Thyrotropin-releasing hormone (TRH) and phorbol myristate acetate decrease TRH receptor messenger RNA in rat pituitary GH3 cells: evidence that protein kinase-C mediates the TRH effect.
In a previous report we showed that TRH-induced down-regulation of the density of its receptors (TRH-Rs) on rat pituitary tumor (GH3) cells was preceded by a decrease in the activity of the mRNA for the TRH-R, as assayed in Xenopus oocytes. Here we report the effects of TRH, elevation of cytoplasmic free Ca2+ concentration, phorbol myristate acetate (PMA), and H-7 [1-(5-isoquinolinesulfonyl)2-methylpiperazine dihydrochloride], an inhibitor of protein kinases, on the levels of TRH-R mRNA, which were measured by Northern analysis and in nuclease protection assays using probes made from mouse pituitary TRH-R cDNA, in GH3 cells. These agents were studied to gain insight into the mechanism of the TRH effect, because signal transduction by TRH involves generation of inositol 1,4,5-trisphosphate and elevation of cytoplasmic free Ca2+ concentration, which leads to activation of Ca2+/calmodulin-dependent protein kinase, and of 1,2-diacylglycerol, which leads to activation of protein kinase-C. TRH (1 microM TRH, a maximally effective dose) caused a marked transient decrease in TRH-R mRNA that attained a nadir of 20-45% of control by 3-6 h, increased after 9 h, but was still below control levels after 24 h. Elevation of the cytoplasmic free Ca2+ concentration had no effect on TRH-R mRNA. A maximally effective dose of PMA (1 microM) caused decreases in TRH-R mRNA that were similar in magnitude and time course to those induced by 1 microM TRH. H-7 (20 microM) blocked the effects of TRH and PMA to lower TRH-R mRNA to similar extents.(ABSTRACT TRUNCATED AT 250 WORDS)
Fujimoto
J
J
Department of Medicine, New York Hospital, Cornell University Medical College, New York 10021.
Straub
R E
RE
Gershengorn
M C
MC
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D015342:DNA Probes; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D010879:Piperazines / Q000494:pharmacology; D010911:Pituitary Neoplasms; D011061:Poly A / Q000235:genetics; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism*; D012313:RNA / Q000235:genetics; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D017981:Receptors, Neurotransmitter / Q000235:genetics*; D018025:Receptors, Thyrotropin-Releasing Hormone; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology*
D015342:DNA Probes; D007546:Isoquinolines; D010879:Piperazines; D012333:RNA, Messenger; D017981:Receptors, Neurotransmitter; D018025:Receptors, Thyrotropin-Releasing Hormone; D011061:Poly A; D013973:Thyrotropin-Releasing Hormone; D012313:RNA; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate