Estrogen receptor regulation of the Na+/H+ exchanger regulatory factor

To better understand the actions of estrogens and antiestrogens in estrogen target cells, we have searched for estrogen-regulated genes in human breas t cancer cells, in which the number of genes known to be directly activated by estrogen is quite small. Using differential display RNA methods, we hav e identified the human homolog of the Na2+-H+ exchanger regulatory factor ( NHE-RF), an approximately 50-kDa protein that is also an ezrin-radixin-moes in-binding phosphoprotein, as being under rapid and direct regulation by es trogen in estrogen receptor (ER)-containing breast cancer cells. Stimulatio n by estrogen of NHE-RF RNA is rapid, being near maximal (similar to 6-fold ) by Ih, and is not blocked by cycloheximide, indicating that it is a prima ry response. Stimulation is selective for estrogen Ligands, with no stimula tion by other classes of steroid hormones, and stimulation by estrogen is s uppressed by the antiestrogens tamoxifen and ICI 182,780. Induction is show n to require an active ER through several approaches, including the use of ER-negative breast cancer cells containing a stably integrated ER. NHE-RF p rotein levels, monitored using antibodies specific for this protein, increa se after estrogen and reach ma;maximal levels at 24-48 h. Interestingly, NH E-RF is a PDZ domain-containing protein that is enriched in polarized epith elia, where it is known to be localized in microvilli. Among various human tissues we have examined, we found that NHE-RF is expressed at a fairly hig h level in mammary tissue. NHE-RF regulates protein kinase A inhibition of the Na+-H+ exchanger and may serve as a scaffold adaptor protein that contr ibutes to the specificity of signal transduction events. Our findings sugge st that the early, known effects of estrogen on cell cytoarchitecture (e.g, increasing microvilli on breast cancer cells) and on some cell signaling p athways (e,g, those involving cAMP) may involve rapid estrogen-mediated cha nges in the production of NHE-RF.