Different effects on fast exocytosis induced by synaptotagmin 1 and 2 isoforms and abundance but not by phosphorylation
Different effects on fast exocytosis induced by synaptotagmin 1 and 2 isoforms and abundance but not by phosphorylation
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DOI：
10.1523/jneurosci.2589-05.2006
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发表时间：
2006-01-11
影响因子：
5.3
通讯作者：
Sorensen, JB
中科院分区：
文献类型：
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作者：
Nagy, G;Kim, JH;Sorensen, JB
突触结合蛋白是一个大的蛋白质家族，其中突触结合蛋白1（Syt 1）是一个快速胞吐的Ca 2+传感器，而它的近亲，突触结合蛋白2（Syt 2），被认为具有类似的功能。嗜铬细胞表达Syt 1，但不表达Syt 2。我们比较了过表达任一Syt亚型的Syt 1缺失小鼠嗜铬细胞的分泌。高时间分辨率的电容测量表明，Syt 1空细胞缺乏对应于囊泡的易释放池（RRP）的胞吐相。与安培信号的比较证实，缺失的胞吐相由含儿茶酚胺的囊泡组成。Syt 1的过表达拯救了RRP，并增加了其大小高于野生型值，而缓慢释放池的大小下降，表明Syt 1的可用性调节两个可释放池的相对大小。RRP也被Syt 2过表达拯救，但融合动力学比表达Syt 1的细胞稍慢。生物化学实验表明，Syt 2具有比Syt 1稍低的Ca 2+对磷脂结合的亲和力，因为C2 A结构域的差异。这些数据构成了Syt 1和Syt 2作为RRP融合的替代但不相同的钙传感器的功能的证据。通过过度表达突变的Syt 1在共享的PKC/钙/钙调蛋白依赖性激酶磷酸化位点，我们表明佛波酯的作用独立和上游的Syt 1调节可释放池的大小。我们的结论是，从小鼠嗜铬细胞的胞吐可以修改的差异表达的Syt亚型和Syt丰度，但不是由磷酸化Syt 1。
Synaptotagmins comprise a large protein family, of which synaptotagmin 1 (Syt1) is a Ca2+ sensor for fast exocytosis, and its close relative, synaptotagmin 2 (Syt2), is assumed to serve similar functions. Chromaffin cells express Syt1 but not Syt2. We compared secretion from chromaffin cells from Syt1 null mice overexpressing either Syt isoform. High time-resolution capacitance measurement showed that Syt1 null cells lack the exocytotic phase corresponding to the readily-releasable pool (RRP) of vesicles. Comparison with the amperometric signal confirmed that the missing phase of exocytosis consists of catecholamine-containing vesicles. Overexpression of Syt1 rescued the RRP and increased its size above wild-type values, whereas the size of the slowly releasable pool decreased, indicating that the availability of Syt1 regulates the relative size of the two releasable pools. The RRP was also rescued by Syt2 overexpression, but the kinetics of fusion was slightly slower than in cells expressing Syt1. Biochemical experiments showed that Syt2 has a slightly lower Ca2+ affinity for phospholipid binding than Syt1 because of a difference in the C2A domain. These data constitute evidence for the function of Syt1 and Syt2 as alternative, but not identical, calcium-sensors for RRP fusion. By overexpression of Syt1 mutated in the shared PKC/calcium/calmodulin-dependent kinase phosphorylation site, we show that phorbol esters act independently and upstream of Syt1 to regulate the size of the releasable pools. We conclude that exocytosis from mouse chromaffin cells can be modified by the differential expression of Syt isoforms and by Syt abundance but not by phosphorylation of Syt1.