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

Abstract

Development of strategies for prevention of breast cancer development requires an understanding of the eects of mammary oncogenes on mammary cells at early stages in neoplastic transformation. As mammary oncogenes wnt-1 and int-2 aect dierent signal transduction pathways, we investigated their eects on established mouse mammary epithelial cell lines (MMECLs) re¯ecting early stages in tumorigenesis. Normal interactions between b-catenin and E-cadherin were abrogated in all three immortalized MMECLs and the cells lacked b-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 con®rmed 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 b-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 eects 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 eects in the immortalized FSK4 and EL12 cells were not sucient to mediate a tumorigenic phenotype. This study showed that the wnt-1 and int-2 oncogenes have similar but distinguishable eects on immortalized MMECLs and that the genetic background of the mammary cells greatly in¯uences the consequences of oncogene expression at early stages of cell transformation. Oncogene (2001) 20, 7645 ± 7657.

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