The vitamin D-responsive element in the rat bone gla protein gene is an imperfect direct repeat that cooperates with other cis-elements in 1,25-dihydroxyvitamin D3-mediated transcriptional activation

Christopher M. Terpening, Carol A. Haussler, Peter W. Jurutka, Michael A. Galligan, Barry S. Komm, Mark R Haussler

Research output: Contribution to journalArticle

115 Citations (Scopus)

Abstract

The gene for rat bone gla protein (BGP) was isolated and 1250 basepairs (bp), including 1100 bp of 5′ flanking DNA, were placed up-stream of the human GH reporter gene. After transient transfection into the osteoblast-like rat osteosarcoma cell line ROS 17/2.8, the BGP promoter demonstrated a low level of basal activity that was increased approximately 10-fold by the addition of 10-8 M 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. A single 250-bp fragment (-523 to -274) was sufficient to confer hormone inducibility upon both heterologous and homologous promoters. Deletion studies, complemented by evaluation with synthetic oligomers, enabled localization of the 1,25-(OH)2D3 response element to within 19 bp (-456 to -438), containing an element with an imperfect direct repeat [GGTGA(N4)GGACA] and homology to other steroid-responsive elements. Gel retardation assays demonstrated that partially purified chick intestinal 1,25-(OH)2D3 receptor bound specifically and with high affinity to a DNA fragment containing the putative 1,25-(OH)2D3 response element, and this binding was perturbed by monoclonal antibodies to the 1,25-(OH)2D3 receptor. Surprisingly, the 250-bp fragment, when linked in an antisense orientation with respect to the BGP promoter, blocked basal and hormone-dependent gene expression. However, a 246-bp fragment 5′ to the 250-bp element (-1100 to -855) restored 20-fold inducibility when linked to the first fragment in the same orientation, suggesting cooperativity between at least two elements to achieve the hormonal regulation observed in this gene.

Original languageEnglish (US)
Pages (from-to)373-385
Number of pages13
JournalMolecular Endocrinology
Volume5
Issue number3
StatePublished - Mar 1991

Fingerprint

Calcitriol
Nucleic Acid Repetitive Sequences
Osteocalcin
Vitamin D
Transcriptional Activation
Response Elements
Hormones
Genes
DNA
Electrophoretic Mobility Shift Assay
Osteosarcoma
Osteoblasts
Reporter Genes
Transfection
Steroids
Monoclonal Antibodies
Gene Expression
Cell Line

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology, Diabetes and Metabolism

Cite this

The vitamin D-responsive element in the rat bone gla protein gene is an imperfect direct repeat that cooperates with other cis-elements in 1,25-dihydroxyvitamin D3-mediated transcriptional activation. / Terpening, Christopher M.; Haussler, Carol A.; Jurutka, Peter W.; Galligan, Michael A.; Komm, Barry S.; Haussler, Mark R.

In: Molecular Endocrinology, Vol. 5, No. 3, 03.1991, p. 373-385.

Research output: Contribution to journalArticle

@article{b8d500750d2a45a4a4754e8cd7bc967f,
title = "The vitamin D-responsive element in the rat bone gla protein gene is an imperfect direct repeat that cooperates with other cis-elements in 1,25-dihydroxyvitamin D3-mediated transcriptional activation",
abstract = "The gene for rat bone gla protein (BGP) was isolated and 1250 basepairs (bp), including 1100 bp of 5′ flanking DNA, were placed up-stream of the human GH reporter gene. After transient transfection into the osteoblast-like rat osteosarcoma cell line ROS 17/2.8, the BGP promoter demonstrated a low level of basal activity that was increased approximately 10-fold by the addition of 10-8 M 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. A single 250-bp fragment (-523 to -274) was sufficient to confer hormone inducibility upon both heterologous and homologous promoters. Deletion studies, complemented by evaluation with synthetic oligomers, enabled localization of the 1,25-(OH)2D3 response element to within 19 bp (-456 to -438), containing an element with an imperfect direct repeat [GGTGA(N4)GGACA] and homology to other steroid-responsive elements. Gel retardation assays demonstrated that partially purified chick intestinal 1,25-(OH)2D3 receptor bound specifically and with high affinity to a DNA fragment containing the putative 1,25-(OH)2D3 response element, and this binding was perturbed by monoclonal antibodies to the 1,25-(OH)2D3 receptor. Surprisingly, the 250-bp fragment, when linked in an antisense orientation with respect to the BGP promoter, blocked basal and hormone-dependent gene expression. However, a 246-bp fragment 5′ to the 250-bp element (-1100 to -855) restored 20-fold inducibility when linked to the first fragment in the same orientation, suggesting cooperativity between at least two elements to achieve the hormonal regulation observed in this gene.",
author = "Terpening, {Christopher M.} and Haussler, {Carol A.} and Jurutka, {Peter W.} and Galligan, {Michael A.} and Komm, {Barry S.} and Haussler, {Mark R}",
year = "1991",
month = "3",
language = "English (US)",
volume = "5",
pages = "373--385",
journal = "Molecular Endocrinology",
issn = "0888-8809",
publisher = "The Endocrine Society",
number = "3",

}

TY - JOUR

T1 - The vitamin D-responsive element in the rat bone gla protein gene is an imperfect direct repeat that cooperates with other cis-elements in 1,25-dihydroxyvitamin D3-mediated transcriptional activation

AU - Terpening, Christopher M.

AU - Haussler, Carol A.

AU - Jurutka, Peter W.

AU - Galligan, Michael A.

AU - Komm, Barry S.

AU - Haussler, Mark R

PY - 1991/3

Y1 - 1991/3

N2 - The gene for rat bone gla protein (BGP) was isolated and 1250 basepairs (bp), including 1100 bp of 5′ flanking DNA, were placed up-stream of the human GH reporter gene. After transient transfection into the osteoblast-like rat osteosarcoma cell line ROS 17/2.8, the BGP promoter demonstrated a low level of basal activity that was increased approximately 10-fold by the addition of 10-8 M 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. A single 250-bp fragment (-523 to -274) was sufficient to confer hormone inducibility upon both heterologous and homologous promoters. Deletion studies, complemented by evaluation with synthetic oligomers, enabled localization of the 1,25-(OH)2D3 response element to within 19 bp (-456 to -438), containing an element with an imperfect direct repeat [GGTGA(N4)GGACA] and homology to other steroid-responsive elements. Gel retardation assays demonstrated that partially purified chick intestinal 1,25-(OH)2D3 receptor bound specifically and with high affinity to a DNA fragment containing the putative 1,25-(OH)2D3 response element, and this binding was perturbed by monoclonal antibodies to the 1,25-(OH)2D3 receptor. Surprisingly, the 250-bp fragment, when linked in an antisense orientation with respect to the BGP promoter, blocked basal and hormone-dependent gene expression. However, a 246-bp fragment 5′ to the 250-bp element (-1100 to -855) restored 20-fold inducibility when linked to the first fragment in the same orientation, suggesting cooperativity between at least two elements to achieve the hormonal regulation observed in this gene.

AB - The gene for rat bone gla protein (BGP) was isolated and 1250 basepairs (bp), including 1100 bp of 5′ flanking DNA, were placed up-stream of the human GH reporter gene. After transient transfection into the osteoblast-like rat osteosarcoma cell line ROS 17/2.8, the BGP promoter demonstrated a low level of basal activity that was increased approximately 10-fold by the addition of 10-8 M 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. A single 250-bp fragment (-523 to -274) was sufficient to confer hormone inducibility upon both heterologous and homologous promoters. Deletion studies, complemented by evaluation with synthetic oligomers, enabled localization of the 1,25-(OH)2D3 response element to within 19 bp (-456 to -438), containing an element with an imperfect direct repeat [GGTGA(N4)GGACA] and homology to other steroid-responsive elements. Gel retardation assays demonstrated that partially purified chick intestinal 1,25-(OH)2D3 receptor bound specifically and with high affinity to a DNA fragment containing the putative 1,25-(OH)2D3 response element, and this binding was perturbed by monoclonal antibodies to the 1,25-(OH)2D3 receptor. Surprisingly, the 250-bp fragment, when linked in an antisense orientation with respect to the BGP promoter, blocked basal and hormone-dependent gene expression. However, a 246-bp fragment 5′ to the 250-bp element (-1100 to -855) restored 20-fold inducibility when linked to the first fragment in the same orientation, suggesting cooperativity between at least two elements to achieve the hormonal regulation observed in this gene.

UR - http://www.scopus.com/inward/record.url?scp=0025761318&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0025761318&partnerID=8YFLogxK

M3 - Article

C2 - 1653893

AN - SCOPUS:0025761318

VL - 5

SP - 373

EP - 385

JO - Molecular Endocrinology

JF - Molecular Endocrinology

SN - 0888-8809

IS - 3

ER -