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  • Original Paper
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Wnt-1 and int-2 mammary oncogene effects on the β-catenin pathway in immortalized mouse mammary epithelial cells are not sufficient for tumorigenesis

Oncogene volume 20, pages 7645–7657 (2001)Cite this article

Abstract

Development of strategies for prevention of breast cancer development requires an understanding of the effects of mammary oncogenes on mammary cells at early stages in neoplastic transformation. As mammary oncogenes wnt-1 and int-2 affect different signal transduction pathways, we investigated their effects on established mouse mammary epithelial cell lines (MMECLs) reflecting early stages in tumorigenesis. Normal interactions between β-catenin and E-cadherin were abrogated in all three immortalized MMECLs and the cells lacked β-catenin-mediated transactivation activity, detectable using a reporter assay, suggesting that alterations in cell adhesion may be very early events in mammary tumorigenesis. Immortalized FSK4 and EL12 cells and hyperplastic TM3 cells were stably transfected with expression vectors encoding wnt-1 or int-2 or the control vector, and drug-selected pooled cells from each line were confirmed by reverse transcription-polymerase chain reaction to express the transfected oncogene; this expression persisted in the cells analysed in vitro and in vivo. Resultant phenotypic changes depended both on the oncogene and the target mammary cell line. In FSK4 cells, expression of wnt-1 or int-2 resulted in proliferative changes in vitro, including reduced contact inhibition, increased β-catenin expression, and decreased p53 transcriptional activity, but neither oncogene conferred upon those cells the ability to produce tumors in vivo. EL12 cells were highly refractory to the effects of both oncogenes, with the only measurable changes being increased E-cadherin levels induced by both oncogenes and increased proliferation of the int-2-transfected cells in the absence of serum. Parental TM3 cells were phenotypically similar to wnt-1- or int-2-transfected FSK4 cells and displayed an increased rate of proliferation in vitro and markedly increased tumorigenicity in vivo following transfection with int-2 but not with wnt-1. These results suggest that wnt-1 signaling is redundant in the hyperplastic TM3 cells and indicate that wnt-1-induced effects in the immortalized FSK4 and EL12 cells were not sufficient to mediate a tumorigenic phenotype. This study showed that the wnt-1 and int-2 oncogenes have similar but distinguishable effects on immortalized MMECLs and that the genetic background of the mammary cells greatly influences the consequences of oncogene expression at early stages of cell transformation.

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Abbreviations

CMV:

cytomegalovirus

ECL:

enhanced chemiluminescence

EGF:

epidermal growth factor

FGF:

fibroblast growth factor

FGFR:

FGF receptor

MMECL:

mouse mammary epithelial cell line

MMTV:

mouse mammary tumor virus

MTT:

3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide

PAGE:

polyacrylamide gel electrophoresis

PBS:

phosphate-buffered saline

RT–PCR:

reverse transcription polymerase chain reaction

SDS:

sodium dodecyl sulfate

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Acknowledgements

The authors thank C Wong, JA Lednicky and PJ Bonilla for assistance and helpful advice. This work was supported in part by research grant CA25215 from the National Cancer Institute and by the Research Training Program in Breast Cancer (BC980058) from the US Army Medical Research and Materiel Command.

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  1. C Annette Hollmann

    Present address: Neuroimmunology Unit, Montreal Neurological Institute, 3801 University St., Montreal, H3A 2B4, PQ, Canada

Authors and Affiliations

  1. Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas, USA

    C Annette Hollmann & Janet S Butel

  2. Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas, USA

    Frances S Kittrell & Daniel Medina

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  1. C Annette Hollmann
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  2. Frances S Kittrell
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Correspondence to Janet S Butel.

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Hollmann, C., Kittrell, F., Medina, D. et al. Wnt-1 and int-2 mammary oncogene effects on the β-catenin pathway in immortalized mouse mammary epithelial cells are not sufficient for tumorigenesis. Oncogene 20, 7645–7657 (2001). https://doi.org/10.1038/sj.onc.1204980

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