JBC, Vol. 251, Issue 23, 7380-7387, Dec, 1976
Chromatin receptors for thyroid hormones. Interactions of the solubilized proteins with DNA
K. M. MacLeod and J. D. Baxter
Thyroid hormone-responsive tissues contain chromatin "receptor" proteins
that are concentrated in chromatin subfractions enriched in DNA. These
receptors appear to be DNA-binding proteins. In the present study, we
utilized a DNA-cellulose binding assay to further examine the interactions
of solubilized receptors with DNA. [125I]Triiodothyronine associates with
receptors bound to DNA-cellulose, whereas free [I]triiodothyronine and
[125I]triiodothyronine associated with other proteins does not. The
DNA-receptor interactions appear to be strong enough to exist at
physiological ionic strength since binding is 50% maximal ag 0.175 M NaCl
and is only partly inhibited by Ca2+ and Mg2+ in the 1 to 5 mM range. Most,
if not all, of the receptors are capable of DNA binding, and there are at
least 80,000 receptor binding sites/diploid DNA (assuming one
triiodothyronine binding site/receptor). Binding of the
receptor-[125I]triiodothyronine complexes to other DNAs and analogs was
examined using a competition assay. There is similar binding by native and
denatured DNA, gy eukaryotic DNA from different species and by prokaryotic
DNA (Bacillus subtilis). Binding by natural DNAs is more avid than by
cytoplasmic RNA, nuclear RNA, poly(dA-dT)-poly(dA-dT), or
poly(dG-dC)-poly(dG-dC). Under these conditions, binding by tRNA and
poly(dA) is insignificant, and the nucleotide monomers ATP and GTP have no
detectable binding. These studies support the idea that the thyroid hormone
receptor is a DNA-binding protein and that the interaction is a major
determinant for receptor localization in chromatin. The competition studies
suggest that the polynucleotide composition and/or conformation can have
marked influences on the binding, and that multiple orders of binding
affinity can exist. The presence of specific sequences cannot be excluded.
However, the finding that receptors bind extensively and tightly to DNA
suggests that receptors in chromatin may randomly bind to any available
DNA, resulting in some of the receptors being at physiologically
unimportant sites. If so, the several thousand hormone receptors present in
each target cell may be required to enhance the possibility that some of
the receptors are present at the actual sites of action.
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