G-alpha-i and G-alpha-q proteins mediate the effects of melatonin on steroid hormone receptor function, gene expression and cell proliferation in breast cancer
Description
The pineal gland hormone melatonin has been shown to inhibit the proliferation of estrogen receptor alpha (ERalpha)-positive human breast cancer. The MT1 melatonin G-protein-coupled receptor (GPCR) mediates, at least some of the growth-suppressive actions of melatonin in breast cancer cells. Therefore, we examined the MT1 gene and its expression at the mRNA level by Southern blot analysis and RT-PCR/Southern blot analysis, respectively, in primary human breast tumor cells. Southern blot analysis revealed polymorphisms, an amplification, and deletion of the MT1 gene of human primary breast tumors using the restriction enzyme Hind III. In addition, we demonstrated a wide range of the MT1 receptor expression in primary human breast tumors. No significance was observed between the alterations in the MT1 gene and the expression of the MT1 receptor. Further analysis of the MT1 receptor mRNA expression in the MCF-7 breast cancer cell line, demonstrated that melatonin or estradiol downregulate the expression of MT1 receptor mRNA in the presence of serum. Co-immunoprecipitation experiments conducted in MCF-7 human breast tumor cells, demonstrated that the MT1 melatonin receptor couples to several different G proteins, including the Galphai2, Galphai3 , Galphaq/11 proteins. Via those different G-proteins, melatonin differentially cross talks with several steroid hormone receptor signaling pathways. These studies demonstrate that melatonin suppresses estradiol-induced ERalpha transcriptional activation via the Galphai2 protein, whereas it stimulates all-trans retinoic acid (atRA)-induced RARalpha transcriptional activity via the Galphaq/11 protein. Given that melatonin can regulate steroid hormone receptor transcription activity, we employed cDNA microarray analysis and identified approximately 300 genes regulated by melatonin. These data confirm that the melatonin/membrane MT1 GPCR signaling system can regulate the transcription of numerous genes in breast cancer. Finally, these studies demonstrate that melatonin inhibits the growth of MCF-7 cells through activation of both Galphai and Galphaq proteins. Based on these studies, we propose that the MT1 melatonin receptor functionally couples to both PTX-sensitive (Galpha i2) and PTX-insensitive G proteins (Galphaq and Galpha 11) in MCF-7 cells, and that via these G-proteins and their associated signaling pathways, melatonin can differentially regulate the transcriptional activity of specific steroid/thyroid hormone receptors to inhibit the growth of human breast cancer cells