The cannabinoid receptor interacting protein CRIP1 binds to glutamate receptors in the CNS: Molecular mechanisms and functional implications

谷氨酸是中枢神经系统（CNS）中最重要的兴奋性神经递质，并激活G蛋白偶联（代谢型）谷氨酸受体（mGluR 1-mGluR 8）。此外，内源性大麻素调节G蛋白偶联受体（CB 1和CB 2），这些受体调节神经元活动并负责大麻的精神病效应。哺乳动物视网膜的光感受器在突触前共表达mGluR 8和CB 1，它们都是调节从光感受器末端释放谷氨酸的负反馈环的一部分。通过这种方式，mGluR 8和CB 1参与调节视网膜在大范围照明下的敏感性的适应性机制。不同神经递质受体类别之间的串扰代表了协调神经元中信号通路的关键机制。CB 1和mGluR类型之间的功能相互作用的报告。此外，腺苷和5-羟色胺的受体与CB 1或mGluR 2形成异源二聚体，并证明了由腺苷、多巴胺、谷氨酸和内源性大麻素受体以不同组合组成的异源三聚体受体复合物的形成。此外，细胞内支架蛋白可以物理连接功能连接的受体types.Recently，我们确定了大麻素受体相互作用蛋白1a CRIP 1a作为mGluR 8a亚型的一个新的结合伴侣。因此，我们建议mGluR 8a和CB 1之间通过CRIP 1的功能/物理相互作用。该假说基于（i）CRIP 1a作为mGluR 8a相互作用物的鉴定，（ii）CRIP 1a、CB 1和mGluR 8a在突触前光感受器末梢中的共表达，（iii）CB 1和mGluR 8a相似的生理功能，（抑制突触前谷氨酸释放），（iv）在CB 1 C-末端中的映射的9个氨基酸长的CRIP 1a结合序列与mGluR 8a C-末端中的6个氨基酸的线性延伸之间的高度同源性，以及（v）最后，根据已知的生物学原理，即受体串扰可以通过细胞内蛋白发生，我们计划阐明介导已鉴定的mGluR 8a/CRIP 1a相互作用的分子机制，以及阐明mGluR 8a和CB 1在CNS中的物理/功能串扰。我们的工作计划将首先分析mGluR 8a，CB 1和CRIP 1a之间结合的分子机制，从而为后续功能研究提供分子工具和机制知识。这些数据将通过解剖学共定位实验进行补充。此后，我们计划调查新发现的蛋白质相互作用的功能后果，首先使用重组表达的蛋白质在HEK-293细胞，然后在photoreceptors.我们的研究旨在描述新的原则，在中枢神经系统的神经递质受体功能，并将具体阐明在哺乳动物视网膜的第一个视觉突触的信号调节的分子机制。

Glutamate is the most important excitatory neurotransmitter in the central nervous system (CNS) and activates G-protein coupled (metabotropic) glutamate receptors (mGluR1-mGluR8). Also endocannabinoids regulate G-protein coupled receptors (CB1 and CB2) that regulate neuronal activity and are responsible for psychotic effects of marijuana. Photoreceptors of the mammalian retina co-express pre-synaptically mGluR8 and CB1 that are both part of negative feedback loops regulating glutamate release from photoreceptor terminals. In this way, mGluR8 and CB1 participate in adaptive mechanisms that regulate the sensitivity of the retina over a large range of illumination.Crosstalk between different neurotransmitter receptor classes represents a key mechanism to coordinate signal pathways in neurons. Functional interactions between CB1 and mGluR types were reported. In addition, receptors for adenosine and serotonin form heterodimers with CB1 or mGluR2, and the formation of heterotrimeric receptor complexes composed of adenosine, dopamine, glutamate and endocannabinoid receptors in different combinations was demonstrated. In addition, intracellular scaffold proteins can physically link functionally connected receptor types.Recently, we identified the cannabinoid receptor interacting protein 1a CRIP1a as a new binding partner of the mGluR8a isoform. Therefore, we suggest a functional/physical interaction between mGluR8a and CB1 via CRIP1. This hypothesis is based on (i) the identification of CRIP1a as a mGluR8a interactor, (ii) the co-expression of CRIP1a, CB1 and mGluR8a in pre-synaptic photoreceptor terminals, (iii) similar physiological functions of CB1 and mGluR8a (inhibition of the pre-synaptic glutamate release), (iv) a high homology between the mapped 9 amino acid long CRIP1a binding sequence in the CB1 C-terminus and a linear stretch of 6 amino acids in the mGluR8a C-terminus and (v) finally on the known biological principle that receptor crosstalk can occur via intracellular proteins.We plan to characterise molecular mechanisms that mediate the identified mGluR8a/CRIP1a interaction, as well as to elucidate a suggested physical/functional crosstalk between mGluR8a and CB1 in the CNS. Our working programme will first analyse molecular mechanisms of the binding between mGluR8a, CB1 and CRIP1a, thereby providing molecular tools and mechanistic knowledge for successive functional studies. These data will be complemented by anatomical co-localization experiments. Thereafter, we plan to investigate functional consequences of the newly identified protein interactions, first using recombinantly expressed proteins in HEK-293 cells and thereafter in photoreceptors.Our studies are designed to describe new principles of neurotransmitter receptor function in the CNS and will specifically elucidate molecular mechanisms that regulate signalling at the first visual synapse in the mammalian retina.