Thyroid hormone can increase estrogen-mediated transcription from a consensus estrogen response element in neuroblastoma cells

Thyroid hormones (T) and estrogens (E) are nuclear receptor ligands with at least two molecular mechanisms of action: (i) relatively slow genomic effects, such as the regulation of transcription by cognate T receptors (TR) and E receptors (ER); and (ii) relatively rapid nongenomic effects, such as kinase activation and calcium release initiated at the membrane by putative membrane receptors. Genomic and nongenomic effects were thought to be disparate and independent. However, in a previous study using a two-pulse paradigm in neuroblastoma cells, we showed that E acting at the membrane could potentiate transcription from an E-driven reporter gene in the nucleus. Because both T and E can have important effects on mood and cognition, it is possible that the two hormones can act synergistically. In this study, we demonstrate that early actions of T via TRalpha1 and TRbeta1 can potentiate E-mediated transcription (genomic effects) from a consensus E response element (ERE)-driven reporter gene in transiently transfected neuroblastoma cells. Such potentiation was reduced by inhibition of mitogen-activated protein kinase. Using phosphomutants of ERalpha, we also show that probable mitogen-activated protein kinase phosphorylation sites on the ERalpha, the serines at position 167 and 118, are important in TRbeta1-mediated potentiation of ERalpha-induced transactivation. We suggest that crosstalk between T and E includes potential interactions through both nuclear and membrane-initiated molecular mechanisms of hormone signaling.

Calcium Flux in Neuroblastoma Cells Is a Coupling Mechanism between Non-Genomic and Genomic Modes of Estrogens

Estrogens have been demonstrated to rapidly modulate calcium levels in a variety of cell types. However, the significance of estrogen-mediated calcium flux in neuronal cells is largely unknown. The relative importance of intra- and extracellular sources of calcium in estrogenic effects on neurons is also not well understood. Previously, we have demonstrated that membrane-limited estrogens, such as E-BSA given before an administration of a 2-hour pulse of 17beta-estradiol (E(2)), can potentiate the transcription mediated by E(2) from a consensus estrogen response element (ERE)-driven reporter gene. Inhibitors to signal transduction cascades given along with E-BSA or E(2) demonstrated that calcium flux is important for E-BSA-mediated potentiation of transcription in a transiently transfected neuroblastoma cell line. In this report, we have used inhibitors to different voltage-gated calcium channels (VGCCs) and to intracellular store receptors along with E-BSA in the first pulse or with E(2) in the second pulse to investigate the relative importance of these channels to estrogen-mediated transcription. Neither L- nor P-type VGCCs seem to play a role in estrogen action in these cells; while N-type VGCCs are important in both the non-genomic and genomic modes of estrogen action. Specific inhibitors also showed that the ryanodine receptor and the inositol trisphosphate receptor are important to E-BSA-mediated transcriptional potentiation. This report provides evidence that while intracellular stores of calcium are required to couple non-genomic actions of estrogen initiated at the membrane to transcription in the nucleus, extracellular sources of calcium are also important in both non-genomic and genomic actions of estrogens. Copyright (c) 2005 S. Karger AG, Basel.

Estrogens and thyroid hormone: Non-genomic effects are coupled to transcription.

Estrogens and thyroid hormones are regulators of important diverse physiological processes such as reproduction, thermogenesis, neural development, neural differentiation and cardiovascular functions. Both are ligands for receptors in the nuclear receptor superfamily, which act as ligand-dependent transcription factors, regulating transcription. However, estrogens and thyroid hormones also rapidly (within minutes or seconds) activate kinase cascades and calcium increases, presumably initiated at the cell membrane. We discuss the relevance of both modes of hormone action, including the membrane estrogen receptor, to physiology, with particular reference to lordosis behavior. We first showed that estrogen restricted to the membrane can, in fact, lead to subsequent increases in transcription from a consensus estrogen response element-based reporter in the neuroblastoma cell line, SK-N-BE(2)C. Using a novel hormonal paradigm, we also showed that the activation of protein kinase A, protein kinase C, mitogen activated protein kinase and increases in calcium were important in the ability of the membrane-limited estrogen to potentiate transcription. We discuss the source of calcium important in transcriptional potentiation. Since estrogens and thyroid hormones have common effects on neuroprotection, cognition and mood, we also hypothesized that crosstalk could occur between the rapid actions of thyroid hormones and the genomic actions of estrogens. In neural cells, we showed that triiodothyronine acting rapidly via MAPK can increase transcription by the nuclear estrogen receptor ERa from a consensus estrogen response element, possibly by the phosphorylation of the ERa. Novel mechanisms that link signals initiated by hormones from the membrane to the nucleus are physiologically relevant and can achieve neuroendocrine integration

Estrogen receptor-mediated transcription involves the activation of multiple kinase pathways in neuroblastoma cells

While many physiological effects of estrogens (E) are due to regulation of gene transcription by liganded estrogen receptors (ERs), several effects are also mediated, at least in part, by rapid non-genomic actions of E. Though the relative importance of rapid versus genomic effects in the central nervous system is controversial, we showed previously that membrane-limited effects of E, initiated by an estradiol bovine serum albumin conjugate (E2-BSA), could potentiate transcriptional effects of 17beta-estradiol from an estrogen response element (ERE)-reporter in neuroblastoma cells. Here, using specific inhibitors and activators in a pharmacological approach, we show that activation of phosphatidylinositol-3-phosphate kinase (PI3K) and mitogen activated protein kinase (MAPK) pathways, dependent on a Galphaq coupled receptor signaling are important in this transcriptional potentiation. We further demonstrate, using ERalpha phospho-deficient mutants, that E2-BSA mediated phosphorylation of ERalpha is one mechanism to potentiate transcription from an ERE reporter construct. This study provides a possible mechanism by which signaling from the membrane is coupled to transcription in the nucleus, providing an integrated view of hormone signaling in the brain.