Binding of T3 in rat liver nuclei.

Abstract

The binding of labeled triiodothyronine has been studied in rat liver nuclei prepared after in vivo administration of hormone. Nuclear pellets prepared from homogenates made in 0.32M sucrose + 1 mm magnesium chloride and centrifuged through 2.3M sucrose density gradients contain, 2 hr after labeling, approximately 9% of liver isotope in normal animals and 18%in surgically thyroidectomized animals. 60% of DNA is recovered in the nuclear pellet so prepared. The fraction of total liver isotope present in the purified nuclear pellets is 6%at 10 min, peaks between 2 and 3.5 hr, and declines gradually over the subsequent 24 hr as total blood, liver, and nuclear isotope rapidly decline. These data suggest relatively free exchange between nucleus and other components of the liver. The nuclear binding of receptor sites in normal and thyroidectomized animals can be saturated by addition of increasing doses of carrier unlabeled triiodothyronine. Scatchard analysis suggests the presence of high affinity binding sites with a capacity of 2.4 ng per g in liver of thyroidectomized animals. These nuclear binding sites would contain approximately 10,400 molecules of T3 per liver cell nucleus, and would be partially saturated at normal rat serum T3 levels. Nuclear labeled T3 can be displaced by unlabeled T3, unlabeled triac, and with less efficiency by L-thyroxine or D-thyroxine. The nuclear T3 can be extracted in vitro by incubation of the nuclei with albumin or other proteins. The nuclear binding proteins can be extracted from the nuclei by 0.4M KC1 and sediment as macromolecules with a sedimentation constant of less than 4S. The binding molecules appear to be protein since the binding activity is destroyed by incubation with protease. The high affinity nuclear T3 binding sites may be important in hormone action, although as yet no data linking these sites to subsequent metabolic events has been adduced. (Endocrinology95: 74, 1974)