kovarik A., Peat N., Wilson D., Gendler S. J., Taylor-Papadimitriou J. Analysis of the tissue-specific promoter of the MUC1 gene. J Biol Chem. 1993 May 5;268(13):9917–9926. [PubMed] [Google Scholar]
  21. Lauerová L., Kovarik J., Bártek J., Rejthar A., Vojtesek B. Novel monoclonal antibodies defining epitope of human cytokeratin 18 molecule. Hybridoma. 1988 Oct;7(5):495–504. doi: 10.1089/hyb.1988.7.495. [DOI] [PubMed] [Google Scholar]
  22. Lu Q. L., Elia G., Lucas S., Thomas J. A. Bcl-2 proto-oncogene expression in Epstein-Barr-virus-associated nasopharyngeal carcinoma. Int J Cancer. 1993 Jan 2;53(1):29–35. doi: 10.1002/ijc.2910530107. [DOI] [PubMed] [Google Scholar]
  23. Lu Q. L., Hanby A. M., Nasser Hajibagheri M. A., Gschmeissner S. E., Lu P. J., Taylor-Papadimitriou J., Krajewski S., Reed J. C., Wright N. A. Bcl-2 protein localizes to the chromosomes of mitotic nuclei and is correlated with the cell cycle in cultured epithelial cell lines. J Cell Sci. 1994 Feb;107(Pt 2):363–371. doi: 10.1242/jcs.107.2.363. [DOI] [PubMed] [Google Scholar]
  24. Lu Q. L., Poulsom R., Wong L., Hanby A. M. Bcl-2 expression in adult and embryonic non-haematopoietic tissues. J Pathol. 1993 Apr;169(4):431–437. doi: 10.1002/path.1711690408. [DOI] [PubMed] [Google Scholar]
  25. Morgenstern J. P., Land H. Advanced mammalian gene transfer: high titre retroviral vectors with multiple drug selection markers and a complementary helper-free packaging cell line. Nucleic Acids Res. 1990 Jun 25;18(12):3587–3596. doi: 10.1093/nar/18.12.3587. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nathan B., Anbazhagan R., Clarkson P., Bartkova J., Gusterson B. Expression of BCL-2 in the developing human fetal and infant breast. Histopathology. 1994 Jan;24(1):73–76. doi: 10.1111/j.1365-2559.1994.tb01273.x. [DOI] [PubMed] [Google Scholar]
  27. Oka H., Shiozaki H., Kobayashi K., Inoue M., Tahara H., Kobayashi T., Takatsuka Y., Matsuyoshi N., Hirano S., Takeichi M. Expression of E-cadherin cell adhesion molecules in human breast cancer tissues and its relationship to metastasis. Cancer Res. 1993 Apr 1;53(7):1696–1701. [PubMed] [Google Scholar]
  28. Osborn M., Debus E., Weber K. Monoclonal antibodies specific for vimentin. Eur J Cell Biol. 1984 May;34(1):137–143. [PubMed] [Google Scholar]
  29. Pellegrini R., Bazzini P., Tosi E., Tagliabue E., Conforti G., Dejana E., Ménard S., Colnaghi M. I. Production and characterization of two monoclonal antibodies directed against the integrin beta 1 chain. Tumori. 1992 Feb 29;78(1):1–4. doi: 10.1177/030089169207800101. [DOI] [PubMed] [Google Scholar]
  30. Pezzella F., Tse A. G., Cordell J. L., Pulford K. A., Gatter K. C., Mason D. Y. Expression of the bcl-2 oncogene protein is not specific for the 14;18 chromosomal translocation. Am J Pathol. 1990 Aug;137(2):225–232. [PMC free article] [PubMed] [Google Scholar]
  31. Reed J. C., Cuddy M., Slabiak T., Croce C. M., Nowell P. C. Oncogenic potential of bcl-2 demonstrated by gene transfer. Nature. 1988 Nov 17;336(6196):259–261. doi: 10.1038/336259a0. [DOI] [PubMed] [Google Scholar]
  32. Reed J. C., Haldar S., Croce C. M., Cuddy M. P. Complementation by BCL2 and C-HA-RAS oncogenes in malignant transformation of rat embryo fibroblasts. Mol Cell Biol. 1990 Aug;10(8):4370–4374. doi: 10.1128/mcb.10.8.4370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Reichmann E., Schwarz H., Deiner E. M., Leitner I., Eilers M., Berger J., Busslinger M., Beug H. Activation of an inducible c-FosER fusion protein causes loss of epithelial polarity and triggers epithelial-fibroblastoid cell conversion. Cell. 1992 Dec 24;71(7):1103–1116. doi: 10.1016/s0092-8674(05)80060-1. [DOI] [PubMed] [Google Scholar]
  34. Shimoyama Y., Hirohashi S., Hirano S., Noguchi M., Shimosato Y., Takeichi M., Abe O. Cadherin cell-adhesion molecules in human epithelial tissues and carcinomas. Cancer Res. 1989 Apr 15;49(8):2128–2133. [PubMed] [Google Scholar]
  35. Strasser A., Harris A. W., Bath M. L., Cory S. Novel primitive lymphoid tumours induced in transgenic mice by cooperation between myc and bcl-2. Nature. 1990 Nov 22;348(6299):331–333. doi: 10.1038/348331a0. [DOI] [PubMed] [Google Scholar]
  36. Takeichi M. Cadherins: a molecular family important in selective cell-cell adhesion. Annu Rev Biochem. 1990;59:237–252. doi: 10.1146/annurev.bi.59.070190.001321. [DOI] [PubMed] [Google Scholar]
  37. Taylor-Papadimitriou J., Berdichevsky F., D'Souza B., Burchell J. Human models of breast cancer. Cancer Surv. 1993;16:59–78. [PubMed] [Google Scholar]
  38. Tenchini M. L., Adams J. C., Gilberty C., Steel J., Hudson D. L., Malcovati M., Watt F. M. Evidence against a major role for integrins in calcium-dependent intercellular adhesion of epidermal keratinocytes. Cell Adhes Commun. 1993 May;1(1):55–66. doi: 10.3109/15419069309095681. [DOI] [PubMed] [Google Scholar]
  39. Tsujimoto Y., Finger L. R., Yunis J., Nowell P. C., Croce C. M. Cloning of the chromosome breakpoint of neoplastic B cells with the t(14;18) chromosome translocation. Science. 1984 Nov 30;226(4678):1097–1099. doi: 10.1126/science.6093263. [DOI] [PubMed] [Google Scholar]
  40. Tsujimoto Y. Overexpression of the human BCL-2 gene product results in growth enhancement of Epstein-Barr virus-immortalized B cells. Proc Natl Acad Sci U S A. 1989 Mar;86(6):1958–1962. doi: 10.1073/pnas.86.6.1958. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Vaux D. L., Cory S., Adams J. M. Bcl-2 gene promotes haemopoietic cell survival and cooperates with c-myc to immortalize pre-B cells. Nature. 1988 Sep 29;335(6189):440–442. doi: 10.1038/335440a0. [DOI] [PubMed] [Google Scholar]
  42. Visser R., van der Beek J. M., Havenith M. G., Cleutjens J. P., Bosman F. T. Immunocytochemical detection of basement membrane antigens in the histopathological evaluation of laryngeal dysplasia and neoplasia. Histopathology. 1986 Feb;10(2):171–180. doi: 10.1111/j.1365-2559.1986.tb02472.x. [DOI] [PubMed] [Google Scholar]
  43. Vleminckx K., Vakaet L., Jr, Mareel M., Fiers W., van Roy F. Genetic manipulation of E-cadherin expression by epithelial tumor cells reveals an invasion suppressor role. Cell. 1991 Jul 12;66(1):107–119. doi: 10.1016/0092-8674(91)90143-m. [DOI] [PubMed] [Google Scholar]
  44. Wayner E. A., Carter W. G. Identification of multiple cell adhesion receptors for collagen and fibronectin in human fibrosarcoma cells possessing unique alpha and common beta subunits. J Cell Biol. 1987 Oct;105(4):1873–1884. doi: 10.1083/jcb.105.4.1873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Willott E., Balda M. S., Heintzelman M., Jameson B., Anderson J. M. Localization and differential expression of two isoforms of the tight junction protein ZO-1. Am J Physiol. 1992 May;262(5 Pt 1):C1119–C1124. doi: 10.1152/ajpcell.1992.262.5.C1119. [DOI] [PubMed] [Google Scholar]
  46. Yunis J. J., Oken M. M., Kaplan M. E., Ensrud K. M., Howe R. R., Theologides A. Distinctive chromosomal abnormalities in histologic subtypes of non-Hodgkin's lymphoma. N Engl J Med. 1982 Nov 11;307(20):1231–1236. doi: 10.1056/NEJM198211113072002. [DOI] [PubMed] [Google Scholar]
  47. Zuk A., Matlin K. S., Hay E. D. Type I collagen gel induces Madin-Darby canine kidney cells to become fusiform in shape and lose apical-basal polarity. J Cell Biol. 1989 Mar;108(3):903–919. doi: 10.1083/jcb.108.3.903. [DOI] [PMC free article] [PubMed] [Google Scholar]

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