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HTRF® Total and Phospho-YAP (Ser127) Cellular Assays

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The Hippo Pathway

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1893))

Abstract

The YAP protein is a co-transcription factor increasing the expression of genes involved in cell proliferation and repressing the expression of genes important for cell differentiation and apoptosis. It is regulated by several inputs, like the Hippo pathway, through the action of kinases that phosphorylate YAP on several residues. The level of phosphorylation of the residues serine 127 (S127) of YAP is generally assessed in cellular models, native tissues, and organs, as a marker of YAP activity and location, and is regulated by numerous partners. This phosphorylation event is classically detected using a western blot technical approach. Here, we describe a novel approach to detect both the relative amount of total YAP (T-YAP assay) and the phosphorylation of the residue S127 of YAP (S127-P-YAP assay) using a HTRF®-based method. This easy-to-run method can easily be miniaturized and allows for a high-throughput analysis in 96/384-well plate format, requiring less cellular material and being more rapid than other approaches.

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Acknowledgment

The work described was made possible through Cisbio Bioassays and the technological pharmacology facilities ARPEGE (Pharmacology Screening Interactome) of Biocampus (IGF). This work has been supported by CisBio (cooperative research team Eidos, CNRS N°039293/CBB DRD-09-04 avenant 4), the Labex EpiGenMed, an “Investissements d’avenir” program (ANR-10-LABX-12-01), the Fondation pour la Recherche Médicale (DEQ20170336747), CNRS, Inserm, and the University of Montpellier.

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Authors and Affiliations

  1. IGF, Univ Montpellier, CNRS, INSERM, Montpellier, France

    Diana Zindel, Claire Vol, Odile Lecha, Isabelle Bequignon, Merve Bilgic, Marion Vereecke, Jean-Philippe Pin, Julie Pannequin & Laurent Prézeau

  2. Cisbio Bioassays, F-30200, Codolet, France

    Fabienne Charrier-Savournin, Maïté Romier, Eric Trinquet, Thomas Roux & Elodie Dupuis

Authors
  1. Diana Zindel
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  2. Claire Vol
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  3. Odile Lecha
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  4. Isabelle Bequignon
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  5. Merve Bilgic
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  6. Marion Vereecke
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  7. Fabienne Charrier-Savournin
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  8. Maïté Romier
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  9. Eric Trinquet
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  10. Jean-Philippe Pin
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  11. Julie Pannequin
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  12. Thomas Roux
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  13. Elodie Dupuis
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  14. Laurent Prézeau
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Corresponding author

Correspondence to Laurent Prézeau .

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Editors and Affiliations

  1. Cancer Institute, University College London, London, UK

    Alexander Hergovich

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Zindel, D. et al. (2019). HTRF® Total and Phospho-YAP (Ser127) Cellular Assays. In: Hergovich, A. (eds) The Hippo Pathway. Methods in Molecular Biology, vol 1893. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8910-2_13

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  • DOI: https://doi.org/10.1007/978-1-4939-8910-2_13

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8909-6

  • Online ISBN: 978-1-4939-8910-2

  • eBook Packages: Springer Protocols

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