Neuroscience Center and Curriculum in Neurobiology, 2 Department of Cell Biology and Physiology, and 3 Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599.
J Cell Biol. 2014 Apr 28;205(2):217-32. doi: 10.1083/jcb.201311003.
Developmental axon branching dramatically increases synaptic capacity and neuronal surface area. Netrin-1 promotes branching and synaptogenesis, but the mechanism by which Netrin-1 stimulates plasma membrane expansion is unknown. We demonstrate that SNARE-mediated exocytosis is a prerequisite for axon branching and identify the E3 ubiquitin ligase TRIM9 as a critical catalytic link between Netrin-1 and exocytic SNARE machinery in murine cortical neurons. TRIM9 ligase activity promotes SNARE-mediated vesicle fusion and axon branching in a Netrin-dependent manner. We identified a direct interaction between TRIM9 and the Netrin-1 receptor DCC as well as a Netrin-1-sensitive interaction between TRIM9 and the SNARE component SNAP25. The interaction with SNAP25 negatively regulates SNARE-mediated exocytosis and axon branching in the absence of Netrin-1. Deletion of TRIM9 elevated exocytosis in vitro and increased axon branching in vitro and in vivo. Our data provide a novel model for the spatial regulation of axon branching by Netrin-1, in which localized plasma membrane expansion occurs via TRIM9-dependent regulation of SNARE-mediated vesicle fusion.
发育中的轴突分支显著增加了突触容量和神经元表面积。Netrin-1 促进分支和突触发生，但 Netrin-1 刺激质膜扩展的机制尚不清楚。我们证明 SNARE 介导的胞吐作用是轴突分支的前提条件，并确定 E3 泛素连接酶 TRIM9 是 Netrin-1 和小鼠皮质神经元中胞吐 SNARE 机制之间的关键催化连接。TRIM9 连接酶活性以 Netrin-1 依赖的方式促进 SNARE 介导的囊泡融合和轴突分支。我们发现 TRIM9 与 Netrin-1 受体 DCC 之间存在直接相互作用，以及 TRIM9 与 SNARE 成分 SNAP25 之间存在 Netrin-1 敏感的相互作用。与 SNAP25 的相互作用负调节 Netrin-1 缺失时的 SNARE 介导的胞吐作用和轴突分支。TRIM9 的缺失增加了体外的胞吐作用，并增加了体外和体内的轴突分支。我们的数据提供了 Netrin-1 调控轴突分支的空间调节的新模型，其中局部质膜扩展是通过 TRIM9 依赖的 SNARE 介导的囊泡融合调节发生的。
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