The Synonymous Ala87 Mutation of Estrogen Receptor Alpha Modifies Transcriptional Activation Through Both ERE and AP1 Sites

  • Tamara Fernández-Calero
  • Gilles Flouriot
  • Mónica Marín
Part of the Methods in Molecular Biology book series (MIMB, volume 1366)

Abstract

Estrogen receptor α (ERα) exerts regulatory actions through genomic mechanisms. In the classical pathway, ligand-activated ERα binds directly to DNA through estrogen response elements (ERE) located in the promoter of target genes. ERα can also exert indirect regulation of transcription via protein-protein interaction with other transcription factors such as AP-1.

Several ERα synonymous polymorphisms have been identified and efforts to understand their implications have been made. Nevertheless effects of synonymous polymorphisms are still neglected. This chapter focuses on the experimental procedure employed in order to characterize the transcriptional activity of a synonymous polymorphism of the ERα (rs746432) called Alanine 87 (Ala87). Activity of both WT and Ala87 ERα isoforms on transcriptional pathways can be analyzed in transiently transfected cells using different reporter constructs. ERα efficiency on the classical genomic pathway can be analyzed by determining its transactivation activity on an ERE-driven thymidine kinase (TK) promoter controlling the expression of the luciferase reporter gene. Transcriptional activity through the indirect genomic pathway can be analyzed by employing an AP-1 DNA response element-driven promoter also controlling the expression of luciferase reporter gene.

Key words

Estrogenreceptoralpha (ERα) Estrogen transcriptional regulation Estrogen-responsive element (ERE) Nonclassical pathway AP-1pathway Estrogenreceptor alanine 87 polymorphism 

Notes

Acknowledgments

This work was partially supported by Fondo Clemente Estable, ANII (Uruguay), and PEDECIBA.

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Tamara Fernández-Calero
    • 1
    • 2
  • Gilles Flouriot
    • 3
  • Mónica Marín
    • 1
  1. 1.Biochemistry-Molecular Biology, Facultad de CienciasUniversidad de la RepúblicaMontevideoUruguay
  2. 2.Bioinformatics UnitInstitut Pasteur MontevideoMontevideoUruguay
  3. 3.Université de Rennes 1, Institut de Recherche en Santé, Environnement et Travail, IRSETINSERM U1085RennesFrance

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