Rapid localization of mutations in the thyroid hormone receptor-β gene by denaturing gradient gel electrophoresis in 18 families with thyroid hormone resistance

K. Takeda, Roy E Weiss, S. Refetoff

Research output: Contribution to journalArticle

68 Citations (Scopus)

Abstract

Generalized resistance to thyroid hormone (GRTH) is an inherited syndrome of reduced tissue responsiveness to thyroid hormone. Point mutations in the human thyroid hormone receptor-β (hTRβ) gene of these patients, causing single amino acid substitutions, appear be different in unrelated individuals affected by the same syndrome. To localize mutations in the hTRβ gene, GC- clamped DNA fragments from affected individuals belonging to 21 families with GRTH were generated by the polymerase chain reaction and analyzed by denaturing gradient gel electrophoresis (DGGE). Putative mutations in the hTRβ gene of 18 unrelated individuals with GRTH were identified, and their nature has been confirmed in 9 by sequencing. All were in the hormone-binding domain of the receptor, and 13 of 18 mutations were in its center (exon 7). In 3 families we were unable to identify mutations in hTRβ, suggesting the existence of mutations at other loci, possibly the hTRα gene or other proteins involved in the thyroid hormone-dependent transactivation system. Sequencing of DNA fragments negative for the presence of putative mutations by DEEG confirmed the absence of sequence differences. DGGE of amplified DNA fragments can rapidly and reliably localize the sites of mutations in the hTRβ gene of patients with GRTH. The procedure enabled mapping the regions in the hTRβ harboring mutations associated with GRTH.

Original languageEnglish (US)
Pages (from-to)712-719
Number of pages8
JournalJournal of Clinical Endocrinology and Metabolism
Volume74
Issue number4
DOIs
StatePublished - 1992
Externally publishedYes

Fingerprint

Thyroid Hormone Resistance Syndrome
Thyroid Hormone Receptors
Denaturing Gradient Gel Electrophoresis
Electrophoresis
Thyroid Hormones
Genes
Gels
Mutation
DNA
Polymerase chain reaction
Amino Acid Substitution
DNA Sequence Analysis
Point Mutation
Exons
Transcriptional Activation
Substitution reactions
Hormones
Tissue
Amino Acids
Polymerase Chain Reaction

ASJC Scopus subject areas

  • Biochemistry
  • Endocrinology, Diabetes and Metabolism

Cite this

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title = "Rapid localization of mutations in the thyroid hormone receptor-β gene by denaturing gradient gel electrophoresis in 18 families with thyroid hormone resistance",
abstract = "Generalized resistance to thyroid hormone (GRTH) is an inherited syndrome of reduced tissue responsiveness to thyroid hormone. Point mutations in the human thyroid hormone receptor-β (hTRβ) gene of these patients, causing single amino acid substitutions, appear be different in unrelated individuals affected by the same syndrome. To localize mutations in the hTRβ gene, GC- clamped DNA fragments from affected individuals belonging to 21 families with GRTH were generated by the polymerase chain reaction and analyzed by denaturing gradient gel electrophoresis (DGGE). Putative mutations in the hTRβ gene of 18 unrelated individuals with GRTH were identified, and their nature has been confirmed in 9 by sequencing. All were in the hormone-binding domain of the receptor, and 13 of 18 mutations were in its center (exon 7). In 3 families we were unable to identify mutations in hTRβ, suggesting the existence of mutations at other loci, possibly the hTRα gene or other proteins involved in the thyroid hormone-dependent transactivation system. Sequencing of DNA fragments negative for the presence of putative mutations by DEEG confirmed the absence of sequence differences. DGGE of amplified DNA fragments can rapidly and reliably localize the sites of mutations in the hTRβ gene of patients with GRTH. The procedure enabled mapping the regions in the hTRβ harboring mutations associated with GRTH.",
author = "K. Takeda and Weiss, {Roy E} and S. Refetoff",
year = "1992",
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AU - Takeda, K.

AU - Weiss, Roy E

AU - Refetoff, S.

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N2 - Generalized resistance to thyroid hormone (GRTH) is an inherited syndrome of reduced tissue responsiveness to thyroid hormone. Point mutations in the human thyroid hormone receptor-β (hTRβ) gene of these patients, causing single amino acid substitutions, appear be different in unrelated individuals affected by the same syndrome. To localize mutations in the hTRβ gene, GC- clamped DNA fragments from affected individuals belonging to 21 families with GRTH were generated by the polymerase chain reaction and analyzed by denaturing gradient gel electrophoresis (DGGE). Putative mutations in the hTRβ gene of 18 unrelated individuals with GRTH were identified, and their nature has been confirmed in 9 by sequencing. All were in the hormone-binding domain of the receptor, and 13 of 18 mutations were in its center (exon 7). In 3 families we were unable to identify mutations in hTRβ, suggesting the existence of mutations at other loci, possibly the hTRα gene or other proteins involved in the thyroid hormone-dependent transactivation system. Sequencing of DNA fragments negative for the presence of putative mutations by DEEG confirmed the absence of sequence differences. DGGE of amplified DNA fragments can rapidly and reliably localize the sites of mutations in the hTRβ gene of patients with GRTH. The procedure enabled mapping the regions in the hTRβ harboring mutations associated with GRTH.

AB - Generalized resistance to thyroid hormone (GRTH) is an inherited syndrome of reduced tissue responsiveness to thyroid hormone. Point mutations in the human thyroid hormone receptor-β (hTRβ) gene of these patients, causing single amino acid substitutions, appear be different in unrelated individuals affected by the same syndrome. To localize mutations in the hTRβ gene, GC- clamped DNA fragments from affected individuals belonging to 21 families with GRTH were generated by the polymerase chain reaction and analyzed by denaturing gradient gel electrophoresis (DGGE). Putative mutations in the hTRβ gene of 18 unrelated individuals with GRTH were identified, and their nature has been confirmed in 9 by sequencing. All were in the hormone-binding domain of the receptor, and 13 of 18 mutations were in its center (exon 7). In 3 families we were unable to identify mutations in hTRβ, suggesting the existence of mutations at other loci, possibly the hTRα gene or other proteins involved in the thyroid hormone-dependent transactivation system. Sequencing of DNA fragments negative for the presence of putative mutations by DEEG confirmed the absence of sequence differences. DGGE of amplified DNA fragments can rapidly and reliably localize the sites of mutations in the hTRβ gene of patients with GRTH. The procedure enabled mapping the regions in the hTRβ harboring mutations associated with GRTH.

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