Figure 1.

The HER4 intracellular domain (4ICD) is necessary and sufficient to potentiate estrogen stimulated gene expression. (A) MCF-7/B (MCF-7 cells with ectopic BCL-2 expression) [22] breast cancer cells were incubated in growth media supplemented with 5% charcoal-stripped FBS (CS-FBS) for 24 hrs and cotransfected with 200 ng of ERE-luc and the indicated HER4 (JM-a, Cyt1 isoform) or 4ICD expression vectors. At 24 hrs post-transfection cells were cultured in the presence or absence of 100 pM 17-β-estradiol (Estrogen) and the luciferase reporter gene assay was performed after 16 hrs as described previously [5]. Each sample was prepared in duplicate and the entire experiment was repeated at least three times. Data represents the mean and standard error of luciferase activity normalized to untreated cells transfected with ERE-Luc alone. (B) Protein lysates prepared from the same experimental treatments were analyzed by western blot for HER4/4ICD (Cell Signaling 111B2) expression with β-tubulin (Upstate DM1A) included as a loading control.