The N-terminal adenosine triphosphate binding domain of Hsp90 is necessary and sufficient for interaction with estrogen receptor

I. Bouhouche-Chatelier, Ahmed Chadli, M. G. Catelli

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

To understand how the molecular chaperone Hsp90 participates in conformational maturation of the estrogen receptor (ER), we analyzed the interaction of immobilized purified avian Hsp90 with mammalian cytosolic ER. Hsp90 was either immunoadsorbed to BF4 antibody-Sepharose or GST-Hsp90 fusion protein (GST.90) was adsorbed to glutathione-Sepharose. GST.90 was able to retain specifically ER, similarly to immunoadsorbed Hsp90. When cells were treated with estradiol and the hormone treatment was maintained during cell homogenization, binding, and washing steps, GST.90 still interacted efficiently with ER, suggesting that ER may form complexes with Hsp90 even after its activation by hormone and salt extraction from nuclei. The GST.90-ER interaction was consistently reduced in the presence of increasing concentrations of potassium chloride or when cytosolic ER-Hsp90 complexes were previously stabilized by molybdate, indicating that GST.90-ER complexes behave like cytosolic Hsp90-ER complexes. A purified thioredoxin-ER fusion protein was also able to form complexes with GST.90, suggesting that the presence of other chaperones is not required. ER was retained only by GST.90 deletion mutants bearing an intact Hsp90 N-terminal region (1-224), the interaction being more efficient when the charged region A was present in the mutant (1-334). The N-terminal fragment 1-334, devoid of the dimeric GST moiety, was also able to interact with ER, pointing to the monomeric N-terminal adenosine triphosphate binding region of Hsp90 (1-224) as the region necessary and sufficient for interaction. These results contribute to understand the Hsp90-dependent process responsible for conformational competence of ER.

Original languageEnglish (US)
Pages (from-to)297-305
Number of pages9
JournalCell Stress and Chaperones
Volume6
Issue number4
DOIs
StatePublished - Dec 19 2001

Fingerprint

Estrogen Receptors
Adenosine Triphosphate
Sepharose
Bearings (structural)
Fusion reactions
Hormones
Thioredoxins
Molecular Chaperones
Potassium Chloride
Mental Competency
Washing
Glutathione
Estradiol
Salts
Proteins
Chemical activation
Antibodies

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

Cite this

The N-terminal adenosine triphosphate binding domain of Hsp90 is necessary and sufficient for interaction with estrogen receptor. / Bouhouche-Chatelier, I.; Chadli, Ahmed; Catelli, M. G.

In: Cell Stress and Chaperones, Vol. 6, No. 4, 19.12.2001, p. 297-305.

Research output: Contribution to journalArticle

@article{b98d2299800646adaf60d7ec4a2fa33b,
title = "The N-terminal adenosine triphosphate binding domain of Hsp90 is necessary and sufficient for interaction with estrogen receptor",
abstract = "To understand how the molecular chaperone Hsp90 participates in conformational maturation of the estrogen receptor (ER), we analyzed the interaction of immobilized purified avian Hsp90 with mammalian cytosolic ER. Hsp90 was either immunoadsorbed to BF4 antibody-Sepharose or GST-Hsp90 fusion protein (GST.90) was adsorbed to glutathione-Sepharose. GST.90 was able to retain specifically ER, similarly to immunoadsorbed Hsp90. When cells were treated with estradiol and the hormone treatment was maintained during cell homogenization, binding, and washing steps, GST.90 still interacted efficiently with ER, suggesting that ER may form complexes with Hsp90 even after its activation by hormone and salt extraction from nuclei. The GST.90-ER interaction was consistently reduced in the presence of increasing concentrations of potassium chloride or when cytosolic ER-Hsp90 complexes were previously stabilized by molybdate, indicating that GST.90-ER complexes behave like cytosolic Hsp90-ER complexes. A purified thioredoxin-ER fusion protein was also able to form complexes with GST.90, suggesting that the presence of other chaperones is not required. ER was retained only by GST.90 deletion mutants bearing an intact Hsp90 N-terminal region (1-224), the interaction being more efficient when the charged region A was present in the mutant (1-334). The N-terminal fragment 1-334, devoid of the dimeric GST moiety, was also able to interact with ER, pointing to the monomeric N-terminal adenosine triphosphate binding region of Hsp90 (1-224) as the region necessary and sufficient for interaction. These results contribute to understand the Hsp90-dependent process responsible for conformational competence of ER.",
author = "I. Bouhouche-Chatelier and Ahmed Chadli and Catelli, {M. G.}",
year = "2001",
month = "12",
day = "19",
doi = "10.1379/1466-1268(2001)006<0297:TNTATB>2.0.CO;2",
language = "English (US)",
volume = "6",
pages = "297--305",
journal = "Cell Stress and Chaperones",
issn = "1355-8145",
publisher = "Springer Netherlands",
number = "4",

}

TY - JOUR

T1 - The N-terminal adenosine triphosphate binding domain of Hsp90 is necessary and sufficient for interaction with estrogen receptor

AU - Bouhouche-Chatelier, I.

AU - Chadli, Ahmed

AU - Catelli, M. G.

PY - 2001/12/19

Y1 - 2001/12/19

N2 - To understand how the molecular chaperone Hsp90 participates in conformational maturation of the estrogen receptor (ER), we analyzed the interaction of immobilized purified avian Hsp90 with mammalian cytosolic ER. Hsp90 was either immunoadsorbed to BF4 antibody-Sepharose or GST-Hsp90 fusion protein (GST.90) was adsorbed to glutathione-Sepharose. GST.90 was able to retain specifically ER, similarly to immunoadsorbed Hsp90. When cells were treated with estradiol and the hormone treatment was maintained during cell homogenization, binding, and washing steps, GST.90 still interacted efficiently with ER, suggesting that ER may form complexes with Hsp90 even after its activation by hormone and salt extraction from nuclei. The GST.90-ER interaction was consistently reduced in the presence of increasing concentrations of potassium chloride or when cytosolic ER-Hsp90 complexes were previously stabilized by molybdate, indicating that GST.90-ER complexes behave like cytosolic Hsp90-ER complexes. A purified thioredoxin-ER fusion protein was also able to form complexes with GST.90, suggesting that the presence of other chaperones is not required. ER was retained only by GST.90 deletion mutants bearing an intact Hsp90 N-terminal region (1-224), the interaction being more efficient when the charged region A was present in the mutant (1-334). The N-terminal fragment 1-334, devoid of the dimeric GST moiety, was also able to interact with ER, pointing to the monomeric N-terminal adenosine triphosphate binding region of Hsp90 (1-224) as the region necessary and sufficient for interaction. These results contribute to understand the Hsp90-dependent process responsible for conformational competence of ER.

AB - To understand how the molecular chaperone Hsp90 participates in conformational maturation of the estrogen receptor (ER), we analyzed the interaction of immobilized purified avian Hsp90 with mammalian cytosolic ER. Hsp90 was either immunoadsorbed to BF4 antibody-Sepharose or GST-Hsp90 fusion protein (GST.90) was adsorbed to glutathione-Sepharose. GST.90 was able to retain specifically ER, similarly to immunoadsorbed Hsp90. When cells were treated with estradiol and the hormone treatment was maintained during cell homogenization, binding, and washing steps, GST.90 still interacted efficiently with ER, suggesting that ER may form complexes with Hsp90 even after its activation by hormone and salt extraction from nuclei. The GST.90-ER interaction was consistently reduced in the presence of increasing concentrations of potassium chloride or when cytosolic ER-Hsp90 complexes were previously stabilized by molybdate, indicating that GST.90-ER complexes behave like cytosolic Hsp90-ER complexes. A purified thioredoxin-ER fusion protein was also able to form complexes with GST.90, suggesting that the presence of other chaperones is not required. ER was retained only by GST.90 deletion mutants bearing an intact Hsp90 N-terminal region (1-224), the interaction being more efficient when the charged region A was present in the mutant (1-334). The N-terminal fragment 1-334, devoid of the dimeric GST moiety, was also able to interact with ER, pointing to the monomeric N-terminal adenosine triphosphate binding region of Hsp90 (1-224) as the region necessary and sufficient for interaction. These results contribute to understand the Hsp90-dependent process responsible for conformational competence of ER.

UR - http://www.scopus.com/inward/record.url?scp=0035202753&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0035202753&partnerID=8YFLogxK

U2 - 10.1379/1466-1268(2001)006<0297:TNTATB>2.0.CO;2

DO - 10.1379/1466-1268(2001)006<0297:TNTATB>2.0.CO;2

M3 - Article

C2 - 11795466

AN - SCOPUS:0035202753

VL - 6

SP - 297

EP - 305

JO - Cell Stress and Chaperones

JF - Cell Stress and Chaperones

SN - 1355-8145

IS - 4

ER -