The expression of estrogen receptor (ER) has a profound influence on the biology of breast carcinoma. Patients with ER-positive tumors tend to have a better prognosis than patients with carcinomas which lack receptor expression. However, little is known about mechanisms which control ER expression. By utilizing a cell line model of breast cancer, preliminary studies have determined that ER expression is controlled through transcriptional regulation. No previous studies have defined the ER promoter.
The specific aim of this application is to characterize the ER promoter and define important transcriptional control regions which are functional in breast carcinoma. These results will then be used to determine the mechanism responsible for the lack of ER expression in ER- negative carcinomas. The ER promoter will be functionally mapped using a luciferase reporter system. A human genomic lambda library will be screened to identify clones containing genomic DNA from the 5' end of the ER gene. Various regions of DNA upstream of the transcriptional start site of the ER gene will be cloned into luciferase reporter plasmids. The first intron from the ER gene is 20 kbp and this region will also be examined for enhancer activity using the luciferase reporter system. These constructs will be transfected into MCF-7 (ER positive) and MDA-MB-231 (ER negative) cells to define what region of the ER gene are necessary for ER expression. One of two possible results seems most likely; either these constructs will function the same in both cell lines or the MCF-7 cell line will demonstrate expression while MDA-MB-231 will not. These results will be useful to determine if the lack of expression in MDA-MB-231 is due to a cis-effect defined by DNA mutations or methylation or a trans-effect possibly due to a different transcription factor(s) functioning in these two cell lines. It is hoped that these experiments will define a mechanism responsible for the varied expression found in these breast cancer cell lines. Finally, the universality of these mechanisms will be explored by examining a variety of other cell lines and fresh cancer isolates.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
1R29CA063251-01
Application #
2104986
Study Section
Reproductive Endocrinology Study Section (REN)
Project Start
1994-05-01
Project End
1999-04-30
Budget Start
1994-05-01
Budget End
1995-04-30
Support Year
1
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Stanford University
Department
Surgery
Type
Schools of Medicine
DUNS #
800771545
City
Stanford
State
CA
Country
United States
Zip Code
94305
Nakatani, K; Sakaue, H; Thompson, D A et al. (1999) Identification of a human Akt3 (protein kinase B gamma) which contains the regulatory serine phosphorylation site. Biochem Biophys Res Commun 257:906-10
Hoch, R V; Thompson, D A; Baker, R J et al. (1999) GATA-3 is expressed in association with estrogen receptor in breast cancer. Int J Cancer 84:122-8
McPherson, L A; Weigel, R J (1999) AP2alpha and AP2gamma: a comparison of binding site specificity and trans-activation of the estrogen receptor promoter and single site promoter constructs. Nucleic Acids Res 27:4040-9
Nakatani, K; Thompson, D A; Barthel, A et al. (1999) Up-regulation of Akt3 in estrogen receptor-deficient breast cancers and androgen-independent prostate cancer lines. J Biol Chem 274:21528-32
Kuang, W W; Thompson, D A; Hoch, R V et al. (1998) Differential screening and suppression subtractive hybridization identified genes differentially expressed in an estrogen receptor-positive breast carcinoma cell line. Nucleic Acids Res 26:1116-23
Thompson, D A; Weigel, R J (1998) Characterization of a gene that is inversely correlated with estrogen receptor expression (ICERE-1) in breast carcinomas. Eur J Biochem 252:169-77
Thompson, D A; Weigel, R J (1998) hAG-2, the human homologue of the Xenopus laevis cement gland gene XAG-2, is coexpressed with estrogen receptor in breast cancer cell lines. Biochem Biophys Res Commun 251:111-6
Thompson, D A; McPherson, L A; Carmeci, C et al. (1997) Identification of two estrogen receptor transcripts with novel 5' exons isolated from a MCF7 cDNA library. J Steroid Biochem Mol Biol 62:143-53
Carmeci, C; Thompson, D A; Ring, H Z et al. (1997) Identification of a gene (GPR30) with homology to the G-protein-coupled receptor superfamily associated with estrogen receptor expression in breast cancer. Genomics 45:607-17
McPherson, L A; Baichwal, V R; Weigel, R J (1997) Identification of ERF-1 as a member of the AP2 transcription factor family. Proc Natl Acad Sci U S A 94:4342-7

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