CRIP1a inhibits endocytosis of G-protein coupled receptors activated by endocannabinoids and glutamate by a common molecular mechanism

Abstract

The excitability of the central nervous system depends largely on the surface density of neurotransmitter receptors. The endocannabinoid receptor 1 (CB1R) and the metabotropic glutamate receptor mGlu8R are expressed pre-synaptically where they reduce glutamate release into the synaptic cleft. Recently, the CB1R interacting protein cannabinoid receptor interacting protein 1a (CRIP1a) was identified and characterized to regulate CB1R activity in neurons. However, underlying molecular mechanisms are largely unknown. Here, we identified a common mechanism used by CRIP1a to regulate the cell surface density of two different types of G-protein coupled receptors, CB1R and mGlu8aR. Five amino acids within the CB1R C-terminus were required and sufficient to reduce constitutive CB1R endocytosis by about 72% in the presence of CRIP1a. Interestingly, a similar sequence is present in mGlu8aR and consistently, endocytosis of mGlu8aR depended on CRIP1a, as well. Docking analysis and molecular dynamics simulations identified a conserved serine in CB1R (S468) and mGlu8aR (S894) that forms a hydrogen bond with the peptide backbone of CRIP1a at position R82. In contrast to mGlu8aR, the closely related mGlu8bR splice-variant carries a lysine (K894) at this position, and indeed, mGlu8bR endocytosis was not affected by CRIP1a. Chimeric constructs between CB1R, mGlu8aR, and mGlu8bR underline the role of the identified five CRIP1a sensitive amino acids. In summary, we suggest that CRIP1a negatively regulates endocytosis of two different G-protein coupled receptor types, CB1R and mGlu8aR. (Figure presented.).