Simak Ali, Kirsty Balachandran and Bert O’Malley
Silvia Ottaviani, Greg N Brooke, Ciara O'Hanlon-Brown, Jonathan Waxman, Simak Ali and Laki Buluwela
The development and growth of prostate cancer is dependent on androgens; thus, the identification of androgen-regulated genes in prostate cancer cells is vital for defining the mechanisms of prostate cancer development and progression and developing new markers and targets for prostate cancer treatment. Glycine N-methyltransferase (GNMT) is a S-adenosylmethionine-dependent methyltransferase that has been recently identified as a novel androgen-regulated gene in prostate cancer cells. Although the importance of this protein in prostate cancer progression has been extensively addressed, little is known about the mechanism of its androgen regulation. Here, we show that GNMT expression is stimulated by androgen in androgen receptor (AR) expressing cells and that the stimulation occurs at the mRNA and protein levels. We have identified an androgen response element within the first exon of the GNMT gene and demonstrated that AR binds to this element in vitro and in vivo. Together, these studies identify GNMT as a direct transcriptional target of the AR. As this is an evolutionarily conserved regulatory element, this highlights androgen regulation as an important feature of GNMT regulation.
Ross S Thomas, Naveed Sarwar, Fladia Phoenix, R Charles Coombes and Simak Ali
Phosphorylation of estrogen receptor-α (ERα) at specific residues in transcription activation function 1 (AF-1) can stimulate ERα activity in a ligand-independent manner. This has led to the proposal that AF-1 phosphorylation and the consequent increase in ERα activity could contribute to resistance to endocrine therapies in breast cancer patients. Previous studies have shown that serine 118 (S118) in AF-1 is phosphorylated by extracellular signal-regulated kinases 1 and 2 (Erk1/2) mitogen-activated protein kinase (MAPK) in a ligand-independent manner. Here, we show that serines 104 (S104) and 106 (S106) are also phosphorylated by MAPK in vitro and upon stimulation of MAPK activity in vivo. Phosphorylation of S104 and S106 can be inhibited by the MAP-erk kinase (MEK)1/2 inhibitor U0126 and by expression of kinase-dead Raf1. Further, we show that, although S118 is important for the stimulation of ERα activity by the selective ER modulator 4-hydroxytamoxifen (OHT), S104 and S106 are also required for the agonist activity of OHT. Acidic amino acid substitution of S104 or S106 stimulates ERα activity to a greater extent than the equivalent substitution at S118, suggesting that phosphorylation at S104 and S106 is important for ERα activity. Collectively, these data indicate that the MAPK stimulation of ERα activity involves the phosphorylation not only of S118 but also of S104 and S106, and that MAPK-mediated hyperphosphorylation of ERα at these sites may contribute to resistance to tamoxifen in breast cancer.