Abstract
Peroxisome proliferator-activated receptors are a family of nuclear hormone receptors that control the expression of genes involved in a variety of physiologic processes, through heterodimerization with retinoid X receptor and complex formation with various cofactors. The specific cofactors recruited to PPAR-RXR complexes in response to different ligands lead to major differences in the transactivation of target genes. We developed a cofactor recruitment assay that is based on an europium-labeled anti-GST antibody and streptavidin-APC leading to a fluorescence resonance energy transfer signal. This assay allows for the determination of unique agonistic profiles in terms of potency and co-activator motif. Hence, it is a valuable drug discovery tool to support hit finding and lead optimization campaigns, enabling the characterization of next generation PPAR agonists.
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References
Berger J, Moller DE (2002) The mechanisms of action of PPARs. Annu Rev Med 53:409–435
Botta M, Audano M, Sahebkar A, Sirtori CR, Mitro N, Ruscica M (2018) PPAR agonists and metabolic syndrome: an established role? Int J Mol Sci 19(4):1197
Burns KA, Van den Heuvel JP (2007) Modulation of PPAR activity via phosphorylation. Biochim Biophys Acta Mol Cell Biol Lipids 1771:952–960
Kodera Y, Takeyama K, Murayama A, Suzawa M, Masuhiro Y, Kato S (2000) Ligand type-specific interactions of peroxisome proliferator-activated receptor gamma with transcriptional co-activators. J Biol Chem 275:33201–33204
Shao G, Heyman RA, Schulman IG (2000) Three amino acids specify co-activator choice by retinoid X receptors. Mol Endocrinol 14:1198–1209
Picard F, Auwerx J (2002) PPAR(gamma) and glucose homeostasis. Annu Rev Nutr 22:167–197
Yamauchi T, Oike Y, Kamon J, Waki H, Komeda K, Tsuchida A, Date Y, Li MX, Miki H, Akanuma Y, Nagai R, Kimura S, Saheki T, Nakazato M, Naitoh T, Yamamura K, Kadowaki T (2002) Increased insulin sensitivity despite lipodystrophy in Crebbp heterozygous mice. Nat Genet 30:221–226
Dietz M, Mohr P, Kuhn B, Maerki HP, Hartman P, Ruf A, Benz J, Grether U, Wright MB (2012) Comparative molecular profiling of the PPARα/γ activator aleglitazar: PPAR selectivity, activity and interaction with cofactors. ChemMedChem 7(6):1101–1111
Acknowledgments
Veronique Schirmer is acknowledged for the generation of PPARα/γ cofactor recruitment assay data.
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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
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Roth, D., Benz, J., Grether, U., Dietz, M. (2023). Homogeneous Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Cofactor Recruitment Assay for PPARα and PPARγ. In: Maccarrone, M. (eds) Endocannabinoid Signaling. Methods in Molecular Biology, vol 2576. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2728-0_12
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DOI: https://doi.org/10.1007/978-1-0716-2728-0_12
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Publisher Name: Humana, New York, NY
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Online ISBN: 978-1-0716-2728-0
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