title: Cannabinoid receptor interacting protein (CRIP1a) attenuates CB1R signaling in neuronal cells.
abstract: CB1 cannabinoid receptors (CB1R) are one of the most abundantly expressed G protein coupled receptors (GPCR) in the CNS and regulate diverse neuronal functions. The identification of GPCR interacting proteins has provided additional insight into the fine-tuning and regulation of numerous GPCRs. The cannabinoid receptor interacting protein 1a (CRIP1a) binds to the distal carboxy terminus of CB1R, and has been shown to alter CB1R-mediated neuronal function [1]. The mechanisms by which CRIP1a regulates CB1R activity have not yet been identified; therefore the focus of this investigation is to examine the cellular effects of CRIP1a on CB1R signaling using neuronal N18TG2 cells stably transfected with CRIP1a over-expressing and CRIP1a knockdown constructs. Modulation of endogenous CRIP1a expression did not alter total levels of CB1R, ERK, or forskolin-activated adenylyl cyclase activity. When compared to WT cells, CRIP1a over-expression reduced basal phosphoERK levels, whereas depletion of CRIP1a augmented basal phosphoERK levels. Stimulation of phosphoERK by the CB1R agonists WIN55212-2, CP55940 or methanandamide was unaltered in CRIP1a over-expressing clones compared with WT. However, CRIP1a knockdown clones exhibited enhanced ERK phosphorylation efficacy in response to CP55940. In addition, CRIP1a knockdown clones displayed a leftward shift in CP55940-mediated inhibition of forskolin-stimulated cAMP accumulation. CB1R-mediated Gi3 and Go activation by CP99540 was attenuated by CRIP1a over-expression, but robustly enhanced in cells depleted of CRIP1a. Conversely, CP55940-mediated Gi1 and Gi2 activation was significant enhanced in cells over-expressing CRIP1a, but not in cells deficient of CRIP1a. These studies suggest a mechanism by which endogenous levels of CRIP1a modulate CB1R-mediated signal transduction by facilitating a Gi/o protein subtype preference for Gi1 and Gi2, accompanied by an overall suppression of G-protein-mediated signaling in neuronal cells.
authors.affiliation: Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 USA.
authors.forename: Lawrence C
authors.initials: LC
authors.lastname: Blume
authors.affiliation: Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 USA.
authors.forename: Khalil
authors.initials: K
authors.lastname: Eldeeb
authors.affiliation: Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 USA.
authors.forename: Caroline E
authors.initials: CE
authors.lastname: Bass
authors.affiliation: Department of Pharmacology and Toxicology, Virginia Commonwealth University, Richmond, VA 23298 USA.
authors.forename: Dana E
authors.initials: DE
authors.lastname: Selley
authors.affiliation: Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 USA.
authors.forename: Allyn C
authors.initials: AC
authors.lastname: Howlett
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D000242:Cyclic AMP / Q000378:metabolism; D003511:Cyclohexanols / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D010880:Piperidines / Q000494:pharmacology; D011720:Pyrazoles / Q000494:pharmacology; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D043884:Receptor, Cannabinoid, CB1 / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D000077285:Rimonabant; D015398:Signal Transduction
chemical_list: C000602666:CRIP1a protein, mouse; D002352:Carrier Proteins; D003511:Cyclohexanols; D010880:Piperidines; D011720:Pyrazoles; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D043884:Receptor, Cannabinoid, CB1; C054649:3-(2-hydroxy-4-(1,1-dimethylheptyl)phenyl)-4-(3-hydroxypropyl)cyclohexanol; D000242:Cyclic AMP; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D000077285:Rimonabant
keywords: CB(1) receptor; CRIP1a; ERK1/2; Endocannabinoid; G protein; cAMP