Effect of growth factors on estrogen receptor mediated gene expression.

The proliferation of mammary carcinoma cells can be stimulated by estrogens and various growth factors such as EGF and IGF-I. Steroid hormones and growth factors are understood to exert their effects via different receptors and signal transduction pathways. Recently, it has been shown that growth factors can utilize the unliganded estrogen receptor (ER) as a transcription factor. This study was aimed at identifying the growth factors that can act via the estrogen receptor, and finding new estrogen antagonists that block this activity. Originally, a transcription assay was used in which HeLa cells had been transiently co-transfected with the expression vector for the human ER and a reporter plasmid EREwtc luc. EGF and, to a lesser extent, insulin stimulated the expression of the reporter gene in the absence of estradiol (E2), whereas IGF-I was inactive. The stimulatory effect of E2 and insulin was suppressed when the ER was blocked by the pure antiestrogen ICI 182,780. In ER-positive MCF-7 cells, transfected transiently with the reporter plasmid, EGF had no stimulatory effect on luciferase expression. IGF-I stimulated the transcription to about 50% of the E2 value. Similar activity was found for insulin. The effect of both growth factors was only partly reversed by the addition of a pure antiestrogen. The combination of E2 and IGF-I or insulin led to a synergistic activation of transcription. Because transiently transfected cells do not allow one to study the influence of chromatin structure on gene expression, an MCF-7 subline (MCF-7/2a) was established, in which the reporter construct had been integrated in the genome. IGF-I stimulated luciferase expression in these cells, but showed no overadditive effect with E2. The effects of both agents were completely suppressed by the pure antiestrogen ICI 182,780. These data suggest the existence of an ER-independent mechanism for the activation of the reporter gene in transiently transfected cells, but not in stable transfectants.