Characterization of the estrogen receptor and its dynamics in MCF-7 human breast cancer cells using a covalently attaching antiestrogen
The authors have used a covalently attaching antiestrogen, tamoxifen aziridine TA to analyze the structure and dynamics of the estrogen receptor in MCF-7 human breast cancer cells. The labeling of receptor with (/sup 3/H)TA is specific, being blocked only by estrogens and antiestrogens, and the labeling is very efficient in that TA labels covalently the same number of receptors that are labeled reversibly by estradiol. In cells exposed to (/sup 3/H)TA for 1 h, most of the covalently associated radioactivity is found in the 0.6 M KCl extract of the nuclear fraction; this receptor has an apparent mol wt of 63,000 +/- 2000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and a pI of 5.7 by gel isoelectric focusing in the presence of 8 M urea. The mol wt and pI of cytosol receptor labeled with (/sup 3/H) TA are identical. In cells labeled with (/sup 3/H)TA (20 nM) for 1 h and then exposed to a chase of 10(-6) M estradiol, (3H)TA-labeled nuclear receptor disappears with a half-life of 4 h. Affinity labeled receptor interacts with several monoclonal antibodies to MCF-7 estrogen receptor, and it can be purified extensively by immunoadsorbent chromatography. The findings of similar mol wt and isoelectric points for soluble cytosol and nuclear extracted receptors under strongly denaturing and disaggregating conditions reveal that nuclear localization of receptor after ligand binding is not associated with major structural alterations in the receptor component labeled by TA.
- Research Organization:
- Univ. of Illinois, Urbana
- OSTI ID:
- 6472430
- Journal Information:
- Endocrinology; (United States), Vol. 115:1
- Country of Publication:
- United States
- Language:
- English
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RECEPTORS
MOLECULAR STRUCTURE
MOLECULAR WEIGHT
ESTROGENS
MAMMARY GLANDS
MONOCLONAL ANTIBODIES
TRITIUM COMPOUNDS
TUMOR CELLS
ANIMAL CELLS
ANTIBODIES
BODY
GLANDS
HORMONES
LABELLED COMPOUNDS
ORGANS
STEROID HORMONES
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