TRANSCRIPTIONAL DOWN-REGULATION BY EPIDERMAL GROWTH-FACTOR OF TRH RECEPTOR MESSENGER-RNA IN RAT PITUITARY-CELLS

To gain insight into the mechanism underlying the epidermal growth fac tor (EGF)-induced changes in responsiveness to TRH and in the numbers of TRH receptors (TRH-Rs) in the pituitary, we investigated the transc riptional 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, wh ile no significant changes were observed in beta-actin mRNA levels. Ad dition of actinomycin D caused an acute increase in the basal TRH-R mR NA 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 T RH-R mRNA was not significantly affected in EGF-treated cells. Incubat ion with cycloheximide also induced an increase in the basal TRH-R mRN A level and completely reversed the EGF-induced reduction of TRH-R mRN A levels. Furthermore, a nuclear run-on assay demonstrated that the ra te of transcription of the TRH-R gene was significantly inhibited in c ells treated with EGF. We conclude that (1) EGF decreases the expressi on of the TRH-R mRNA largely by reducing its rate of transcription, an d this action requires the synthesis of new proteins, and (2) inhibito rs of protein and RNA synthesis cause a significant increase in the ba sal TRH-R mRNA level, suggesting that there may be a short-lived prote in suppressing the TRH-R mRNA level in the pituitary.