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Published on: May 27, 2016
Identification of a novel estrogen response element in the breast cancer resistance protein (ABCG2) gene
Pui Lai Rachel Ee1, Sitharthan Kamalakaran, Debra Tonetti
1Department of Biopharmaceutical Sciences, University of Illinois at Chicago, 833 South Wood, Chicago, IL 60612, USA.
Abstract:
The breast cancer resistance protein (BCRP) is an ATP-binding cassette half transporter that confers resistance to anticancer drugs such as mitoxantrone, anthracyclines, topotecan, and SN-38. Initial characterization of the BCRP promoter revealed that it is TATA-less with 5 putative Sp1 sites downstream from a putative CpG island and several AP1 sites (K. J. Bailey-Dell et al., Biochim. Biophys. Acta, 1520: 234-241, 2001). Here, we examined the sequence of the 5'-flanking region of the BCRP gene and found a putative estrogen response element (ERE). We showed that estrogen enhanced the expression of BCRP mRNA in the estrogen receptor (ER)-positive T47D:A18 cells and PA-1 cells stably expressing ERalpha. In BCRP promoter-luciferase assays, sequential deletions of the BCRP promoter showed that the region between -243 and -115 is essential for the ER effect. Mutation of the ERE found within this region attenuated the estrogen response, whereas deletion of the site completely abrogated the estrogen effect. Furthermore, electrophoretic mobility shift assays revealed specific binding of ERalpha to the BCRP promoter through the identified ERE. Taken together, we provide evidence herein for a novel ERE in the BCRP promoter.
Insights
Estrogen enhances breast cancer resistance protein (BCRP) expression by binding to a newly identified estrogen response element (ERE) in the BCRP promoter, impacting drug resistance. This discovery reveals a novel regulatory mechanism for BCRP.
Area of Science:
- Molecular Biology
- Genetics
- Cancer Research
Background:
- Breast cancer resistance protein (BCRP) is a transporter protein involved in multidrug resistance.
- BCRP confers resistance to various anticancer drugs, including mitoxantrone and anthracyclines.
- Previous studies identified Sp1 and AP1 sites in the BCRP promoter, but its regulation by other factors was unclear.
Purpose of the Study:
- To investigate the regulation of the breast cancer resistance protein (BCRP) gene expression by estrogen.
- To identify potential regulatory elements in the 5'-flanking region of the BCRP gene.
- To elucidate the mechanism by which estrogen influences BCRP expression.
Main Methods:
- Sequence analysis of the BCRP 5'-flanking region to identify putative regulatory elements.
- Analysis of BCRP mRNA expression in response to estrogen in ER-positive cell lines.
- BCRP promoter-luciferase assays with sequential deletions and mutations.
- Electrophoretic mobility shift assays (EMSA) to assess ERalpha binding to the BCRP promoter.
Main Results:
- A putative estrogen response element (ERE) was identified in the BCRP promoter.
- Estrogen significantly enhanced BCRP mRNA expression in ER-positive cells.
- A specific region (-243 to -115) of the BCRP promoter was crucial for the estrogenic effect.
- Mutation or deletion of the identified ERE abolished the estrogen-induced BCRP expression.
- ERalpha was shown to bind directly to the BCRP promoter at the identified ERE.
Conclusions:
- A novel estrogen response element (ERE) in the breast cancer resistance protein (BCRP) promoter has been identified.
- Estrogen receptor alpha (ERalpha) directly binds to this ERE, leading to enhanced BCRP gene expression.
- This finding reveals a new mechanism for regulating BCRP, potentially influencing drug resistance in cancer therapy.
