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 microM) 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.