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Characterization of a thyroid-hormone-binding site on nuclear envelopes and nuclear matrices of the male-rat liver
Abstract:
Nuclear envelopes and nuclear matrices were isolated from the male-rat liver. Incubation of 125I-labelled 3,3',5-tri-iodothyronine (T3) with the nuclear-envelope fraction resulted in specific binding of T3 to the membranes. Maximum specific binding occurred at 30 degrees C after 2h incubation. Storage for 1 week at -80 degrees C resulted in no loss of binding. Scatchard analysis revealed a class of binding sites with KD 86 nM. 3,3',5'-Tri-iodothyronine was as effective a competitor of [125I]T3 binding to nuclear envelopes as was L-T3 itself, and tri-iodothyroacetic acid was 70% as potent as T3. L- and D-thyronine did not compete for [125I]T3 binding. Incubation of nuclear envelopes with 0.6 M-NaCl before addition of T3 resulted in the complete loss of specific binding sites, whereas exposure of the membranes to 2.0 M-NaCl after incubation with T3 did not extract binding sites. Nuclear matrices, after incubation with [125I]T3 under the same conditions, were shown to possess a class of binding sites with a similar KD but with approx. 30% of the maximum binding capacity. Nuclear envelopes from hypothyroid animals may possess slightly lower numbers of binding sites compared with nuclear envelopes from the intact animal, whereas nuclear matrices from hypothyroid animals have the same number of binding sites as do nuclear envelopes from the intact animal. In conclusion, nuclear envelopes and nuclear matrices have a class of binding sites with relatively high affinity for T3. It is distinct from nuclear and cytosolic binding sites.
Insights
Male rat liver nuclear envelopes and matrices bind triiodothyronine (T3) with high affinity. These specific T3 binding sites are located within the nuclear envelope and matrix, distinct from other known cellular binding sites.
Area of Science:
- Endocrinology
- Molecular Biology
- Cell Biology
Background:
- Thyroid hormones, such as triiodothyronine (T3), play crucial roles in regulating cellular metabolism and gene expression.
- The precise mechanisms of T3 transport and binding within the cell nucleus are still under investigation.
- Nuclear envelopes and matrices are key structural components of the nucleus involved in various cellular processes.
Purpose of the Study:
- To investigate the presence and characteristics of T3 binding sites within isolated rat liver nuclear envelopes and nuclear matrices.
- To determine the affinity and specificity of these binding sites for T3 and related compounds.
- To compare the binding properties of T3 in nuclear envelopes and matrices, and in animals with different thyroid statuses.
Main Methods:
- Isolation of nuclear envelopes and nuclear matrices from male rat liver.
- Incubation of isolated fractions with 125I-labelled T3.
- Scatchard analysis to characterize binding sites.
- Competition assays using various thyroid hormone analogues.
- Treatment of fractions with different salt concentrations to assess binding site stability.
Main Results:
- Specific binding of T3 to nuclear envelope membranes was observed, with maximum binding at 30°C after 2h.
- Scatchard analysis revealed a class of binding sites with a dissociation constant (KD) of 86 nM in nuclear envelopes.
- Nuclear matrices also exhibited T3 binding sites with similar KD but lower capacity (approx. 30%).
- 3,3',5'-Triiodothyronine and triiodothyroacetic acid competed effectively for T3 binding, while thyronine isomers did not.
- High salt concentrations (0.6 M NaCl) removed binding sites from nuclear envelopes, indicating their proteinaceous nature.
Conclusions:
- Nuclear envelopes and nuclear matrices possess a distinct class of high-affinity T3 binding sites.
- These binding sites are different from previously identified nuclear and cytosolic T3 receptors.
- The findings suggest a potential role for nuclear envelope and matrix-associated binding sites in thyroid hormone action.
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