Influence of Exogenous Tissue Factor on Estrogen Receptor α Expression in Breast Cancer Cells: Involvement of β1-Integrin, PAR2, and Mitogen-Activated Protein Kinase Activation

Increased expression of tissue factor (TF) has been associated with invasive forms of breast cancer. Conversely, the loss of estrogen receptor alpha (ER alpha) is associated with increased cell invasiveness. We have examined the influence of exogenous truncated recombinant TF (rTF) on ER alpha expression and cell invasiveness and investigated the mechanism of rTF signaling. The influence of rTF on ERa expression in MCF-7 and T47D cell lines was investigated using reverse transcription-PCR and ELISA. Cell invasion was measured using Boyden chamber-based invasion assays. Additionally, the interaction of fluorescein-labeled rTF with the surface of MCF-7 cells and particularly with beta(1)-integrin was examined. Treatment of cells with rTF resulted in the down-regulation of ER alpha mRNA and protein over 24 h, which required beta(1)-integrin and involved the mitogen-activated protein kinase pathway but did not require PAR2 activation. The addition of rTF reduced estradiol-mediated cell proliferation as well as increased cell invasiveness requiring both PAR2 and beta(1)-integrin activation. Fluorescein-labeled rTF was shown to bind to the surface of MCF-7 cells within 5 min and peaked at 15 min. The bound rTF colocalized with cellular beta(1)-integrin and was disrupted in the presence of excess unlabeled rTF and an anti-beta(1) polyclonal antibody. Finally, affinity purification of beta(1)-integrin using rTF-conjugated agarose showed a requirement for the presence of divalent cations but not factor Vila. The results indicate that rTF is capable of down-regulating ER alpha expression in breast cancer cells, resulting in decreases in estrogen-mediated cell proliferation and increased invasiveness. Furthermore, the mechanisms by which rTF induces these changes involve both PAR2 and beta(1)-integrin. (Mol Cancer Res 2008;6(12):1807-18)