Binding characteristics of L-triiodothyronine to isolated rat liver chromatin.
The binding characteristics of [125I]-labeled L-triiodothyronine (T3) to chromatin isolated from rat liver nuclei were investigated. Binding of T3 to chromatin showed temperature-, incubation time-, and DNA concentration-dependence. According to Scatchard analysis, the apparent equilibrium dissociation constant was 225 pM, with a maximum binding capacity of about 0.2 pmoles per mg DNA. Displacement studies with unlabeled thyroxine (T4) and T3 showed that the binding sites for T4 might be the same as T3 but the binding affinity of the former was less than that of the latter. The binding was completely inhibited by the eukaryotic RNA polymerase inhibitor, rifampicin AF/021, but not by the prokaryotic inhibitor, rifampicin and alpha-amanitin. These observations indicate that the receptors for T3 have certain properties in common with RNA polymerase or other enzyme proteins which are sensitive to the rifampicin derivative. The hormone--chromatin fragments complex was solubilized from residual chromatin by digestion with DNase, but not with RNase, suggesting that the T3 receptors localize in the DNase-sensitive regions of DNA in the chromatin. This provides a useful method to use to investigate the localization of the receptor proteins in the chromatin and the interaction of the hormone-receptor complex with DNA.
D000546:Amanitins / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D002843:Chromatin / Q000187:drug effects / Q000378:metabolism*; D003851:Deoxyribonucleases; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D009994:Osmolar Concentration; D051381:Rats; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism*; D012260:Ribonucleases; D012293:Rifampin / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D013974:Thyroxine / Q000378:metabolism; D014284:Triiodothyronine / Q000378:metabolism*
D000546:Amanitins; D002843:Chromatin; D011956:Receptors, Cell Surface; D014284:Triiodothyronine; D012965:Sodium Chloride; D003851:Deoxyribonucleases; D012260:Ribonucleases; D013974:Thyroxine; D012293:Rifampin