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

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 mGlu(8)R 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 mGlu(8a)R. 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 mGlu(8a)R and consistently, endocytosis of mGlu(8a)R depended on CRIP1a, as well. Docking analysis and molecular dynamics simulations identified a conserved serine in CB1R (S468) and mGlu(8a)R (S894) that forms a hydrogen bond with the peptide backbone of CRIP1a at position R82. In contrast to mGlu(8a)R, the closely related mGlu(8b)R splice-variant carries a lysine (K894) at this position, and indeed, mGlu(8b)R endocytosis was not affected by CRIP1a. Chimeric constructs between CB1R, mGlu(8a)R, and mGlu(8b)R 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 mGlu(8a)R.