ATP-induced activation of purified rat hepatic glucocorticoid receptors.
We have utilized unactivated rat hepatic glucocorticoid receptor complexes purified to near homogeneity by a three-step scheme which includes affinity chromatography, gel filtration and anion exchange chromatography, to demonstrate for the first time that ATP can interact directly with the receptor protein in stimulating activation. This stimulation is reflected by an increase in DNA-cellulose binding as well as by a shift in the elution profile of the purified receptor complexes from DEAE-cellulose. A concentration of 10 mM Na2MoO4 is able to block both of these effects. ATP stimulates activation in a dose-dependent manner (maximally at 10 mM), and elicits maximal activation within 30 min at 15 degrees C. There appears to be no nucleotide specificity since GTP, CTP and UTP, as well as ADP and GDP also stimulate activation. All of these observations closely parallel data obtained from similar activation experiments performed with crude rat hepatic receptors. ATP does not appear to stimulate activation of receptors (crude or purified) by initiating a phosphorylation reaction since hydrolysis-resistant analogues of ATP are also effective. Pyrophosphate (PPi) is as effective as ATP in promoting receptor activation, since it elicits similar increases in DNA-cellulose binding, shifts in elution patterns from DEAE-cellulose, and dose-response relationships. None of the compounds tested stimulate activation indirectly by pH or ionic strength effects. Despite the fact that high ATP concentrations (3-4-fold higher than those present in vivo) are necessary to stimulate maximal activation, a physiological role of ATP in directly regulating in vivo activation of glucocorticoid receptors cannot be ruled out.
Diehl
E E
EE
Department of Physiology and Biophysics, College of Medicine, University of Iowa, Iowa City 52242.
Schmidt
T J
TJ
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology*; D000315:Adrenalectomy; D000818:Animals; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D009994:Osmolar Concentration; D051381:Rats; D011919:Rats, Inbred Strains; D011965:Receptors, Glucocorticoid / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D014222:Triamcinolone Acetonide / Q000378:metabolism
D011965:Receptors, Glucocorticoid; D000255:Adenosine Triphosphate; D014222:Triamcinolone Acetonide