Regulation of endogenous thyrotropin-releasing hormone receptor messenger RNA in GH4C1 cells: roles of protein and RNA synthesis.
In previous studies on the regulation of the endogenous TRH receptor (TRHR) in GH4C1 cells, we have shown that the synthetic glucocorticoid dexamethasone (Dex) increased TRHR abundance on the plasma membrane of these cells as well as increasing the concentration of TRHR mRNA. In the present investigation, we determined whether the action of Dex on TRHR mRNA was direct or whether it involved the synthesis of intermediary protein(s). We found that the protein synthesis inhibitors cycloheximide or anisomycin alone enhanced TRHR mRNA accumulation and, in the presence of Dex, caused a 10- to 20-fold superinduction of TRHR mRNA. The superinduction required inhibition of protein synthesis. Results of nuclear run-on assays showed that superinduction was accompanied by a large increase in transcription of TRHR mRNA. Experiments designed to examine the stability of TRHR mRNA transcripts revealed that inhibitors of RNA synthesis (actinomycin D and dichlorobenzimidazole riboside) caused an increase in endogenous TRHR mRNA concentrations by themselves, complicating their use to estimate TRHR mRNA half-life. We conclude that Dex stimulates transcription of endogenous TRHR mRNA by a mechanism that does not require enhanced synthesis of new proteins. In fact, ongoing protein synthesis reduces steady state TRHR mRNA concentrations consistent with a role for a labile protein(s) in suppressing TRHR gene transcription; an additional action of a labile protein to enhance TRHR mRNA degradation is also possible.
Yang
J
J
Department of Molecular and Cellular Toxicology, Harvard School of Public Health, Boston, Massachusetts 02115.
Tashjian
A H
AH
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D004004:Dichlororibofuranosylbenzimidazole / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D007700:Kinetics; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010911:Pituitary Neoplasms; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis / Q000378:metabolism*; D012337:RNA, Ribosomal, 18S / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000096:biosynthesis* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D012337:RNA, Ribosomal, 18S; D018025:Receptors, Thyrotropin-Releasing Hormone; D003609:Dactinomycin; D004004:Dichlororibofuranosylbenzimidazole; D003907:Dexamethasone; D003513:Cycloheximide