Complex hormonal, growth factor and complimentary receptor interactions regulate proliferative and differentiative changes in normal cycling uterine tissues and in estrogen-dependent tumor cells and tissues. Estrogen induction, repression and depression of various nuclear genes are known to mediate these proliferative and differentiative events. How antiestrogens, like Tamoxifen, regulate early events in estrogen induction is totally undetermined. Consequently, the major objective of this study will be to investigate the regulatory effects of estrogen and Tamoxifen on specific genes known to play key roles in proliferation and/or differentiation in rodent uterine tissue. Specifically, the probes used in these studies to evaluate proliferation and differentiation will include growth factors (EGF, IGF-1, IGF-2 Exon 2, IGF-2 Exon 4, TGFBeta-types 1,2,3), growth factor receptors (EGFR, IGF- 1R, IGF-2R, TGFBeta-2R,3R), hormone receptors (ER,hPR and protooncogenes (c-myc; c-fos; c-ras; c-Jun). In this temporal study, polyA+ RNA will be isolated from estrogen and Tamoxifen-treated uteri at 0',15,30', 1hr, 6hr, 24hr, 48hr, and 72hr. Expression studies will be analyzed by dot/Northern blot, and dot blot/hybridization with the above mentioned probes. Autoradiograms will be analyzed by densitometric scans. In related studies we will determine if the peroxidase gene is under the direct control of gene products activated by the E2-ER complex. To answer this question, dot/slot studies using recombinant DNA clones from a genomic library will be used to identify estrogen-induced peroxidase (EIP) mRNAs. Cloned DNA fragments homologous with the EIP mRNA will be sequenced and hybridized to chromatin from uteri in an attempt to localize the EIP gene.
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