Recruitment of Epac2A to Insulin Granule Docking Sites Regulates Priming for Exocytosis
Recruitment of Epac2A to Insulin Granule Docking Sites Regulates Priming for Exocytosis
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DOI：
10.2337/db17-0050
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发表时间：
2017-10-01
期刊：
影响因子：
7.7
通讯作者：
Tengholm, Anders
中科院分区：
文献类型：
--
作者：
Alenkvist, Ida;Gandasi, Nikhil R.;Tengholm, Anders
Epac是一种cAMP激活的鸟嘌呤核苷酸交换因子，在包括-细胞在内的各种类型的细胞中介导cAMP信号传导，参与胰岛素分泌的控制。激活后，蛋白质重新分配到质膜，但潜在的分子机制和功能后果尚不清楚。使用定量高分辨率显微镜，我们发现cAMP升高导致Epac2A与细胞质膜快速结合，在那里它特异性地积聚在分泌颗粒上，使它们更容易发生胞吐。camp依赖的膜结合需要高亲和力的环核苷酸结合（CNB）和Ras结合结构域，而不需要disheveled-Egl-10-pleckstrin结构域。尽管n端低亲和CNB结构域（CNB- a）对于细胞膜的易位是必不可少的，但它对于将Epac2A引导到颗粒位点至关重要。缺乏CNB-A结构域的Epac1被招募到质膜上，但没有在颗粒中积累。我们得出结论，Epac2A通过结合胞吐机制来控制分泌颗粒的释放，这种作用被先前的camp依赖性蛋白在质膜上的积累所增强。
Epac is a cAMP-activated guanine nucleotide exchange factor that mediates cAMP signaling in various types of cells, including -cells, where it is involved in the control of insulin secretion. Upon activation, the protein redistributes to the plasma membrane, but the underlying molecular mechanisms and functional consequences are unclear. Using quantitative high-resolution microscopy, we found that cAMP elevation caused rapid binding of Epac2A to the -cell plasma membrane, where it accumulated specifically at secretory granules and rendered them more prone to undergo exocytosis. cAMP-dependent membrane binding required the high-affinity cyclic nucleotide-binding (CNB) and Ras association domains, but not the disheveled-Egl-10-pleckstrin domain. Although the N-terminal low-affinity CNB domain (CNB-A) was dispensable for the translocation to the membrane, it was critical for directing Epac2A to the granule sites. Epac1, which lacks the CNB-A domain, was recruited to the plasma membrane but did not accumulate at granules. We conclude that Epac2A controls secretory granule release by binding to the exocytosis machinery, an effect that is enhanced by prior cAMP-dependent accumulation of the protein at the plasma membrane.