title: PSD-95 regulates CRFR1 localization, trafficking and β-arrestin2 recruitment.
abstract: Corticotropin-releasing factor (CRF) is a neuropeptide commonly associated with the hypothalamic-pituitary adrenal axis stress response. Upon release, CRF activates two G protein-coupled receptors (GPCRs): CRF receptor 1 (CRFR1) and CRF receptor 2 (CRFR2). Although both receptors contribute to mood regulation, CRFR1 antagonists have demonstrated anxiolytic and antidepressant-like properties that may be exploited in the generation of new pharmacological interventions for mental illnesses. Previous studies have demonstrated CRFR1 capable of heterologously sensitizing serotonin 2A receptor (5-HT2AR) signaling: another GPCR implicated in psychiatric disease. Interestingly, this phenomenon was dependent on Postsynaptic density 95 (PSD-95)/Disc Large/Zona Occludens (PDZ) interactions on the distal carboxyl termini of both receptors. In the current study, we demonstrate that endogenous PSD-95 can be co-immunoprecipitated with CRFR1 from cortical brain homogenate, and this interaction appears to be primarily via the PDZ-binding motif. Additionally, PSD-95 colocalizes with CRFR1 within the dendritic projections of cultured mouse neurons in a PDZ-binding motif-dependent manner. In HEK 293 cells, PSD-95 overexpression inhibited CRFR1 endocytosis, whereas PSD-95 shRNA knockdown enhanced CRFR1 endocytosis. Although PSD-95 does not appear to play a significant role in CRF-mediated cAMP or ERK1/2 signaling, PSD-95 was demonstrated to suppress β-arrestin2 recruitment: providing a potential mechanism for PSD-95's inhibition of endocytosis. In revisiting previously documented heterologous sensitization, PSD-95 shRNA knockdown did not prevent CRFR1-mediated enhancement of 5-HT2AR signaling. In conclusion, we have identified and characterized a novel functional relationship between CRFR1 and PSD-95 that may have implications in the design of new treatment strategies for mental illness.
authors.affiliation: J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, and the Department of Physiology and Pharmacology, University of Western Ontario, 100 Perth Dr. London, Ontario N6A 5K8, Canada; Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, 451 Smyth Dr. Ottawa, Ontario K1H 8M5, Canada.
authors.forename: Henry A
authors.initials: HA
authors.lastname: Dunn
authors.affiliation: J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, and the Department of Physiology and Pharmacology, University of Western Ontario, 100 Perth Dr. London, Ontario N6A 5K8, Canada.
authors.forename: Harpreet S
authors.initials: HS
authors.lastname: Chahal
authors.affiliation: J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, and the Department of Physiology and Pharmacology, University of Western Ontario, 100 Perth Dr. London, Ontario N6A 5K8, Canada.
authors.forename: Fabiana A
authors.initials: FA
authors.lastname: Caetano
authors.affiliation: J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, and the Department of Physiology and Pharmacology, University of Western Ontario, 100 Perth Dr. London, Ontario N6A 5K8, Canada.
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Holmes
authors.affiliation: J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, and the Department of Physiology and Pharmacology, University of Western Ontario, 100 Perth Dr. London, Ontario N6A 5K8, Canada.
authors.forename: George Y
authors.initials: GY
authors.lastname: Yuan
authors.affiliation: J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, and the Department of Physiology and Pharmacology, University of Western Ontario, 100 Perth Dr. London, Ontario N6A 5K8, Canada.
authors.forename: Ruchi
authors.initials: R
authors.lastname: Parikh
authors.affiliation: Department of Microbiology and Immunology, University of Western Ontario, London, Ontario N6A 3K7, Canada.
authors.forename: Bryan
authors.initials: B
authors.lastname: Heit
authors.affiliation: Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, 451 Smyth Dr. Ottawa, Ontario K1H 8M5, Canada. Electronic address: sferguso@uottawa.ca.
authors.forename: Stephen S G
authors.initials: SSG
authors.lastname: Ferguson
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D000076122:Disks Large Homolog 4 Protein; D004705:Endocytosis; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D051528:Guanylate Kinases / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D057907:Post-Synaptic Density / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D021381:Protein Transport; D018019:Receptors, Corticotropin-Releasing Hormone / Q000737:chemistry / Q000378:metabolism*; D015398:Signal Transduction; D000071560:beta-Arrestin 2 / Q000378:metabolism*; D000099052:CRF Receptor, Type 1
chemical_list: D000076122:Disks Large Homolog 4 Protein; C501447:Dlg4 protein, mouse; D008565:Membrane Proteins; D018019:Receptors, Corticotropin-Releasing Hormone; D000071560:beta-Arrestin 2; D000099052:CRF Receptor, Type 1; D000242:Cyclic AMP; D048049:Extracellular Signal-Regulated MAP Kinases; D051528:Guanylate Kinases
keywords: Arrestin; CRFR1; GPCR; PDZ; PSD-95; Trafficking