The major goal of this investigation is to elucidate the role of specific intracellular receptor proteins in the estrogenic regulation of cell proliferation and function in hormone-responsive tissues and cancers derived from these tissues. The proposal focuses on the use of biochemical, immunochemical and recombinant DNA techniques to determine the structure, composition and intracellular interactions and dynamics of the estrogen receptor and progesterone receptor from human or rodent sources. We are especially interested in the molecular mechanisms by which activity and expression of these receptors are regulated by estrogens, progestins and their antagonists in breast cancer cell lines. This work is facilitated by the availability in our laboratory of purified, or partially purified receptor proteins, monoclonal antibodies specific for both receptors, and cloned cDNAs that correspond to all of the translated mRNA for human ER from MCF-7 breast cancer cells and 90% of the translated mRNA for human PR from T47D breast cancer cells.
Specific aims i nclude: 1) To isolate, sequence and characterize cDNA clones corresponding to the entire mRNA for T47D human PR; 2) to isolate, sequence and characterize genomic clones for human ER and PR, including upstream regulatory sequences; 3) to characterize the regulation of ER and PR expression in breast cancer cells and rat uterus by estrogens, progestins and their antagonists; 4) to locate and characterize ER and any PR binding sites (responses elements) on the chromosomal genes for both receptors; 5) to dissect and characterize the functional domains of both receptors by in vitro mutagenesis, including domain switching between ER and PR, coupled with transient expression assays in heterologous cells containing estrogen- or progestin-responsive reporter genes; 6) to isolate mutant fragments either from transfected cells or from in vitro translations of transcripts and study the steroid-binding and specific DNA-binding properties of these fragments; 7) to overexpress human ER and PR cDNAs in suitable cell lines, purify expressed receptor and carry out detailed structural and biochemical studies on full length receptor or genetically engineered receptor fragments; 8) to determine the amino acids involved in the binding of both agonists and antagonists, and the location, role and hormonal regulation of phosphorylation sites in rat and human ER and PR.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA002897-36
Application #
3163127
Study Section
Biochemical Endocrinology Study Section (BCE)
Project Start
1977-04-01
Project End
1994-03-31
Budget Start
1992-04-01
Budget End
1994-03-31
Support Year
36
Fiscal Year
1992
Total Cost
Indirect Cost
Name
University of Chicago
Department
Type
Schools of Medicine
DUNS #
225410919
City
Chicago
State
IL
Country
United States
Zip Code
60637
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Landel, C C; Kushner, P J; Greene, G L (1995) Estrogen receptor accessory proteins: effects on receptor-DNA interactions. Environ Health Perspect 103 Suppl 7:23-8
Pertschuk, L P; Schaeffer, H; Feldman, J G et al. (1995) Immunostaining for prostate cancer androgen receptor in paraffin identifies a subset of men with a poor prognosis. Lab Invest 73:302-5
Landel, C C; Kushner, P J; Greene, G L (1994) The interaction of human estrogen receptor with DNA is modulated by receptor-associated proteins. Mol Endocrinol 8:1407-19
Nardulli, A M; Greene, G L; Shapiro, D J (1993) Human estrogen receptor bound to an estrogen response element bends DNA. Mol Endocrinol 7:331-40

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