A combination of three distinct trafficking signals mediates axonal targeting and presynaptic clustering of GAD65
A combination of three distinct trafficking signals mediates axonal targeting and presynaptic clustering of GAD65
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DOI：
10.1083/jcb.200205053
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发表时间：
2002-09-30
影响因子：
7.8
通讯作者：
Baekkeskov, S
中科院分区：
文献类型：
--
作者：
Kanaani, J;El-Husseini, AED;Baekkeskov, S
涉及轴突运输和突触前聚集的信号定义不明确。在这里，我们证明了γ-氨基丁酸合成酶谷氨酸脱羧酶65(GAD65)的靶向突触前簇是由其棕榈酰化的60-AA NH2-末端结构域介导的，该区域可以靶向其他可溶性蛋白及其相关的伙伴到突触前末端。这一过程需要AA1-23中的高尔基体定位信号和棕榈酰化基序上游的膜锚定信号，并介导GAD65靶向高尔基体膜的胞质小叶，这是轴突分离的必要第一步。高尔基体靶向不需要膜锚定信号下游的第三个运输信号的棕榈酰化。然而，半胱氨酸30和45的棕榈酰化对于高尔基体后GAD65运输到突触前部位及其相对树突状排斥是至关重要的。细胞胆固醇水平的降低导致棕榈酰化GAD65突触前聚集的抑制，这表明该蛋白选择性地靶向突触前末端依赖于对富含胆固醇的膜微域的分选。GAD65的棕榈酰化NH2末端区域是第一个被发现的可以靶向其他蛋白质突触前簇的蛋白质区域。
he signals involved in axonal trafficking and presynaptic clustering are poorly defined. Here we show that targeting of the gamma-aminobutyric acid-synthesizing enzyme glutamate decarboxylase 65 (GAD65) to presynaptic clusters is mediated by its palmitoylated 60-aa NH2-terminal domain and that this region can target other soluble proteins and their associated partners to presynaptic termini. A Golgi localization signal in aa 1-23 followed by a membrane anchoring signal upstream of the palmitoylation motif are required for this process and mediate targeting of GAD65 to the cytosolic leaflet of Golgi membranes, an obligatory first step in axonal sorting. Palmitoylation of a third trafficking signal downstream of the membrane anchoring signal is not required for Golgi targeting. However, palmitoylation of cysteines 30 and 45 is critical for post-Golgi trafficking of GAD65 to presynaptic sites and for its relative dendritic exclusion. Reduction of cellular cholesterol levels resulted in the inhibition of presynaptic clustering of palmitoylated GAD65, suggesting that the selective targeting of the protein to presynaptic termini is dependent on sorting to cholesterol-rich membrane microdomains. The palmitoylated NH2-terminal region of GAD65 is the first identified protein region that can target other proteins to presynaptic clusters.