Extraction of DNA-cellulose-bound glucocorticoid-receptor complexes with sodium tungstate.
Glucocorticoid-receptor complex from rat liver cytosol, activated by warming at 23 degrees C or fractionation with (NH4)2SO4, was adsorbed over DNA-cellulose. This DNA-cellulose-bound [3H]triamcinolone acetonide-receptor complex was extracted in a dose-dependent manner by incubation with different concentrations of sodium tungstate. A 50% recovery of receptor was achieved with 5 mM sodium tungstate. Almost the entire glucocorticoid-receptor complex bound to DNA-cellulose could be extracted with 20 mM sodium tungstate. The [3H]triamcinolone acetonide released from DNA-cellulose following tungstate and molybdate treatment was found to be associated with a macromolecule, as seen by analysis on a Sephadex G-75 column. The glucocorticoid-receptor complex extracted by both the compounds sedimented as a 4 S entity of 5-20% sucrose gradients under low- and high-salt conditions. Addition of tungstate or molybdate to the preparations containing activated receptor had no effect on the sedimentation rate of receptor. However, addition of tungstate to non-activated receptor preparation caused aggregates of larger size. The tungstate-extracted glucocorticoid-receptor complex failed to rebind to DNA-cellulose even after extensive dialysis, whereas receptor in molybdate-extract retained its DNA-cellulose binding capacity.
D000818:Animals; D002482:Cellulose / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D004247:DNA / Q000378:metabolism*; D008099:Liver / Q000032:analysis*; D008297:Male; D008982:Molybdenum / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000302:isolation & purification*; D011987:Receptors, Steroid / Q000302:isolation & purification*; D014222:Triamcinolone Acetonide / Q000378:metabolism; D014414:Tungsten; D017973:Tungsten Compounds
D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; D017973:Tungsten Compounds; C025399:sodium tungstate(VI); D011189:Potassium Chloride; D008982:Molybdenum; D002482:Cellulose; D004247:DNA; C024687:sodium molybdate(VI); D014222:Triamcinolone Acetonide; D014414:Tungsten