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Co-immunoprecipitation (Co-IP) of G Protein-Coupled Receptor (GPCR)-Receptor Tyrosine Kinase (RTK) Complexes from the Dorsal Hippocampus of the Rat Brain

  • Michael Di Palma
  • Manuel Narvaez
  • Mariana Pita-Rodríguez
  • Chiara Contri
  • Martina Zannoni
  • Riccardo Cuppini
  • Kjell Fuxe
  • Patrizia Ambrogini
  • Dasiel O. Borroto-Escuela
Protocol
Part of the Neuromethods book series (NM, volume 144)

Abstract

A large body of evidence indicates that G protein-coupled receptors (GPCRs) and receptor tyrosine kinases (RTKs) can form heteroreceptor complexes. In these complexes, the signaling from each interacting protomer is modulated to produce an integrated and therefore novel response upon agonist(s) activation. In the GPCR-RTK heteroreceptor complexes, GPCRs can activate RTK in the absence of added growth factor through the use of RTK signaling molecules. This integrative phenomenon is reciprocal and can place also RTK signaling downstream of GPCR. The existence of either stable or transient GPCR-RTK homo- and heteroreceptor complexes with allosteric receptor-receptor interactions increases the diversity of receptor function including recognition, trafficking, and signaling. The isolation and characterization of GPCR-RTK heteroreceptor complexes are therefore important to understand these processes. Co-immunoprecipitation (Co-IP) is a straightforward technique to study in vivo GPCR-RTK interactions and can identify interacting proteins or heteroreceptor complexes present in cell extracts. Here, we present detailed protocol for Co-IP of GPCR-RTK heteroreceptor complexes from brain membrane preparations using as an example the study of A2AR-TrkB heteroreceptor complexes in the rat dorsal hippocampus.

Key words

G protein-coupled receptors (GPCRs) Receptor tyrosine kinases (RTKs) Heteroreceptor complexes GPCR-RTK heteroreceptor complexes Receptor-receptor interactions Homodimerization Heterodimerization Co-immunoprecipitation 

Notes

Acknowledgments

This work has been supported by the Karolinska Institutets Forskningsstiftelser 2017 to D.O.B-E, the Swedish Medical Research Council (62X-00715-50-3), and Hjärnfonden 2018 to D.O.B-E. D.O.B-E belongs to Academia de Biólogos Cubanos.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Michael Di Palma
    • 1
  • Manuel Narvaez
    • 2
  • Mariana Pita-Rodríguez
    • 2
    • 3
  • Chiara Contri
    • 4
    • 5
  • Martina Zannoni
    • 4
    • 5
  • Riccardo Cuppini
    • 1
  • Kjell Fuxe
    • 5
  • Patrizia Ambrogini
    • 1
  • Dasiel O. Borroto-Escuela
    • 5
    • 6
  1. 1.Section of Physiology, Department of Biomolecular ScienceUniversity of UrbinoUrbinoItaly
  2. 2.Facultad de Medicina, Instituto de Investigación Biomédica de MálagaUniversidad de MálagaMálagaSpain
  3. 3.Neurogenetics DepartmentInstitute of Neurology and NeurosurgeryHavanaCuba
  4. 4.Department of Life Science and BiotechnologyUniversity of FerraraFerraraItaly
  5. 5.Department of NeuroscienceKarolinska InstitutetStockholmSweden
  6. 6.Observatorio Cubano de NeurocienciasYaguajayCuba

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