The CNTNAP2-CASK complex modulates GluA1 subcellular distribution in interneurons
The CNTNAP2-CASK complex modulates GluA1 subcellular distribution in interneurons
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DOI：
10.1016/j.neulet.2019.02.025
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发表时间：
2019-05-14
影响因子：
2.5
通讯作者：
Penzes, Peter
中科院分区：
文献类型：
--
作者：
Gao, Ruoqi;Zaccard, Colleen R.;Penzes, Peter
GABA能中间神经元在多种神经发育障碍的病理生理学中是重要的底物，包括自闭症谱系障碍、精神分裂症、智力残疾和癫痫。中间神经元兴奋活性受2-氨基-3-（3-羟基-5-甲基-异恶唑-4-基）丙酸受体（AMPAR）影响，其进而影响中枢神经系统中的兴奋性传递。然而，神经元间AMPAR的调节异常如何明显地促进神经生物学疾病的分子基础是彻底探索不足。接触素相关蛋白样2（CNTNAP 2）是一种神经毒素相关的粘附分子，介导AMPAR的亚细胞分布，而钙/钙调蛋白依赖性丝氨酸蛋白激酶（CASK）是一种多功能支架，参与谷氨酸受体的运输。这两个基因的突变有重叠的疾病关联，包括自闭症谱系障碍，智力残疾和癫痫，从而表明兴奋/抑制平衡的会聚扰动。我们的实验室以前已经表明，CNTNAP 2通过CASK稳定中间神经元树突状动脉，CNTNAP 2调节兴奋性神经元中AMPAR亚基GluA 1的运输。然而，这三种蛋白质之间的相互作用尚未在中间神经元中研究。利用生物化学技术、结构照明显微镜（SIM）技术和shRNA技术，我们首先证实了这三种蛋白在小鼠脑中相互作用，然后研究了CNTNAP 2、CASK和GluA 1在成熟中间神经元中的相互关系。使用SIM，我们确定了一个大部分的内源性CNTNAP 2，CASK，和GluA 1分子集体共定位在一起，在一个三方的方式。最后，CNTNAP 2或CASK的单独敲除类似地改变GluA 1水平和定位。这些研究结果提供了深入了解的分子机制的GluA 1调控intereuron。
GABAergic intemeurons are emerging as prominent substrates in the pathophysiology of multiple neurodevelopmental disorders, including autism spectrum disorders, schizophrenia, intellectual disability, and epilepsy. Interneuron excitatory activity is influenced by 2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl) propanoic acid receptors (AMPARs), which in turn affects excitatory transmission in the central nervous system. Yet how dysregulation of interneuronal AMPARs distinctly contributes to the molecular underpinning of neurobiological disease is drastically underexplored. Contactin-associated protein-like 2 (CNTNAP2) is a neurexin-related adhesion molecule shown to mediate AMPAR subcellular distribution while calcium/calmodulin-dependent serine protein kinase (CASK) is a multi-functional scaffold involved with glutamate receptor trafficking. Mutations in both genes have overlapping disease associations, including autism spectrum disorders, intellectual disability, and epilepsy, thus suggesting converging perturbations of excitatory/inhibitory balance. Our lab has previously shown that CNTNAP2 stabilizes interneuron dendritic arbors through CASK and that CNTNAP2 regulates AMPAR subunit GluA1 trafficking in excitatory neurons. The interaction between these three proteins, however, has not been studied in interneurons. Using biochemical techniques, structured illumination microscopy (SIM) and shRNA technology, we first confirm that these three proteins interact in mouse brain, and then examined relationship between CNTNAP2, CASK and GluA1 in mature interneurons. Using SIM, we ascertain that a large fraction of endogenous CNTNAP2, CASK, and GluA1 molecules collectively colocalize together in a tripartite manner. Finally, individual knockdown of either CNTNAP2 or CASK similarly alter GluA1 levels and localization. These findings offer insight to molecular mechanisms underlying GluA1 regulation in intemeurons.