Thyroid hormone causes mitogen-activated protein kinase-dependent phosphorylation of the nuclear estrogen receptor

Activated by thyroid hormone, the MAPK (ERK1/2) signaling pathway causes serine phosphorylation by MAPK of several nucleoproteins, including the nuclear thyroid hormone receptor β1. Because estrogen can activate MAPK and cause MAPK-dependent serine phosphorylation of nuclear estrogen receptor (ER)α, we studied whether thyroid hormone also promoted MAPK-mediated ERα phosphorylation. Human breast cancer (MCF-7) cells were incubated with physiological concentrations of L-T₄ or 17β-estradiol (E ₂) for 15 min to 24 h, and nuclear ERα and serine-118- phosphorylated ERα were identified by Western blotting. Serine-118-phosphorylated ERα was recovered at 15 min in nuclei of MCF-7 cells exposed to either T₄ or E₂. The T₄ effect was apparent at 15 min and peaked at 2 h, whereas the E₂ effect was maximal at 4-6 h. T₄-agarose was as effective as T₄ in causing phosphorylation of ERα. T₄ action on ERα was inhibited by PD 98059, an inhibitor of ERK1/2 phosphorylation, and by tetraiodothyroacetic acid, a T₄ analog that blocks cell surface-initiated actions of T₄ but is not itself an agonist. Electrophoretic mobility shift assay of nuclear extracts from T ₄-treated and E₂-treated cells showed similar specific protein-DNA-binding. Indexed by [³H]thymidine incorporation and nuclear proliferating cell nuclear antigen, MCF-7 cell proliferation was stimulated by T₄ and T₄-agarose to an extent comparable with the effect of E₂. This T₄ effect was blocked by either PD 98059 or ICI 182,780, an ER antagonist. Thus, T₄, like E₂, causes phosphorylation by MAPK of nuclear ERα at serine-118 in MCF-7 cells and promotes cell proliferation through the ER by a MAPK-dependent pathway.