Transrepression of Estrogen Receptor β Signaling by Nuclear Factor-κB in Ovarian Granulosa Cells

Abstract

Estrogen receptor (ER) beta is the predominant ER in granulosa cells of the ovary. ERbeta is expressed at high levels in granulosa cell tumors (GCT) and in the human GCT-derived cell lines, COV434 and KGN. To gain insight into ERbeta function in granulosa cells and in GCT, we have used the COV434 and KGN cell lines. Although the cells bind estradiol (E2), transcriptional activation of a transfected estrogen-responsive reporter vector construct (ERE2-luc) by E2 was not observed. Transactivation was also not observed with cotransfected ERalpha or beta. This transcriptional resistance is specific to steroid receptor transactivation; reporter plasmids that are activated by the transcription factors activator protein 1 (AP-1) and nuclear factor kappaB (NF-kappaB) demonstrate both constitutive and inducible transactivation. AP-1 and NF-kappaB are known to cause transrepression of both ERalpha- and glucocorticoid receptor-mediated transcription. We therefore examined the possibility that activation of these pathways was responsible for the lack of a response to estrogen by using inhibitors of AP-1 or NF-kappaB. The AP-1 inhibitors alone had no effect, whereas inhibition of NF-kappaB signaling allowed a 3- to 4-fold E2-mediated induction of ERE2-luc. This response was both ligand and ER dependent. Repression of ERbeta signaling by NF-kappaB has not previously been reported. Recent evidence suggests that ERbeta may function to promote differentiation. The inhibition of ERbeta in combination with the antiapoptotic properties of NF-kappaB may therefore contribute to pathogenesis of GCT.

The FOXL2 C134W mutation is characteristic of adult granulosa cell tumors of the ovary

“…20 COV434 cells were maintained in DMEM and KGN cells in DMEM/F12 both supplemented with 10% FCS at 37 1C in a 95% air/5% CO 2 humidified incubator.…”

Jun proteins modulate the ovary-specific promoter of aromatase gene in ovarian granulosa cells via a cAMP-responsive element

“…Previous characterization of this cell line clearly shows that it maintains most of the physiological functions of granulosa cells in vivo such as steroidogenesis, cell growth, and apoptosis (Nishi et al, 2001;Mukasa et al, 2003;Chu et al, 2004;Morinaga et al, 2004), thus validating the utility of this cell line in studying ovarian gene regulation. Our study revealed a potent activity of Jun proteins in downregulating the ovary-specific promoter (PII) of the aromatase gene.…”

“…The KGN granulosalike tumor cell line has been used in previous studies of granulosa cell differentiation and steroidogenesis (Nishi et al, 2001;Mukasa et al, 2003;Chu et al, 2004;Morinaga et al, 2004). Asynchronously growing cells were treated with either vehicle or 25 mM forskolin, a known activator of adenylyl cyclase, for various periods of time.…”

Soy Promotes Juvenile Granulosa Cell Tumor Development in Mice and in the Human Granulosa Cell Tumor-Derived COV434 Cell Line1

“…A few number of studies have focused on the effects of estrogen signaling in human GCT [37] or in human COV434 and KGN cell lines, and estradiol signaling in these cell lines is defective [38]. To examine the effect of genistein treatment in JGCT, we treated COV434 cells with genistein (5, 25, or 50 lM) and monitored cell viability for 72 h. The number of viable cells increased significantly at all doses of genistein as early as 24 h posttreatment (3%-9%), with a further induction occurring after 48 h (20%-34%) and 72 h (up to 51%) ( Fig.…”