{"doi":"10.1089/can.2021.0223","title":"The NPXXY Motif Regulates β-Arrestin Recruitment by the CB1 Cannabinoid Receptor","abstract":"Background: Activation of signaling effectors by G-protein coupled receptors (GPCRs) depends on different molecular mechanisms triggered by conserved amino acid residues. Although studies have focused on the G-protein signaling state, the mechanism for β-arrestin signaling by CB1 is not yet well defined. Studies have indicated that transmembrane helix 7 (TMH7) and the highly conserved NPXXY motif can be subject to different conformational changes in response to biased ligands and could therefore participate in a molecular mechanism to trigger β-arrestin recruitment. Objective: To investigate the effect of mutations in the NPXXY motif on different signaling pathways activated by the CB1 receptor. Materials and Methods: Point mutations of the NPXXY motif and associated residues were generated in the CB1 receptor using site-directed mutagenesis and transfection into HEK-293 cells. Signaling by wild-type and mutant receptors was analyzed by quantifying inhibition of cAMP, and by β-arrestin recruitment assays. Results: We found that N7.49 and Y7.53 are essential for β-arrestin recruitment by CB1. N7.49A and Y7.53F impair β-arrestin signaling, with no effect on G-protein signaling. We found a regulatory role for residue I2.43; I2.43 interacts with Y7.53, affecting its positioning. Reducing steric bulk at I2.43 (I2.43A) enhances β-arrestin1 recruitment, while introducing a polar residue (I2.43T) reduces β-arrestin recruitment. Conclusions: These findings point to a novel mechanism for β-arrestin recruitment, implicating amino acids in the NPXXY motif as critical for the putative β-arrestin biased conformational state of Class A GPCRs.","journal":"Cannabis and Cannabinoid Research","year":2022,"id":270979,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":13,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9599,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2022-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":935757,"name":"Rufaida Al-Zoubi","orcid":"0000-0001-8417-4769","position":1,"is_corresponding":false},{"id":431619,"name":"Dow P. Hurst","orcid":"0000-0002-2382-0031","position":2,"is_corresponding":false},{"id":936386,"name":"Anna P. Stephan","orcid":null,"position":3,"is_corresponding":false},{"id":790544,"name":"Pingwei Zhao","orcid":null,"position":4,"is_corresponding":false},{"id":422361,"name":"Douglas G. Tilley","orcid":"0000-0002-9681-3377","position":5,"is_corresponding":false},{"id":935758,"name":"Elke Miess","orcid":"0000-0003-3820-6058","position":6,"is_corresponding":false},{"id":443582,"name":"Stefan Schulz","orcid":"0000-0002-5997-8885","position":7,"is_corresponding":false},{"id":634395,"name":"Mary E. Abood","orcid":"0000-0002-1744-2820","position":8,"is_corresponding":false},{"id":432929,"name":"Patricia H. Reggio","orcid":null,"position":9,"is_corresponding":false},{"id":634394,"name":"Luciana M. Leo","orcid":"0000-0002-9676-937X","position":0,"is_corresponding":true}],"reference_count":73,"raw_metadata":null,"created_at":"2026-07-19T00:27:35.206187Z","pmid":"35792570","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}