Human thyroxine-binding globulin gene: Complete sequence and transcriptional regulation

Yoshitaka Hayashi, Yuichi Mori, Onno E. Janssen, Thongkum Sunthornthepvarakul, Roy E Weiss, Kyoko Takeda, Michael Weinberg, Hisao Seo, Graeme I. Bell, Samuel Refetoff

Research output: Contribution to journalArticle

56 Citations (Scopus)

Abstract

T4-binding globulin (TBG) is a glycoprotein of hepatic origin which transports thyroid hormone in serum. To characterize the human TBG (hTBG) gene, we studied its genomic organization, promoter activity, and regulation. To this purpose, we isolated from liver a complete hTBG cDNA clone containing the 5′-untranslated region and localized the transcription start site (TSS). The analysis of genomic clones revealed that the hTBG gene consists of five exons and that its exon-intron organization is similar to that of other members of the serine protease inhibitor family. The first exon (exon 0) is a short noncoding sequence located 1.62 kilobase pairs (kbp) upstream from exon 1. Potential cis-acting transcriptional regulatory elements including a TATA box, a CAAT box, and a hepatocyte nuclear factor-1 binding motif were identified in the upstream region. A reporter gene in which 3.2 kbp of the 5′-flanking region, including exon 0, was inserted upstream of the bacterial chloramphenicol acetyltransferase gene showed significant activity when transfected into a hepatoblastoma-derived (HepG2) cell line. The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate, down-regulated the promoter activity by more than 80% and completely inhibited hTBG synthesis, whereas thyroid hormone, glucocorticoid, estrogen, and nicotinic acid had little, if any, effect. A series of 5′-deletions revealed that the fragment -218 to +4 from the TSS had the highest promoter activity, nearly 1000-fold greater than the promoterless chloramphenicol acetyltransferase construct. When nonhepatocyte-derived cell lines (CV-1 and CHO) were tested, promoter activity was reduced by a factor of 100, showing that the promoter works in liver-specific manner. The region -218 to -102 contains liver-specific enhancer elements, since deletion to nucleotide -101 resulted in a profound re-duction of the promoter activity in HepG2 cells but not in CV-1 or CHO cells. On the other hand, mutational disruption of the putative hepatocyte nuclear factor-1 site (located 65 bp upstream of the TSS) completely abolished the promoter activity in all cell lines, indicating that this site is absolutely required for the transcription of the hTBG gene.

Original languageEnglish (US)
Pages (from-to)1049-1060
Number of pages12
JournalMolecular Endocrinology
Volume7
Issue number8
StatePublished - 1993
Externally publishedYes

Fingerprint

Exons
Transcription Initiation Site
Hepatocyte Nuclear Factor 1
Genes
Chloramphenicol O-Acetyltransferase
Liver
Globulins
Hep G2 Cells
Thyroid Hormones
Cell Line
Clone Cells
Transcriptional Regulatory Elements
Hepatoblastoma
TATA Box
Serine Proteinase Inhibitors
CHO Cells
5' Flanking Region
Niacin
5' Untranslated Regions
Phorbol Esters

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology, Diabetes and Metabolism

Cite this

Hayashi, Y., Mori, Y., Janssen, O. E., Sunthornthepvarakul, T., Weiss, R. E., Takeda, K., ... Refetoff, S. (1993). Human thyroxine-binding globulin gene: Complete sequence and transcriptional regulation. Molecular Endocrinology, 7(8), 1049-1060.

Human thyroxine-binding globulin gene : Complete sequence and transcriptional regulation. / Hayashi, Yoshitaka; Mori, Yuichi; Janssen, Onno E.; Sunthornthepvarakul, Thongkum; Weiss, Roy E; Takeda, Kyoko; Weinberg, Michael; Seo, Hisao; Bell, Graeme I.; Refetoff, Samuel.

In: Molecular Endocrinology, Vol. 7, No. 8, 1993, p. 1049-1060.

Research output: Contribution to journalArticle

Hayashi, Y, Mori, Y, Janssen, OE, Sunthornthepvarakul, T, Weiss, RE, Takeda, K, Weinberg, M, Seo, H, Bell, GI & Refetoff, S 1993, 'Human thyroxine-binding globulin gene: Complete sequence and transcriptional regulation', Molecular Endocrinology, vol. 7, no. 8, pp. 1049-1060.
Hayashi Y, Mori Y, Janssen OE, Sunthornthepvarakul T, Weiss RE, Takeda K et al. Human thyroxine-binding globulin gene: Complete sequence and transcriptional regulation. Molecular Endocrinology. 1993;7(8):1049-1060.
Hayashi, Yoshitaka ; Mori, Yuichi ; Janssen, Onno E. ; Sunthornthepvarakul, Thongkum ; Weiss, Roy E ; Takeda, Kyoko ; Weinberg, Michael ; Seo, Hisao ; Bell, Graeme I. ; Refetoff, Samuel. / Human thyroxine-binding globulin gene : Complete sequence and transcriptional regulation. In: Molecular Endocrinology. 1993 ; Vol. 7, No. 8. pp. 1049-1060.
@article{2ca7370ac4484be99b3598defae94a0d,
title = "Human thyroxine-binding globulin gene: Complete sequence and transcriptional regulation",
abstract = "T4-binding globulin (TBG) is a glycoprotein of hepatic origin which transports thyroid hormone in serum. To characterize the human TBG (hTBG) gene, we studied its genomic organization, promoter activity, and regulation. To this purpose, we isolated from liver a complete hTBG cDNA clone containing the 5′-untranslated region and localized the transcription start site (TSS). The analysis of genomic clones revealed that the hTBG gene consists of five exons and that its exon-intron organization is similar to that of other members of the serine protease inhibitor family. The first exon (exon 0) is a short noncoding sequence located 1.62 kilobase pairs (kbp) upstream from exon 1. Potential cis-acting transcriptional regulatory elements including a TATA box, a CAAT box, and a hepatocyte nuclear factor-1 binding motif were identified in the upstream region. A reporter gene in which 3.2 kbp of the 5′-flanking region, including exon 0, was inserted upstream of the bacterial chloramphenicol acetyltransferase gene showed significant activity when transfected into a hepatoblastoma-derived (HepG2) cell line. The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate, down-regulated the promoter activity by more than 80{\%} and completely inhibited hTBG synthesis, whereas thyroid hormone, glucocorticoid, estrogen, and nicotinic acid had little, if any, effect. A series of 5′-deletions revealed that the fragment -218 to +4 from the TSS had the highest promoter activity, nearly 1000-fold greater than the promoterless chloramphenicol acetyltransferase construct. When nonhepatocyte-derived cell lines (CV-1 and CHO) were tested, promoter activity was reduced by a factor of 100, showing that the promoter works in liver-specific manner. The region -218 to -102 contains liver-specific enhancer elements, since deletion to nucleotide -101 resulted in a profound re-duction of the promoter activity in HepG2 cells but not in CV-1 or CHO cells. On the other hand, mutational disruption of the putative hepatocyte nuclear factor-1 site (located 65 bp upstream of the TSS) completely abolished the promoter activity in all cell lines, indicating that this site is absolutely required for the transcription of the hTBG gene.",
author = "Yoshitaka Hayashi and Yuichi Mori and Janssen, {Onno E.} and Thongkum Sunthornthepvarakul and Weiss, {Roy E} and Kyoko Takeda and Michael Weinberg and Hisao Seo and Bell, {Graeme I.} and Samuel Refetoff",
year = "1993",
language = "English (US)",
volume = "7",
pages = "1049--1060",
journal = "Molecular Endocrinology",
issn = "0888-8809",
publisher = "The Endocrine Society",
number = "8",

}

TY - JOUR

T1 - Human thyroxine-binding globulin gene

T2 - Complete sequence and transcriptional regulation

AU - Hayashi, Yoshitaka

AU - Mori, Yuichi

AU - Janssen, Onno E.

AU - Sunthornthepvarakul, Thongkum

AU - Weiss, Roy E

AU - Takeda, Kyoko

AU - Weinberg, Michael

AU - Seo, Hisao

AU - Bell, Graeme I.

AU - Refetoff, Samuel

PY - 1993

Y1 - 1993

N2 - T4-binding globulin (TBG) is a glycoprotein of hepatic origin which transports thyroid hormone in serum. To characterize the human TBG (hTBG) gene, we studied its genomic organization, promoter activity, and regulation. To this purpose, we isolated from liver a complete hTBG cDNA clone containing the 5′-untranslated region and localized the transcription start site (TSS). The analysis of genomic clones revealed that the hTBG gene consists of five exons and that its exon-intron organization is similar to that of other members of the serine protease inhibitor family. The first exon (exon 0) is a short noncoding sequence located 1.62 kilobase pairs (kbp) upstream from exon 1. Potential cis-acting transcriptional regulatory elements including a TATA box, a CAAT box, and a hepatocyte nuclear factor-1 binding motif were identified in the upstream region. A reporter gene in which 3.2 kbp of the 5′-flanking region, including exon 0, was inserted upstream of the bacterial chloramphenicol acetyltransferase gene showed significant activity when transfected into a hepatoblastoma-derived (HepG2) cell line. The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate, down-regulated the promoter activity by more than 80% and completely inhibited hTBG synthesis, whereas thyroid hormone, glucocorticoid, estrogen, and nicotinic acid had little, if any, effect. A series of 5′-deletions revealed that the fragment -218 to +4 from the TSS had the highest promoter activity, nearly 1000-fold greater than the promoterless chloramphenicol acetyltransferase construct. When nonhepatocyte-derived cell lines (CV-1 and CHO) were tested, promoter activity was reduced by a factor of 100, showing that the promoter works in liver-specific manner. The region -218 to -102 contains liver-specific enhancer elements, since deletion to nucleotide -101 resulted in a profound re-duction of the promoter activity in HepG2 cells but not in CV-1 or CHO cells. On the other hand, mutational disruption of the putative hepatocyte nuclear factor-1 site (located 65 bp upstream of the TSS) completely abolished the promoter activity in all cell lines, indicating that this site is absolutely required for the transcription of the hTBG gene.

AB - T4-binding globulin (TBG) is a glycoprotein of hepatic origin which transports thyroid hormone in serum. To characterize the human TBG (hTBG) gene, we studied its genomic organization, promoter activity, and regulation. To this purpose, we isolated from liver a complete hTBG cDNA clone containing the 5′-untranslated region and localized the transcription start site (TSS). The analysis of genomic clones revealed that the hTBG gene consists of five exons and that its exon-intron organization is similar to that of other members of the serine protease inhibitor family. The first exon (exon 0) is a short noncoding sequence located 1.62 kilobase pairs (kbp) upstream from exon 1. Potential cis-acting transcriptional regulatory elements including a TATA box, a CAAT box, and a hepatocyte nuclear factor-1 binding motif were identified in the upstream region. A reporter gene in which 3.2 kbp of the 5′-flanking region, including exon 0, was inserted upstream of the bacterial chloramphenicol acetyltransferase gene showed significant activity when transfected into a hepatoblastoma-derived (HepG2) cell line. The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate, down-regulated the promoter activity by more than 80% and completely inhibited hTBG synthesis, whereas thyroid hormone, glucocorticoid, estrogen, and nicotinic acid had little, if any, effect. A series of 5′-deletions revealed that the fragment -218 to +4 from the TSS had the highest promoter activity, nearly 1000-fold greater than the promoterless chloramphenicol acetyltransferase construct. When nonhepatocyte-derived cell lines (CV-1 and CHO) were tested, promoter activity was reduced by a factor of 100, showing that the promoter works in liver-specific manner. The region -218 to -102 contains liver-specific enhancer elements, since deletion to nucleotide -101 resulted in a profound re-duction of the promoter activity in HepG2 cells but not in CV-1 or CHO cells. On the other hand, mutational disruption of the putative hepatocyte nuclear factor-1 site (located 65 bp upstream of the TSS) completely abolished the promoter activity in all cell lines, indicating that this site is absolutely required for the transcription of the hTBG gene.

UR - http://www.scopus.com/inward/record.url?scp=0027291279&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0027291279&partnerID=8YFLogxK

M3 - Article

C2 - 8232304

AN - SCOPUS:0027291279

VL - 7

SP - 1049

EP - 1060

JO - Molecular Endocrinology

JF - Molecular Endocrinology

SN - 0888-8809

IS - 8

ER -