title: Impact of fusion to Gα(i2) and co-expression with RGS proteins on pharmacological properties of human cannabinoid receptors CB₁R and CB₂R.
abstract: OBJECTIVE
  G protein coupled receptor (GPCR)-Gα fusion proteins are often employed to investigate receptor/G protein interaction. In this study, the impact of Gα fusion proteins on pharmacology of CBRs, both mediating signals through Gα(i) proteins, were investigated. Gα(i2) was fused to the C-terminus of the CBRs or co-expressed with non-fused Gα(i2) in Sf9 cells, always together with Gβ₁γ₂. Furthermore, the impact of RGS proteins on CBR signaling in combination with the CBR fusion approach was examined, using RGS4 and RGS19 as paradigms.
  
  
  METHODS
  CBR ligands were characterized in the steady-state GTPase assay and pharmacological properties of ligands in the different test systems were correlated.
  
  
  RESULTS
  Fusion of CBRs to Gα(i2) enhanced the maximal stimulatory effects of ligands compared to the co-expression system, especially for CB₂R. RGS4, but not RGS19, behaved as a GTPase-activating protein at CBRs in the Gα(i2) co-expression and fusion system. Fusion of GPCR, most prominently CB₂R, to Gα(i2) , and co-expression with RGS4 altered the pharmacological properties of ligands.
  
  
  CONCLUSIONS
  Our data suggest that fusion of CB₂R to Gα(i2) and co-expression with RGS4 impedes with conformational changes. Moreover, our results support the concept of ligand-specific receptor conformations. Finally, this paper describes the most sensitive CBR test system currently available.
authors: Department of Pharmaceutical Biology, Institute of Pharmacy, University of Regensburg, Regensburg Institute of Pharmacology, Medical School of Hannover, Hannover, Germany.
authors: Sarah
authors: S
authors: Sutor
authors: Jörg
authors: J
authors: Heilmann
authors: Roland
authors: R
authors: Seifert
mesh_terms: D002460:Cell Line; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D011485:Protein Binding; D020710:RGS Proteins / Q000378:metabolism*; D043882:Receptors, Cannabinoid / Q000737:chemistry / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: D044385:GTP-Binding Protein alpha Subunits; D008024:Ligands; D020710:RGS Proteins; D043882:Receptors, Cannabinoid; D043562:Receptors, G-Protein-Coupled; D011993:Recombinant Fusion Proteins; D020558:GTP Phosphohydrolases