Effect of macromolecular-translocation inhibitor-III on binding of activated glucocorticoid-receptor complex to specific DNA.
We previously purified macromolecular-translocation inhibitor-III (MTI-III), which inhibits the binding of the activated glucocorticoid-receptor complex (GR) to nuclei, to homogeneity from rat liver, and we found that the purified MTI-III bound to partially purified activated GR under low salt conditions at slightly acidic pH [Liu, G., Okamoto, K., and Isohashi, F. (1993) Eur. J. Biochem. 218, 679-687]. This was the first direct evidence that the inhibitor acts through a direct interaction with the activated GR. In this study, we examined whether the purified MTI-III could interfere with the binding of GR to a DNA fragment containing a specific glucocorticoid-response element (GRE). Under nearly isotonic salt conditions at neutral pH, the activated GR bound to the GRE but not to nonspecific DNA. Under similar conditions, the activated GR also bound to the purified MTI-III. The resulting GR/MTI-III complex did not bind to the GRE. We also found that addition of MTI-III to the GR/GRE complex resulted in time-dependent disruption of the GR/GRE complex and formation of the GR/MTI-III complex. The half-life of the GR/GRE complex in the presence of MTI-III was about 13 min. These results suggest that MTI-III enhances the release of GR from the GR/GRE complex and immediately forms a stable GR/MTI-III complex.
Okamoto
K
K
Department of Biochemistry, St Marianna University School of Medicine, Kanagawa.
Liu
G
G
Yu
W G
WG
Ochiai
T
T
Isohashi
F
F
D000818:Animals; D001483:Base Sequence; D002457:Cell Extracts; D004247:DNA / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008099:Liver / Q000378:metabolism; D046911:Macromolecular Substances; D008297:Male; D008969:Molecular Sequence Data; D011485:Protein Binding; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid; D014222:Triamcinolone Acetonide / Q000378:metabolism
D002457:Cell Extracts; D046911:Macromolecular Substances; D011965:Receptors, Glucocorticoid; C027048:receptor macromolecular translocation inhibitor; D004247:DNA; D014222:Triamcinolone Acetonide