title: 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.
authors.affiliation: Institut für Biochemie (Emil-Fischer-Zentrum), Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Mascia
authors.affiliation: Institut für Biochemie (Emil-Fischer-Zentrum), Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Lisa
authors.initials: L
authors.lastname: Klotz
authors.affiliation: Institut für Biochemie (Emil-Fischer-Zentrum), Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Judith
authors.initials: J
authors.lastname: Lerch
authors.affiliation: Department of Medicinal Chemistry, Virginia Commonwealth University, Richmond, Virginia, USA.
authors.forename: Mostafa H
authors.initials: MH
authors.lastname: Ahmed
authors.affiliation: Department of Medicinal Chemistry, Virginia Commonwealth University, Richmond, Virginia, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Institut für Biochemie (Emil-Fischer-Zentrum), Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Ralf
authors.identifier: 0000-0001-9219-5295
authors.initials: R
authors.lastname: Enz
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D063385:Cannabinoid Receptor Modulators / Q000494:pharmacology*; D002352:Carrier Proteins / Q000494:pharmacology*; D063388:Endocannabinoids / Q000494:pharmacology*; D004705:Endocytosis / Q000187:drug effects*; D005971:Glutamates / Q000494:pharmacology*; D057809:HEK293 Cells; D006801:Humans; D006860:Hydrogen Bonding; D008565:Membrane Proteins; D051379:Mice; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D011487:Protein Conformation; D017207:Rats, Sprague-Dawley; D043884:Receptor, Cannabinoid, CB1 / Q000187:drug effects / Q000235:genetics; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors*; D018094:Receptors, Metabotropic Glutamate / Q000378:metabolism; D013045:Species Specificity
chemical_list: C000618264:CNRIP1 protein, human; D063385:Cannabinoid Receptor Modulators; D002352:Carrier Proteins; D063388:Endocannabinoids; D005971:Glutamates; D008565:Membrane Proteins; D043884:Receptor, Cannabinoid, CB1; D043562:Receptors, G-Protein-Coupled; D018094:Receptors, Metabotropic Glutamate
keywords: CB1R; CRIP1a; endocannabinoid receptor; endocytosis; mGlu8R; metabotropic glutamate receptor