Amphiphysin I is associated with coated endocytic intermediates and undergoes stimulation-dependent dephosphorylation in nerve terminals
Amphiphysin I is associated with coated endocytic intermediates and undergoes stimulation-dependent dephosphorylation in nerve terminals
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DOI：
10.1074/jbc.272.49.30984
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发表时间：
1997-12-05
影响因子：
4.8
通讯作者：
De Camilli, P
中科院分区：
文献类型：
--
作者：
Bauerfeind, R;Takei, K;De Camilli, P
双囊腺垂体素I是一种丰富的突触前蛋白，其通过其COOH末端src同源3（SH 3）结构域与GT3发动蛋白I和肌醇-5-磷酸酶突触Janin相互作用。动力蛋白I和synaptojanin I都在突触囊泡循环中起作用，并在去极化刺激刺激下分泌的大鼠脑突触体中经历快速的去磷酸化。我们在这里发现，两栖physin I也经历组成性磷酸化和刺激依赖性去磷酸化。两栖physin I的去磷酸化需要细胞外Ca ~（2+），并且不受破伤风毒素预处理突触体的影响。而不是突触囊泡胞吐作用。与发动蛋白I和突触Janin I的去磷酸化一样，两性生理蛋白I的去磷酸化被环孢菌素A和FK-506（0.5 μ M）抑制，这两种药物特异性阻断Ca 2 +/钙调蛋白依赖性磷酸酶2B钙调神经磷酸酶，但不被冈田酸（1 μ M）抑制，冈田酸阻断蛋白磷酸酶1和2B。我们还通过免疫胶体金电子显微镜免疫细胞化学显示，两栖蛋白I定位于神经终末细胞基质中，并部分与内吞中间体相关，这些中间体包括网格蛋白包被的芽和动力蛋白包被的小管，它们在鸟苷5 '-3-O-（硫代）三磷酸存在下孵育的神经终末膜中积累。这些数据支持这一假设，即两栖physin I，发动蛋白I和syaptojanin I是突触囊泡内吞作用的某些步骤中的生理伙伴。我们假设，两栖physin I及其两个主要结合蛋白的平行的Ca 2+依赖性钙调神经磷酸酶依赖性去磷酸化是一个过程的一部分，该过程引发神经末梢响应于胞吐的爆发而进行内吞作用。
Amphiphysin I is an abundant presynaptic protein that interacts via its COOH-terminal src homology 3 (SH3) domain with the GTPase dynamin I and the inositol-5-phosphatase synaptojanin., Both dynamin I and synaptojanin I have a putative role in synaptic vesicle recycling and undergo rapid dephosphorylation in rat brain synaptosomes stimulated to secrete by a depolarizing stimulus, We show here that amphiphysin I also undergoes constitutive phosphorylation and stimulationdependent dephosphorylation, Dephosphorylation of amphiphysin I requires extracellular Ca2+ and is unaffected by pretreatment of synaptosomes with tetanus toxin, Thus, Ca2+ influx, but not synaptic vesicle exocytosis, is required for dephosphorylation. Dephosphorylation of amphiphysin I, like dephosphorylation of dynamin I and synaptojanin I, is inhibited by cyclosporin A and FK-506 (0.5 mu M), two drugs that specifically block the Ca2+/calmodulin-dependent phosphatase 2B calcineurin, but not by okadaic acid (1 mu M), which blocks protein phosphatases 1 and 2B. We also show by immunogold electron microscopy immunocytochemistry that amphiphysin I is localized in the nerve terminal cytomatrix and is partially associated with endocytic intermediates, These include the clathrin-coated buds and dynamin-coated tubules, which accumulate in nerve terminal membranes incubated in the presence of guanosine 5'-3-O-(thio)triphosphate. These data support the hypothesis that amphiphysin I, dynamin I, and syaptojanin I are physiological partners in some step(s) of synaptic vesicle endocytosis, We hypothesize that the parallel Ca2+-dependent calcineurin-dependent dephosphorylation of amphiphysin I and of its two major binding proteins is part of a process that primes the nerve terminal for endocytosis in response to a burst of exocytosis.