Proteomic Studies of a Single CNS Synapse Type: The Parallel Fiber/Purkinje Cell Synapse

Precise neuronal networks underlie normal brain function and require distinct classes of synaptic connections. Although it has been shown that certain individual proteins can localize to different classes of synapses, the biochemical composition of specific synapse types is not known. Here, we have used a combination of genetically engineered mice, affinity purification, and mass spectrometry to profile proteins at parallel fiber/Purkinje cell synapses. We identify approximately 60 candidate postsynaptic proteins that can be classified into 11 functional categories. Proteins involved in phospholipid metabolism and signaling, such as the protein kinase MRCK��, are major unrecognized components of this synapse type. We demonstrate that MRCK�� can modulate maturation of dendritic spines in cultured cortical neurons, and that it is localized specifically to parallel fiber/Purkinje cell synapses in vivo. Our data identify a novel synapse-specific signaling pathway, and provide an approach for detailed investigations of the biochemical complexity of central nervous system synapse types.精准的神经元网络是正常脑功能的核心基础，其正常运作依赖于不同类别的突触连接。尽管已有研究证实，部分单个蛋白质可定位于不同类别突触，但特定突触类型的生化组成仍未明晰。本研究联合运用基因工程小鼠（genetically engineered mice）、亲和纯化（affinity purification）与质谱分析法（mass spectrometry），对平行纤维/浦肯野细胞突触（parallel fiber/Purkinje cell synapses）的蛋白质组进行了表征分析。我们共鉴定出约60个候选突触后蛋白（postsynaptic proteins），可划分为11个功能类别。其中，参与磷脂代谢与信号转导的蛋白质（如蛋白激酶MRCKβ）是该类突触此前未被发现的主要组成成分。我们证实，MRCKβ可调控体外培养皮层神经元的树突棘成熟，且在活体状态下特异性定位于平行纤维/浦肯野细胞突触。本研究不仅鉴定出一条全新的突触特异性信号通路，更为深入解析中枢神经系统不同突触类型的生化复杂性提供了可行的研究策略。