Particulate untransformed glucocorticoid-receptor complexes from HeLa cells crosslinked in vivo.
When control HeLa cells were incubated at 2 degrees C in the presence of tritiated dexamethasone, most glucocorticoid-receptor complexes were found in cytosolic extracts as untransformed forms. Chemical crosslinking of intact HeLa cells resulted in the immobilization of 50% of the total cellular glucocorticoid-receptor complexes in the nuclear fraction. Under these conditions the redistribution of total protein, RNA and lactate dehydrogenase activity between cytosol and nuclei was negligible, indicating that glucocorticoid binding in the nuclear fraction was not due to a methodological artifact. High levels of glucocorticoid receptor were also found in the nuclear fraction of crosslinked cells which were not exposed to glucocorticoids. Nuclear receptor complexes could be released in soluble forms by DNase I and sonication. Evaluations of DNA binding and ionic properties of glucocorticoid receptors prepared from control and crosslinked cells maintained at 2 degrees C revealed that most of the hormone-receptor complex in cytosols and nuclear extracts behaved as untransformed forms. As opposed to glucocorticoid receptors prepared from control cells, heat treatment of extracts obtained from crosslinked cells did not result in increased DNA binding and changes in ionic properties of receptor complexes. I conclude that untransformed glucocorticoid receptors are present in both cytosol and nuclei of intact cells.
Rossini
G P
GP
Istituto di Chimica Biologica, Università di Modena, Italy.
D002467:Cell Nucleus / Q000032:analysis*; D002848:Chromatography, DEAE-Cellulose; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003600:Cytosol / Q000032:analysis*; D004247:DNA / Q000378:metabolism; D005557:Formaldehyde / Q000494:pharmacology; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007477:Ions; D011965:Receptors, Glucocorticoid / Q000187:drug effects / Q000378:metabolism*; D013347:Subcellular Fractions / Q000032:analysis
D003432:Cross-Linking Reagents; D007477:Ions; D011965:Receptors, Glucocorticoid; D005557:Formaldehyde; D004247:DNA