Cannabinoid receptor interacting protein suppresses agonist-driven CB₁ receptor internalization and regulates receptor replenishment in an agonist-biased manner

详细信息    查看全文

• 作者：Lawrence C. Blume ; Sandra Leone-Kabler ; Deborah J. Luessen ; Glen S. Marrs ; Erica Lyons ; Caroline E. Bass ; Rong Chen ; Dana E. Selley and Allyn C. Howlett
• 刊名：Journal of Neurochemistry
• 出版年：2016
• 出版时间：November 2016
• 年：2016
• 卷：139
• 期：3
• 页码：396-407
• 全文大小：1453K
• ISSN：1471-4159

文摘

Cannabinoid receptor interacting protein 1a (CRIP1a) is a CB₁ receptor (CB₁R) distal C-terminus-associated protein that modulates CB₁R signaling via G proteins, and CB₁R down-regulation but not desensitization (Blume et al. [2015] Cell Signal., 27, 716–726; Smith et al. [2015] Mol. Pharmacol., 87, 747–765). In this study, we determined the involvement of CRIP1a in CB₁R plasma membrane trafficking. To follow the effects of agonists and antagonists on cell surface CB₁Rs, we utilized the genetically homogeneous cloned neuronal cell line N18TG2, which endogenously expresses both CB₁R and CRIP1a, and exhibits a well-characterized endocannabinoid signaling system. We developed stable CRIP1a-over-expressing and CRIP1a-siRNA-silenced knockdown clones to investigate gene dose effects of CRIP1a on CB₁R plasma membrane expression. Results indicate that CP55940 or WIN55212-2 (10 nM, 5 min) reduced cell surface CB₁R by a dynamin- and clathrin-dependent process, and this was attenuated by CRIP1a over-expression. CP55940-mediated cell surface CB₁R loss was followed by a cycloheximide-sensitive recovery of surface receptors (30–120 min), suggesting the requirement for new protein synthesis. In contrast, WIN55212-2-mediated cell surface CB₁Rs recovered only in CRIP1a knockdown cells. Changes in CRIP1a expression levels did not affect a transient rimonabant (10 nM)-mediated increase in cell surface CB₁Rs, which is postulated to be as a result of rimonabant effects on ‘non-agonist-driven’ internalization. These studies demonstrate a novel role for CRIP1a in agonist-driven CB₁R cell surface regulation postulated to occur by two mechanisms: 1) attenuating internalization that is agonist-mediated, but not that in the absence of exogenous agonists, and 2) biased agonist-dependent trafficking of de novo synthesized receptor to the cell surface.