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To gain insight into the mechanism underlying the epidermal growth factor (EGF)-induced changes in responsiveness to TRH and in the numbers of TRH receptors (TRH-Rs) in the pituitary, we investigated the transcriptional regulation by EGF of the TRH-R gene in GH4C1 cells. Northern blot analyses and binding studies revealed that EGF reduced both TRH binding and TRH-R mRNA levels in a dose- and time-dependent manner, while no significant changes were observed in β-actin mRNA levels. Addition of actinomycin D caused an acute increase in the basal TRH-R mRNA level, and the rate of decrease of the TRH-R mRNA was identical in control and EGF-treated groups, suggesting that the stability of the TRH-R mRNA was not significantly affected in EGF-treated cells. Incubation with cycloheximide also induced an increase in the basal TRH-R mRNA level and completely reversed the EGF-induced reduction of TRH-R mRNA levels. Furthermore, a nuclear run-on assay demonstrated that the rate of transcription of the TRH-R gene was significantly inhibited in cells treated with EGF. We conclude that (1) EGF decreases the expression of the TRH-R mRNA largely by reducing its rate of transcription, and this action requires the synthesis of new proteins, and (2) inhibitors of protein and RNA synthesis cause a significant increase in the basal TRH-R mRNA level, suggesting that there may be a short-lived protein suppressing the TRH-R mRNA level in the pituitary.

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