Examination of the potential functional role of conserved cysteine residues in the hormone binding domain of the human 1,25-dihydroxyvitamin D3 receptor

Shigeo Nakajima, Jui Cheng Hsieh, Peter Jurutka, Michael A. Galligan, Carol A. Haussler, G. Kerr Whitfield, Mark R. Haussler

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

The significance of conserved cysteines at positions 288, 337, and 369 in the hormone binding domain of the human vitamin D receptor was evaluated by individual site-directed mutagenesis to glycine. Neither nuclear localization nor heterodimerization with retinoid X receptors in binding to the vitamin D- responsive element was appreciably affected by altering these cysteines, but vitamin D hormone (1,25-(OH)2D3) activated transcription was compromised significantly in the C288G and C337G mutants. Only the C288G mutant displayed depressed (3-fold) 1,25-(OH)2D3 ligand binding affinity at 4 °C, in vitro, although at elevated temperatures (23-37 °C), ligand binding was attenuated severely in C288G, moderately in C337G and very mildly in C369G. The degree of impairment of ligand binding at physiologic temperatures correlated with the requirement for increased concentrations of 1,25-(OH)2D3 ligand to maximally stimulate transcriptional activity in co-transfected COS-7 cells. Thus cysteine 288 and, to a lesser extent, cysteine 337 are important for high affinity hormone binding to the vitamin D receptor, which ultimately leads to ligand-dependent transcriptional activation.

Original languageEnglish (US)
Pages (from-to)5143-5149
Number of pages7
JournalJournal of Biological Chemistry
Volume271
Issue number9
DOIs
StatePublished - Mar 1 1996

Fingerprint

Calcitriol Receptors
Cysteine
Hormones
Ligands
Vitamin D
Retinoid X Receptors
Mutagenesis
Temperature
COS Cells
Transcription
Site-Directed Mutagenesis
Glycine
Transcriptional Activation
Chemical activation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Examination of the potential functional role of conserved cysteine residues in the hormone binding domain of the human 1,25-dihydroxyvitamin D3 receptor. / Nakajima, Shigeo; Hsieh, Jui Cheng; Jurutka, Peter; Galligan, Michael A.; Haussler, Carol A.; Whitfield, G. Kerr; Haussler, Mark R.

In: Journal of Biological Chemistry, Vol. 271, No. 9, 01.03.1996, p. 5143-5149.

Research output: Contribution to journalArticle

Nakajima, Shigeo ; Hsieh, Jui Cheng ; Jurutka, Peter ; Galligan, Michael A. ; Haussler, Carol A. ; Whitfield, G. Kerr ; Haussler, Mark R. / Examination of the potential functional role of conserved cysteine residues in the hormone binding domain of the human 1,25-dihydroxyvitamin D3 receptor. In: Journal of Biological Chemistry. 1996 ; Vol. 271, No. 9. pp. 5143-5149.
@article{7429d7377c2a494ebc454bd88c0d30e3,
title = "Examination of the potential functional role of conserved cysteine residues in the hormone binding domain of the human 1,25-dihydroxyvitamin D3 receptor",
abstract = "The significance of conserved cysteines at positions 288, 337, and 369 in the hormone binding domain of the human vitamin D receptor was evaluated by individual site-directed mutagenesis to glycine. Neither nuclear localization nor heterodimerization with retinoid X receptors in binding to the vitamin D- responsive element was appreciably affected by altering these cysteines, but vitamin D hormone (1,25-(OH)2D3) activated transcription was compromised significantly in the C288G and C337G mutants. Only the C288G mutant displayed depressed (3-fold) 1,25-(OH)2D3 ligand binding affinity at 4 °C, in vitro, although at elevated temperatures (23-37 °C), ligand binding was attenuated severely in C288G, moderately in C337G and very mildly in C369G. The degree of impairment of ligand binding at physiologic temperatures correlated with the requirement for increased concentrations of 1,25-(OH)2D3 ligand to maximally stimulate transcriptional activity in co-transfected COS-7 cells. Thus cysteine 288 and, to a lesser extent, cysteine 337 are important for high affinity hormone binding to the vitamin D receptor, which ultimately leads to ligand-dependent transcriptional activation.",
author = "Shigeo Nakajima and Hsieh, {Jui Cheng} and Peter Jurutka and Galligan, {Michael A.} and Haussler, {Carol A.} and Whitfield, {G. Kerr} and Haussler, {Mark R.}",
year = "1996",
month = "3",
day = "1",
doi = "10.1074/jbc.271.9.5143",
language = "English (US)",
volume = "271",
pages = "5143--5149",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "9",

}

TY - JOUR

T1 - Examination of the potential functional role of conserved cysteine residues in the hormone binding domain of the human 1,25-dihydroxyvitamin D3 receptor

AU - Nakajima, Shigeo

AU - Hsieh, Jui Cheng

AU - Jurutka, Peter

AU - Galligan, Michael A.

AU - Haussler, Carol A.

AU - Whitfield, G. Kerr

AU - Haussler, Mark R.

PY - 1996/3/1

Y1 - 1996/3/1

N2 - The significance of conserved cysteines at positions 288, 337, and 369 in the hormone binding domain of the human vitamin D receptor was evaluated by individual site-directed mutagenesis to glycine. Neither nuclear localization nor heterodimerization with retinoid X receptors in binding to the vitamin D- responsive element was appreciably affected by altering these cysteines, but vitamin D hormone (1,25-(OH)2D3) activated transcription was compromised significantly in the C288G and C337G mutants. Only the C288G mutant displayed depressed (3-fold) 1,25-(OH)2D3 ligand binding affinity at 4 °C, in vitro, although at elevated temperatures (23-37 °C), ligand binding was attenuated severely in C288G, moderately in C337G and very mildly in C369G. The degree of impairment of ligand binding at physiologic temperatures correlated with the requirement for increased concentrations of 1,25-(OH)2D3 ligand to maximally stimulate transcriptional activity in co-transfected COS-7 cells. Thus cysteine 288 and, to a lesser extent, cysteine 337 are important for high affinity hormone binding to the vitamin D receptor, which ultimately leads to ligand-dependent transcriptional activation.

AB - The significance of conserved cysteines at positions 288, 337, and 369 in the hormone binding domain of the human vitamin D receptor was evaluated by individual site-directed mutagenesis to glycine. Neither nuclear localization nor heterodimerization with retinoid X receptors in binding to the vitamin D- responsive element was appreciably affected by altering these cysteines, but vitamin D hormone (1,25-(OH)2D3) activated transcription was compromised significantly in the C288G and C337G mutants. Only the C288G mutant displayed depressed (3-fold) 1,25-(OH)2D3 ligand binding affinity at 4 °C, in vitro, although at elevated temperatures (23-37 °C), ligand binding was attenuated severely in C288G, moderately in C337G and very mildly in C369G. The degree of impairment of ligand binding at physiologic temperatures correlated with the requirement for increased concentrations of 1,25-(OH)2D3 ligand to maximally stimulate transcriptional activity in co-transfected COS-7 cells. Thus cysteine 288 and, to a lesser extent, cysteine 337 are important for high affinity hormone binding to the vitamin D receptor, which ultimately leads to ligand-dependent transcriptional activation.

UR - http://www.scopus.com/inward/record.url?scp=0029928026&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029928026&partnerID=8YFLogxK

U2 - 10.1074/jbc.271.9.5143

DO - 10.1074/jbc.271.9.5143

M3 - Article

C2 - 8617794

AN - SCOPUS:0029928026

VL - 271

SP - 5143

EP - 5149

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 9

ER -