Regulation of releasable vesicle pool sizes by protein kinase A-dependent phosphorylation of SNAP-25
Regulation of releasable vesicle pool sizes by protein kinase A-dependent phosphorylation of SNAP-25
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DOI：
10.1016/s0896-6273(04)00038-8
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发表时间：
2004-02-05
期刊：
影响因子：
16.2
通讯作者：
Sorensen, JB
中科院分区：
文献类型：
--
作者：
Nagy, G;Reim, K;Sorensen, JB
蛋白激酶 A (PKA) 是神经分泌的关键调节因子，但分子靶标仍然难以捉摸。我们将激酶和磷酸酶活性的药理学操作与驱动嗜铬细胞中含有儿茶酚胺的囊泡融合的胞吐机制的突变研究结合起来。我们发现，组成型 PKA 活性对于维持大量囊泡处于释放就绪状态（所谓的启动状态）是必要的，而钙调神经磷酸酶（蛋白磷酸酶 2B）活性则拮抗这种作用。 PKA 磷酸化位点 (Thr-138) 突变的 SNARE 蛋白 SNAP-25a 的过度表达消除了 PKA 抑制剂对囊泡启动过程的影响。另一个未识别的 PKA 靶标调节两个不同的引发囊泡池的相对大小，这两个囊泡池的区别在于其释放动力学。 SNAP-25b 同工型的过度表达使两个引发的囊泡库的大小增加了两倍，并且保守的 Thr-138 位点的突变与 a 同工型具有相似的效果。
Protein kinase A (PKA) is a key regulator of neurosecretion, but the molecular targets remain elusive. We combined pharmacological manipulations of kinase and phosphatase activities with mutational studies on the exocytotic machinery driving fusion of catecholamine-containing vesicles from chromaffin cells. We found that constitutive PKA activity was necessary to maintain a large number of vesicles in the release-ready, so-called primed, state, whereas calcineurin (protein phosphatase 2B) activity antagonized this effect. Overexpression of the SNARE protein SNAP-25a mutated in a PKA phosphorylation site (Thr-138) eliminated the effect of PKA inhibitors on the vesicle priming process. Another, unidentified, PKA target regulated the relative size of two different primed vesicle pools that are distinguished by their release kinetics. Overexpression of the SNAP-25b isoform increased the size of both primed vesicle pools by a factor of two, and mutations in the conserved Thr-138 site had similar effects as in the a isoform.