The second intracellular loop of the human cannabinoid CB2 receptor governs G protein coupling in coordination with the carboxyl terminal domain

PLoS One. 2013 May 7;8(5):e63262. doi: 10.1371/journal.pone.0063262. Print 2013.


The major effects of cannabinoids and endocannabinoids are mediated via two G protein-coupled receptors, CB1 and CB2, elucidation of the mechanism and structural determinants of the CB2 receptor coupling with G proteins will have a significant impact on drug discovery. In the present study, we systematically investigated the role of the intracellular loops in the interaction of the CB2 receptor with G proteins using chimeric receptors alongside the characterization of cAMP accumulation and ERK1/2 phosphorylation. We provided evidence that ICL2 was significantly involved in G protein coupling in coordination with the C-terminal end. Moreover, a single alanine substitution of the Pro-139 in the CB2 receptor that corresponds to Leu-222 in the CB1 receptor resulted in a moderate impairment in the inhibition of cAMP accumulation, whereas mutants P139F, P139M and P139L were able to couple to the Gs protein in a CRE-driven luciferase assay. With the ERK activation experiments, we further found that P139L has the ability to activate ERK through both Gi- and Gs-mediated pathways. Our findings defined an essential role of the second intracellular loop of the CB2 receptor in coordination with the C-terminal tail in G protein coupling and receptor activation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenylyl Cyclase Inhibitors
  • Adenylyl Cyclases / metabolism
  • Amino Acid Sequence
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Enzyme Activation / drug effects
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • GTP-Binding Proteins / metabolism*
  • HEK293 Cells
  • Humans
  • Molecular Sequence Data
  • Mutant Proteins / chemistry
  • Mutant Proteins / metabolism
  • Proline / metabolism
  • Protein Binding / drug effects
  • Protein Kinase Inhibitors / pharmacology
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Receptor, Cannabinoid, CB1 / chemistry
  • Receptor, Cannabinoid, CB1 / metabolism
  • Receptor, Cannabinoid, CB2 / agonists
  • Receptor, Cannabinoid, CB2 / chemistry*
  • Receptor, Cannabinoid, CB2 / metabolism*
  • Recombinant Proteins / metabolism
  • Signal Transduction* / drug effects
  • Structure-Activity Relationship


  • Adenylyl Cyclase Inhibitors
  • Mutant Proteins
  • Protein Kinase Inhibitors
  • Receptor, Cannabinoid, CB1
  • Receptor, Cannabinoid, CB2
  • Recombinant Proteins
  • Proline
  • Cyclic AMP-Dependent Protein Kinases
  • Extracellular Signal-Regulated MAP Kinases
  • GTP-Binding Proteins
  • Adenylyl Cyclases