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Title: 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 isoelectricmore » 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.« less
Authors:
; ; ; ;
Publication Date:
OSTI Identifier:
6472430
Resource Type:
Journal Article
Resource Relation:
Journal Name: Endocrinology; (United States); Journal Volume: 115:1
Research Org:
Univ. of Illinois, Urbana
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 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 550301* -- Cytology-- Tracer Techniques; 550201 -- Biochemistry-- Tracer Techniques