Cell Type-Specific Tandem Affinity Purification of the Mouse Hippocampal CB1 Receptor-Associated Proteome.
G protein coupled receptors (GPCRs) exert their effects through multiprotein signaling complexes. The cannabinoid receptor type 1 (CB1) is among the most abundant GPCRs in the mammalian brain and involved in a plethora of physiological functions. We used a combination of viral-mediated cell type-specific expression of a tagged CB1 fusion protein (CB1-SF), tandem affinity purification (TAP) and proteomics on hippocampal mouse tissue to analyze the composition and differences of CB1 protein complexes in glutamatergic neurons and in GABAergic interneurons. Purified proteins underwent tryptic digestion and were identified using deep-coverage data-independent acquisition with ion mobility separation-enhanced mass spectroscopy, leading to the identification of 951 proteins specifically enriched in glutamatergic and GABAergic CB1-SF TAP samples as compared to controls. Gene Ontology and protein network analyses showed an enrichment of single proteins and functional clusters of proteins involved in already well described domains of CB1 functions. Supported by this consistent data set we could confirm already known CB1 interactors, reveal new potentially interacting proteins and differences in cell type-specific signaling properties of CB1, thereby providing the foundation for further functional studies on differential CB1 signaling.
Institute of Physiological Chemistry, University Medical Center of the Johannes Gutenberg-University Mainz , Duesbergweg 6, 55128 Mainz, Germany.
Tobias
T
Mattheus
Institute of Physiological Chemistry, University Medical Center of the Johannes Gutenberg-University Mainz , Duesbergweg 6, 55128 Mainz, Germany.
Katharina
K
Kukla
Institute of Physiological Chemistry, University Medical Center of the Johannes Gutenberg-University Mainz , Duesbergweg 6, 55128 Mainz, Germany.
Tina
T
Zimmermann
Institute for Immunology, University Medical Center of the Johannes Gutenberg-University Mainz , Langenbeckstrasse 1, 55131 Mainz, Germany.
Stefan
S
Tenzer
Institute of Physiological Chemistry, University Medical Center of the Johannes Gutenberg-University Mainz , Duesbergweg 6, 55128 Mainz, Germany.
Beat
B
Lutz
D000818:Animals; D002846:Chromatography, Affinity; D018698:Glutamic Acid / Q000378:metabolism; D006624:Hippocampus / Q000737:chemistry*; D051379:Mice; D009474:Neurons / Q000737:chemistry; D011485:Protein Binding; D060066:Protein Interaction Maps; D020543:Proteome / Q000032:analysis*; D043884:Receptor, Cannabinoid, CB1 / Q000378:metabolism*; D015398:Signal Transduction; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D020543:Proteome; D043884:Receptor, Cannabinoid, CB1; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid
CB1; ECS; GPCR; TAP; cannabinoid receptor type 1; endocannabinoid system; protein interactions; proteomics; signaling; tandem affinity purification