Abstract
The role of calcium in the regulation of GnRH gene expression and the mechanism for its effects were examined in the present study. Using the immortalized hypothalamic GT1-7 cell line, which synthesizes and secretes GnRH, we demonstrated by ribonuclease protection assay and Northern blot analysis that these cells respond to treatment with the calcium ionophores ionomycin and A23187 with an inhibition of transcription of the GnRH gene and decreases in GnRH messenger RNA (mRNA) levels. Ionomycin treatment caused the GnRH mRNA half-life to decrease from 25 to 9 h, concomitant with a decrease in mRNA poly(A) tail length, suggesting that ionomycin causes a decrease in GnRH mRNA stability. The ionomycin inhibitory effect on GnRH cytoplasmic mRNA levels was significantly inhibited in the presence of cycloheximide or the RNA synthesis inhibitor 5,6-dichloro-1beta-ribofuranosylbenzimidazole, indicating that novel protein/RNA synthesis is obligatory for this effect. We conclude that an increase in calcium levels caused by ionomycin inhibits GnRH gene expression at multiple levels, including GnRH gene transcription and mRNA stability in GT1-7 cells.
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