Differential solubilization of nuclear glucocorticoid receptors by DNAase I and DNAase II.
This study analyzes the sensitivity of nuclear bound glucocorticoid receptors to solubilization from nuclei by DNAase I and DNAase II. Thymocytes were incubated with 10(-8) M [3H]dexamethasone, [3H]cortisol or [3H]triamcinolone acetonide, without or with 10(-6) M unlabelled dexamethasone, for 30 min at 37 degrees C and nuclei from these cells were digested with either DNAase I and DNAase II. DNAase I for 2 h at 3 degrees C leads to solubilization of 60% of the nuclear DNA and release of 10--20% triamcinolone acetonide-receptor, 30--40% dexamethasone-receptor and 85--90% cortisol-receptor. DNAase II at the same enzymatic concentration solubilizes only 10--20% of the nuclear DNA, but releases 40--50% triamcinolone-receptor, 60--70% dexamethasone-receptor and 100% cortisol-receptor. Release of nuclear bound dexamethasone-receptor by DNAase I parallels the solubilization of DNA, reaching maximum values by 2 h at 3 degrees C, whereas maximal release by DNAase II is obtained within 45 min when DNA solubilization is not complete. When nuclei initially extracted with DNAase I are re-extracted with DNAase II, greater than 65% of the DNAase I residual dexamethasone-receptors are solubilized, whereas DNAase I is ineffective in solubilizing DNAase II residual dexamethasone-receptors. DNAase I solubilizes only 30% of the 0.4 M KCl residual dexamethasone-receptor whereas DNAase II digests over 90% of this fraction. DNAase I extracts of nuclear dexamethasone-receptor chromatograph on G-100 Sephadex as a single radioactive peak just after the void volume, whereas DNAase II extracts of nuclear dexamethasone-receptor chromatograph as two peaks of radioactivity, one which is similar to the DNAase I solubilized receptor and a second broad peak of macromolecular bound radioactivity which is smaller in size.
D000818:Animals; D002467:Cell Nucleus / Q000201:enzymology; D003850:Deoxyribonuclease I; D003851:Deoxyribonucleases / Q000378:metabolism*; D003907:Dexamethasone / Q000378:metabolism; D004706:Endodeoxyribonucleases; D004720:Endonucleases / Q000378:metabolism*; D006854:Hydrocortisone / Q000378:metabolism; D008297:Male; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000502:physiology*; D011987:Receptors, Steroid / Q000502:physiology*; D012995:Solubility; D013950:Thymus Gland / Q000166:cytology; D014222:Triamcinolone Acetonide / Q000378:metabolism
D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; D003907:Dexamethasone; D003851:Deoxyribonucleases; D004706:Endodeoxyribonucleases; D004720:Endonucleases; D003850:Deoxyribonuclease I; C019329:deoxyribonuclease II; D014222:Triamcinolone Acetonide; D006854:Hydrocortisone