title: The heat-stable cytosolic factor that promotes glucocorticoid receptor binding to DNA is neither thioredoxin nor ribonuclease.
abstract: Treatment of rat liver cytosol containing temperature-transformed [3H]dexamethasone-bound receptors at 0 degree C with the sulfhydryl modifying reagent methyl methanethiosulfonate (MMTS) inhibits the DNA-binding activity of the receptor, and DNA-binding activity is restored after addition of dithiothreitol (DTT). However, transformed receptors that are treated with MMTS and then separated from low Mr components of cytosol by passage through a column of Sephadex G-50 have very little DNA-binding activity when DTT is added to regenerate sulfhydryl moities. The receptors will bind to DNA if whole liver cytosol or boiled liver cytosol is added in addition to DTT. The effect of boiled cytosol is mimicked by purified rat thioredoxin or bovine RNase A in a manner that does not reflect the reducing activity of the former or the catalytic activity of the latter. This suggests that the reported ability of each of these heat-stable peptides to stimulate DNA binding by glucocorticoid receptors is not a biologically relevant action. We suggest that stimulation of DNA binding of partially purified receptors by boiled cytosol does not constitute a reconstitution of a complete cytosolic system in which the dissociated receptor must associate with a specific heat-stable accessory protein required for DNA binding, as has been suggested in the "two-step" model of receptor transformation recently proposed by Schmidt et al. (Schmidt T.J., Miller-Diener, A., Webb M.L. and Litwack G. (1985) J. biol. Chem. 260, 16255-16262).
authors: Department of Pharmacology, University of Michigan Medical School, Ann Arbor 48109-0010.
authors: W
authors: W
authors: Tienrungroj
authors: S E
authors: SE
authors: Pratt
authors: J F
authors: JF
authors: Grippo
authors: A
authors: A
authors: Holmgren
authors: W B
authors: WB
authors: Pratt
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000502:physiology*; D003600:Cytosol / Q000502:physiology*; D004247:DNA / Q000378:metabolism*; D003907:Dexamethasone / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D006358:Hot Temperature; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008741:Methyl Methanesulfonate / Q000031:analogs & derivatives / Q000494:pharmacology; D010455:Peptides / Q000502:physiology*; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D012260:Ribonucleases / Q000502:physiology*; D013879:Thioredoxins / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D010455:Peptides; D011965:Receptors, Glucocorticoid; C014674:methyl methanethiosulfonate; D013879:Thioredoxins; D003907:Dexamethasone; D004247:DNA; D008741:Methyl Methanesulfonate; D012260:Ribonucleases; D004229:Dithiothreitol