Endocrinology, Vol 100, 648-655, Copyright © 1977 by Endocrine Society
Thyroid hormone receptors: release of receptor to the medium during in vitro incubation of isolated rat liver nuclei
J Bernal and LJ Degroot
Department of Medicine, University of Chicago, Illinois.
Isolated rat liver nuclei show a substantial amount of T3 receptor release
to the medium during in vitro incubation. This has been shown to be a
general feature of nuclei compared after several methods of isolation and
incubation. About 50% of nuclear receptors are released to the medium when
incubated in sucrose-MgCl2-Tris, pH 7.85, for 2 h at 20 C.DNA, histones,
and non-histone proteins (NHP) are also released. CaCl2 inhibits about 90%
of the release of DNA and histones, but has less effect on inhibiting
leakage of NHP and nuclear T3-binding protein (NTBP). The highest leakage
for each fraction was found when incubating nuclei in the presence of EDTA.
The receptor released to the medium has an affinity virtually identical to
the receptor remaining in the nuclei. At least 1 mM dithiothreitol is
needed to avoid degradation of the receptor. The NTBP has a sedimentation
constant of 4.5 S when studied in low ionic strength gradients. Increasing
KCl concentration decreases progressively its sedimentation constant, and
in gradients containing 0.4 M KCl the receptor sediments as a single peak
of 3.4 S. Since release of receptor to incubation medium decreases free T3
concentration, it must be taken into account in calculating receptor
affinity. Total nuclear capacity in vitro is obviously underestimated,
unless receptor released to medium is measured. Receptor exchange between
cytosol and nucleus may be of physiologic significance.
