Differential effects of agonists and antagonists on autoregulation of glucocorticoid receptors in a rat colonic adenocarcinoma cell line

Abstract The relative abilities of a potent glucocorticoid receptor (GR) agonist (RU 28362), a weak GR agonist (aldosterone) and a potent GR antagonist (RU 38486) to promote in vivo activation/transformation and subsequent down-regulation of GR mRNA and protein levels were compared using the PROb rat colonic adenocarcinoma cell line. In vivo activation, which is followed immediately by nuclear translocation, by these ligands (1 μM) was evaluated in terms of their abilities to deplete cytoplasmic GR protein levels after a 30 min incubation period. Western blotting experiments with the anti-GR monoclonal antibody BuGR2 demonstrated that a brief incubation with RU 28362 resulted in nearly complete depletion of cytoplasmic GR, whereas incubation with aldosterone resulted in a 50% depletion of the cytoplasmic GR protein. Incubation with RU 38486 was even more effective than aldosterone in promoting this key step in the GR pathway. Prolonged treatment (18 h) with RU 28362 resulted in significant down-regulation of GR mRNA and total cellular GR protein levels. Similar incubation with aldosterone resulted in a transient decrease in the GR mRNA level and also down-regulated the total GR protein level. Although prolonged incubation with RU 38486 did not result in detectable down-regulation of the GR mRNA level, this antagonist very effectively down-regulated total cellular GR protein levels. Taken collectively, these data demonstrate that agonists capable of promoting in vivo activation (and subsequent nuclear translocation) of GR are also effective at down-regulating GR at both the mRNA and protein levels. Although the antagonist RU 38486 is also capable of down-regulating GR protein levels by shortening the half-life of the receptor, it appears to be incapable of altering the rate of transcription of the GR gene. Glucocorticoid target tissue sensitivity may thus be decreased via two independent mechanisms: agonist-induced repression of GR gene transcription; and/or ligand-induced degradation of total cellular GR protein levels.

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