Peroxisome Proliferator-Activated Receptors (PPARs) Methods and Protocols

All three peroxisome proliferator-activated receptor (PPAR) subtypes share a high degree of structural homology but differ in function, tissue distribution and ligand specificity. PPARs play critical roles as regulators of numerous physiological as well as pathophysiological pathways, and efforts are currently underway to fully characterize their functioning and to develop safer and more effective PPAR modulators to treat a myriad of diseases and conditions. In Peroxisome proliferator-Activated Receptors: Methods and Protocols, renowned experts in the PPAR arena provide detailed protocols for investigating these receptors. Chapters contain methods ranging from the cloning of receptors to their knockdown, to protocols exploring posttranslational modifications of PPARs and coactivators, as well as receptor subcellular localization. Also assembled are methods to evaluate the involvement of these receptors in behavior functions, an emerging facet in PPAR research. Written in the successful Methods in Molecular Biology™ series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls.

With its well-honed methodologies, Peroxisome proliferator-Activated Receptors: Methods and Protocols,  will be a useful resource for all seeking to advance their knowledge of this field.

1. PPARs: History and Advances

            Jihan Youssef and Mostafa Badr

2. PPAR-Alpha Cloning, Expression and Characterization

            Suong N. T. Ngo and Ross McKinnon

3. PPARGC1A and PPARG Genotyping. Beginner’s Guide to Genotyping with Unlabeled Probes

            Viera Habalova and Josef Židzik

4. Generation of an Inducible, Cardiomyocyte-Specific Transgenic Mouse Model with PPARß/d Overexpression

Teayoum Kim, Olga Zhelyabovska, Jian Liu, and Qinglin Yang

5. Specific knockdown of PPARd Gene in Colon Cancer Cells by Lentivirus-Mediated RNA Interfering

            Lie Yang, Zong-Guang Zhou, and Xiao-Feng Sun

6. Dominant-Negative and Knockdown Approaches to Studying PPAR activity

            Sudhir Singh and Robert G. Bennett

7. Producing PPARgamma2 Knockdown in Mouse Liver

            Tomomi Yamazaki and Osamu Ezaki

8. Adipose Tissue-Specific PPARg Gene Targeting

            Weimin He

9. Site-Directed Mutagenesis to Study the Role of Specific Amino Acids in the Ligand Binding Domain of PPARs

            Nico Mitro, Frederica Gilardi, Marco Giudici, Cristina Godio, Elena Scotti, and Maurizio Crestani

10. PPAR SUMOylation: Some Useful Experimental Tips

            Benoit Pourcet, Bart Staels, and Corine Glineur

11. Analyzing Phosphorylation-Dependent Regulation of Subcellular Transcriptional Activity of Transcriptional Coactivator NT-PGC-1a

            Ji Suk Chang and Thomas W. Gettys

12. In vivo Studies of PPAR-Chromatin Interactions: Chromatin Immunoprecipitation for Single-Locus and Genomewide Analyses

            Till Adhikary and Rolf Müller

13. FISH Analysis Using PPARg  Specific Probes for Detection of PAX8-PPARg Translocation in Follicular Thyroid Neoplasms

            Sharifah Noor Akmal Syed Hussain, Chia Wai Kit, and Zubaidah Zakaraia

14. Immunohistochemical Techniques to Identify and Localize Proteins of Interest in Paraffin-Embedded Tissue Sections

            Carolyn M. Komar and Mary Jane Long

15. Determination of PPAR Expression by Western-Blot

            Aziz Gullich and Catherine Coirault

16. Fluorescence Resonance Energy Transfer Techniques to Study Ligand-Mediated Interactions of PPARs with Coregulators

            Nico Mitro, Cristina Godio, and Maurizio Crestani

17. Estimation of the PPARa Agonism of Fibrates by a Combined MM-Docking Approach

            Fabio Lannutti, Alessandro Marrone, and Nazzareno Re

18. Combined Biophysical and Cell-based Approaches for the Assessment of Ligand Binding to PPARg

            Silvia Zorrilla and Dolores Pérez-Sala

19. Exploring PPAR Modulation in Experimental Mice

            Anna Rull, Gerard Aragonès, Raúl Beltrán-Debón, Esther Rodríguez-Gallego, Jordi Camps, Jorge Joven

20. Induction of Adipogenic Differentiation in Three-Dimensional Culture Model on a Novel Microfabricated Scaffold

            Yoshitaka Miyagawa, Hajime Okita, and Nobutaka Kiyokawa

21. Analyzing PPAa/Ligand Interactions by Chemical Cross-Linking and High-Resolution Mass Spectrometry

            Mathias Muller, Christian Ihling, and Andrea Sinz

22. Synthesis, Mass Spectrometric Characterization, and Analysis of the PPARd Agonist GW1516 and its Major Human Metabolites – Targets in Sports Drug Testing

            Mario Thevis, Ines Möller, Simon Beuk, and Wihelm Schänzer

23. LC-MS-Based Method for the Qualitative and Quantitative Analysis of the Novel PPARg Agonist KR-62980

            Mi-Sun Kim and Myung Ae Bae

24. Behavioral Paradigms to Evaluate PPAR Modulation in Animal Models of Brain   Injury

            Dana Greene-Schloesser, Caroline I. Schnegg, and Mike E. Robbins