The pineal hormone, melatonin, inhibits tumorigenesis and functions as an oncostatic and antiproliferative agent in endocrine responsive breast tumors. Physiological concentrations of melatonin have been shown to inhibit the proliferation of estrogen receptor (ER)-positive MCF-7 human breast cancer cells and to modulate the expression of both ER mRNA and protein. Melatonin inhibition of MCF-7 cell proliferation is serum dependent, indicating an interaction between melatonin and serum components. To examine the effects of melatonin on ER activity, ER transactivation assays were performed using an estrogen response element (ERE)-luciferase reporter construct transiently transfected into MCF-7 cells. Pre-treatment of MCF-7 cells with melatonin for as little as 5 min followed by either epidermal growth factor (EGF) or insulin resulted in the estrogen independent transactivation of the ER. This effect appears to depend on the sequential treatment with melatonin and EGF or insulin, since none of these compounds individually transactivated the ER. This modulation of ER transactivation was associated with changes in ER phosphorylation levels; treatment with melatonin followed by either EGF or insulin increased phosphorylation of the ER, however, melatonin alone decreased basal ER phosphorylation. Here we report that the membrane bound G-protein coupled melatonin receptor, Mel$\sb{\rm 1a}$, is expressed in MCF-7 cells. Further, melatonin modulation of ER transactivation was blocked by pertussis toxin, a G$\sb{\rm\alpha i}$-protein coupled receptor inhibitor, suggesting that the ligand independent transactivation of the ER in MCF-7 cells could result from crosstalk between the G-protein coupled melatonin receptor pathway and the EGF/insulin tyrosine kinase receptor pathways
