University of Louisville
Tamoxifen Increases Nuclear Respiratory Factor-1 Transcription by Activating Estrogen Receptors Alpha and Beta Interaction with an Estrogen Response Element in the Gene Promoter
Institution
University of Louisville
Faculty Advisor/ Mentor
Carolyn Klinge; Margarita Ivanova
Abstract
Estrogens promote breast tumorigenesis by binding to estrogen receptors (ERs), encoded by two genes, alpha and beta. Tamoxifen competes with estradiol (E2) for binding ERs and is used clinically to inhibit breast cancer progression. E2-ER alpha increases nuclear respiratory factor1 (NRF-1) transcription in MCF-7 breast cancer cells. Dysregulation of NRF-1 and its downstream targets are associated with poor prognosis in breast cancer. Surprisingly, the active metabolite of tamoxifen, 4-hydroxytamoxifen (4-OHT), increased NRF-1 transcription in MCF-7 cells. To determine the mechanism for 4-OHT’s induction of NRF-1 transcription, the 5’ -1100 bp promoter of the human NRF-1 gene was transiently transfected with ER alpha or ER beta in ER-null COS7 cells. Both E2 and 4-OHT increased luciferase reporter activity with ER alpha and ER beta and the transcriptional activity of 4-OHT was blocked by the antiestrogen ICI 182,780, confirming that the response was ER-mediated. Mutation of the ERE in the -1100 promoter of the NRF-1 gene blocked E2 or 4-OHT- induction of luciferase activity. Together these data indicate that both ER alpha and ER beta are involved in E2 and 4-OHT upregulation of NRF-1 gene transcription. Because TFAM is a downstream target of NRF-1, an additional experiment examined the effect of E2 or 4-OHT on TFAM expression in MCF-7 cells. Results of quantitative PCR analysis showed E2 induced higher TFAM expression than 4-OHT. Thus, despite 4-OHT’s induction of NRF-1, there is less NRF-1-mediated transcription in the 4-OHTtreated cells. This is likely due to induction of apoptosis in MCF-7 cells by 4-OHT treatment.
Tamoxifen Increases Nuclear Respiratory Factor-1 Transcription by Activating Estrogen Receptors Alpha and Beta Interaction with an Estrogen Response Element in the Gene Promoter
Estrogens promote breast tumorigenesis by binding to estrogen receptors (ERs), encoded by two genes, alpha and beta. Tamoxifen competes with estradiol (E2) for binding ERs and is used clinically to inhibit breast cancer progression. E2-ER alpha increases nuclear respiratory factor1 (NRF-1) transcription in MCF-7 breast cancer cells. Dysregulation of NRF-1 and its downstream targets are associated with poor prognosis in breast cancer. Surprisingly, the active metabolite of tamoxifen, 4-hydroxytamoxifen (4-OHT), increased NRF-1 transcription in MCF-7 cells. To determine the mechanism for 4-OHT’s induction of NRF-1 transcription, the 5’ -1100 bp promoter of the human NRF-1 gene was transiently transfected with ER alpha or ER beta in ER-null COS7 cells. Both E2 and 4-OHT increased luciferase reporter activity with ER alpha and ER beta and the transcriptional activity of 4-OHT was blocked by the antiestrogen ICI 182,780, confirming that the response was ER-mediated. Mutation of the ERE in the -1100 promoter of the NRF-1 gene blocked E2 or 4-OHT- induction of luciferase activity. Together these data indicate that both ER alpha and ER beta are involved in E2 and 4-OHT upregulation of NRF-1 gene transcription. Because TFAM is a downstream target of NRF-1, an additional experiment examined the effect of E2 or 4-OHT on TFAM expression in MCF-7 cells. Results of quantitative PCR analysis showed E2 induced higher TFAM expression than 4-OHT. Thus, despite 4-OHT’s induction of NRF-1, there is less NRF-1-mediated transcription in the 4-OHTtreated cells. This is likely due to induction of apoptosis in MCF-7 cells by 4-OHT treatment.