Potential mechanisms underlying autoregulation of glucocorticoid receptor mRNA levels in the DHD/K12/PROb rat colonic adenocarcinoma cell line.
The DHD/K12/PROb rat colonic epithelial cell line, which was originally derived from a chemically induced adenocarcinoma, expresses functional glucocorticoid receptors (GR) and has been reported to be growth inhibited by glucocorticoid agonists. In the present study the potential mechanisms underlying corticosteroid-mediated autoregulation of GR mRNA levels in this colonic cell line were investigated. The GR mRNA levels in the various treatment groups were quantitated via the ribonuclease protection assay using a specific 32P-cRNA probe. Time-course experiments demonstrated that in contrast to several other cell lines that are also growth inhibited by glucocorticoids, treatment of confluent monolayers of PROb cells with the pure GR agonist RU 28362 (1 microM) elicits a rapid and significant (65%) down-regulation of GR mRNA levels that is sustained for at least 36 h. This down-regulation, which is also elicited to a lesser extent by weaker GR agonists including corticosterone and aldosterone, is blocked by the GR antagonist RU 38486. The protein synthesis inhibitor cycloheximide was utilized to demonstrate that the initial phase (6 h) of agonist-mediated down-regulation occurs independently of ongoing protein synthesis, although new protein synthesis, perhaps of the GR protein itself, is required to maintain this down-regulation. Although agonist-mediated down-regulation in these cells probably occurs primarily at the level of GR gene transcription, inhibition of ongoing RNA synthesis with actinomycin D or 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) demonstrated that during the initial phase (1 h) of this down-regulation, but not following maximal (18 h) down-regulation, RU 28362 treatment also significantly reduces the stability of the GR mRNA.
D000230:Adenocarcinoma; D000732:Androstanols / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003110:Colonic Neoplasms; D046508:Culture Techniques / Q000379:methods; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D004848:Epithelium; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006207:Half-Life; D006706:Homeostasis; D007700:Kinetics; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis*; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000096:biosynthesis*; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000732:Androstanols; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D003609:Dactinomycin; C049154:11,17-dihydroxy-6-methyl-17-(1-propynyl)androsta-1,4,6-triene-3-one; D003513:Cycloheximide