1,25-Dihydroxyvitamin D3 receptors in parathyroid glands. Preliminary characterization of cytoplasmic and nuclear binding components

M. R. Hughes, Mark R Haussler

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Abstract

Macromolecular components which specifically bind the sterol hormone 1,25 dihydroxyvitamin D3 were identified both in vivo and in vitro in the parathyroid glands of chicks, bovine, and human parathyroid adenoma. The binding molecules from D deficient chicks were isolated and characterized in vitro by sucrose gradient centrifugation, enzymatic digestion, agarose gel filtration, and DEAE cellulose chromatography. The interaction of 1,25 dihydroxy[3H]vitamin D3 with the cytoplasmic binding component occurs at low concentration of hormone (< 10-8 M) with high affinity (K(d) = 4.2 x 10-9 M in 0.3 M KCl at 0°), and can be reversed by mild heating or by excess unlabeled 1,25 dihydroxyvitamin D3. The participation of protein in the sterol binding site is inferred from the destruction of the complex by pronase but not ribo or deoxyribonucleases. When the cytoplasmic hormone macromolecular complex is formed by administration of 1,25 dihydroxy[3H]vitamin D3 to chicks in vivo or by incubation of parathyroid cytosol with tritiated hormone in vitro, it exhibits a sedimentation coefficient of 3.1 to 3.7 S in 0.3 M KCl sucrose gradients. Agarose gel filtration of the cytoplasmic component indicates a Stokes molecular radius of 29 Å and an apparent molecular weight of 37,000. In vitro subcellular distribution of 1,25 dihydroxy[3H]vitamin D3 demonstrates a time and temperature dependent movement of hormone from the cytoplasm to the nucleus. This subcellular migration of the hormone corresponds with the predominant nuclear localization of the sterol which is observed in parathyroid glands after administration of 1,25 dihydroxy[3H]vitamin D3 to reveals in vivo. Analysis of hormone bound to nuclei eveals that it is associated with the chromatin fraction via a 3.1 to 3.7 S macromolecule indistinguishable from the cytosol receptor for 1,25 dihydroxyvitamin D3. The cytoplasmic/nuclear receptor for this hormone is not detected in chick adrenal, testis, liver, or muscle, although a similar binding protein for 1,25 dihydroxyvitamin D3 has been found previously in chick intestine. A functional role of these receptors in the parathyroid gland is unknown but their presence at this endocrine site is consistent with the sterol's possible involvement in the regulation of parathyroid hormone synthesis and secretion.

Original languageEnglish (US)
Pages (from-to)1065-1073
Number of pages9
JournalJournal of Biological Chemistry
Volume253
Issue number4
StatePublished - 1978

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Calcitriol Receptors
Parathyroid Glands
Hormones
Cholecalciferol
Sterols
Calcitriol
Cytosol
Sepharose
Gel Chromatography
Sucrose
Gels
Macromolecular Substances
DEAE-Cellulose Chromatography
DEAE-Cellulose
Parathyroid Neoplasms
Pronase
Deoxyribonucleases
Centrifugation
Cytoplasmic and Nuclear Receptors
Chromatography

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "1,25-Dihydroxyvitamin D3 receptors in parathyroid glands. Preliminary characterization of cytoplasmic and nuclear binding components",
abstract = "Macromolecular components which specifically bind the sterol hormone 1,25 dihydroxyvitamin D3 were identified both in vivo and in vitro in the parathyroid glands of chicks, bovine, and human parathyroid adenoma. The binding molecules from D deficient chicks were isolated and characterized in vitro by sucrose gradient centrifugation, enzymatic digestion, agarose gel filtration, and DEAE cellulose chromatography. The interaction of 1,25 dihydroxy[3H]vitamin D3 with the cytoplasmic binding component occurs at low concentration of hormone (< 10-8 M) with high affinity (K(d) = 4.2 x 10-9 M in 0.3 M KCl at 0°), and can be reversed by mild heating or by excess unlabeled 1,25 dihydroxyvitamin D3. The participation of protein in the sterol binding site is inferred from the destruction of the complex by pronase but not ribo or deoxyribonucleases. When the cytoplasmic hormone macromolecular complex is formed by administration of 1,25 dihydroxy[3H]vitamin D3 to chicks in vivo or by incubation of parathyroid cytosol with tritiated hormone in vitro, it exhibits a sedimentation coefficient of 3.1 to 3.7 S in 0.3 M KCl sucrose gradients. Agarose gel filtration of the cytoplasmic component indicates a Stokes molecular radius of 29 {\AA} and an apparent molecular weight of 37,000. In vitro subcellular distribution of 1,25 dihydroxy[3H]vitamin D3 demonstrates a time and temperature dependent movement of hormone from the cytoplasm to the nucleus. This subcellular migration of the hormone corresponds with the predominant nuclear localization of the sterol which is observed in parathyroid glands after administration of 1,25 dihydroxy[3H]vitamin D3 to reveals in vivo. Analysis of hormone bound to nuclei eveals that it is associated with the chromatin fraction via a 3.1 to 3.7 S macromolecule indistinguishable from the cytosol receptor for 1,25 dihydroxyvitamin D3. The cytoplasmic/nuclear receptor for this hormone is not detected in chick adrenal, testis, liver, or muscle, although a similar binding protein for 1,25 dihydroxyvitamin D3 has been found previously in chick intestine. A functional role of these receptors in the parathyroid gland is unknown but their presence at this endocrine site is consistent with the sterol's possible involvement in the regulation of parathyroid hormone synthesis and secretion.",
author = "Hughes, {M. R.} and Haussler, {Mark R}",
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T1 - 1,25-Dihydroxyvitamin D3 receptors in parathyroid glands. Preliminary characterization of cytoplasmic and nuclear binding components

AU - Hughes, M. R.

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N2 - Macromolecular components which specifically bind the sterol hormone 1,25 dihydroxyvitamin D3 were identified both in vivo and in vitro in the parathyroid glands of chicks, bovine, and human parathyroid adenoma. The binding molecules from D deficient chicks were isolated and characterized in vitro by sucrose gradient centrifugation, enzymatic digestion, agarose gel filtration, and DEAE cellulose chromatography. The interaction of 1,25 dihydroxy[3H]vitamin D3 with the cytoplasmic binding component occurs at low concentration of hormone (< 10-8 M) with high affinity (K(d) = 4.2 x 10-9 M in 0.3 M KCl at 0°), and can be reversed by mild heating or by excess unlabeled 1,25 dihydroxyvitamin D3. The participation of protein in the sterol binding site is inferred from the destruction of the complex by pronase but not ribo or deoxyribonucleases. When the cytoplasmic hormone macromolecular complex is formed by administration of 1,25 dihydroxy[3H]vitamin D3 to chicks in vivo or by incubation of parathyroid cytosol with tritiated hormone in vitro, it exhibits a sedimentation coefficient of 3.1 to 3.7 S in 0.3 M KCl sucrose gradients. Agarose gel filtration of the cytoplasmic component indicates a Stokes molecular radius of 29 Å and an apparent molecular weight of 37,000. In vitro subcellular distribution of 1,25 dihydroxy[3H]vitamin D3 demonstrates a time and temperature dependent movement of hormone from the cytoplasm to the nucleus. This subcellular migration of the hormone corresponds with the predominant nuclear localization of the sterol which is observed in parathyroid glands after administration of 1,25 dihydroxy[3H]vitamin D3 to reveals in vivo. Analysis of hormone bound to nuclei eveals that it is associated with the chromatin fraction via a 3.1 to 3.7 S macromolecule indistinguishable from the cytosol receptor for 1,25 dihydroxyvitamin D3. The cytoplasmic/nuclear receptor for this hormone is not detected in chick adrenal, testis, liver, or muscle, although a similar binding protein for 1,25 dihydroxyvitamin D3 has been found previously in chick intestine. A functional role of these receptors in the parathyroid gland is unknown but their presence at this endocrine site is consistent with the sterol's possible involvement in the regulation of parathyroid hormone synthesis and secretion.

AB - Macromolecular components which specifically bind the sterol hormone 1,25 dihydroxyvitamin D3 were identified both in vivo and in vitro in the parathyroid glands of chicks, bovine, and human parathyroid adenoma. The binding molecules from D deficient chicks were isolated and characterized in vitro by sucrose gradient centrifugation, enzymatic digestion, agarose gel filtration, and DEAE cellulose chromatography. The interaction of 1,25 dihydroxy[3H]vitamin D3 with the cytoplasmic binding component occurs at low concentration of hormone (< 10-8 M) with high affinity (K(d) = 4.2 x 10-9 M in 0.3 M KCl at 0°), and can be reversed by mild heating or by excess unlabeled 1,25 dihydroxyvitamin D3. The participation of protein in the sterol binding site is inferred from the destruction of the complex by pronase but not ribo or deoxyribonucleases. When the cytoplasmic hormone macromolecular complex is formed by administration of 1,25 dihydroxy[3H]vitamin D3 to chicks in vivo or by incubation of parathyroid cytosol with tritiated hormone in vitro, it exhibits a sedimentation coefficient of 3.1 to 3.7 S in 0.3 M KCl sucrose gradients. Agarose gel filtration of the cytoplasmic component indicates a Stokes molecular radius of 29 Å and an apparent molecular weight of 37,000. In vitro subcellular distribution of 1,25 dihydroxy[3H]vitamin D3 demonstrates a time and temperature dependent movement of hormone from the cytoplasm to the nucleus. This subcellular migration of the hormone corresponds with the predominant nuclear localization of the sterol which is observed in parathyroid glands after administration of 1,25 dihydroxy[3H]vitamin D3 to reveals in vivo. Analysis of hormone bound to nuclei eveals that it is associated with the chromatin fraction via a 3.1 to 3.7 S macromolecule indistinguishable from the cytosol receptor for 1,25 dihydroxyvitamin D3. The cytoplasmic/nuclear receptor for this hormone is not detected in chick adrenal, testis, liver, or muscle, although a similar binding protein for 1,25 dihydroxyvitamin D3 has been found previously in chick intestine. A functional role of these receptors in the parathyroid gland is unknown but their presence at this endocrine site is consistent with the sterol's possible involvement in the regulation of parathyroid hormone synthesis and secretion.

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