SNAREs in native plasma membranes are active and readily form core complexes with endogenous and exogenous SNAREs
SNAREs in native plasma membranes are active and readily form core complexes with endogenous and exogenous SNAREs
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DOI：
10.1083/jcb.200203088
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发表时间：
2002-08-19
影响因子：
7.8
通讯作者：
Jahn, R
中科院分区：
文献类型：
--
作者：
Lang, T;Margittai, M;Jahn, R
在神经元胞吐期间，囊泡结合的可溶性NSF附着蛋白（SNAP）受体（SNARE）小突触泡蛋白2与质膜结合的SNARE突触融合蛋白1A和SNAP 25形成复合物以启动融合反应。然而，尚不清楚天然膜中的SNARE是否具有组成性活性，或者它们是否不能进入SNARE复合物，除非在膜融合前被激活。在此，我们使用标记的重组SNARE与由内而外载体支持的PC 12细胞质膜片的结合来探测内源性SNARE的活性。结合是特异性的，饱和的，并依赖于膜居民陷阱合作伙伴的存在。我们的数据表明，几乎所有的内源性突触融合蛋白I和SNAP-25是高度反应性的，很容易形成SNARE复合物与外源性添加的SNARE。此外，当膜新鲜制备时，内源性SNARE之间的复合物是不可检测的，但它们在体外长时间孵育后缓慢形成。我们的结论是，膜驻留的SNARE的活性不下调控制蛋白质，但组成性活跃，即使不参与融合事件。
During neuronal exocytosis, the vesicle-bound soluble NSF attachment protein (SNAP) receptor (SNARE) synaptobrevin 2 forms complexes with the plasma membrane-bound SNAREs syntaxin 1A and SNAP25 to initiate the fusion reaction. However, it is not known whether in the native membrane SNAREs are constitutively active or whether they are unable to enter SNARE complexes unless activated before membrane fusion. Here we used binding of labeled recombinant SNAREs to inside-out carrier supported plasma membrane sheets of PC12 cells to probe for the activity of endogenous SNAREs. Binding was specific, saturable, and depended on the presence of membrane-resident SNARE partners. Our data show that virtually all of the endogenous syntaxin I and SNAP-25 are highly reactive and readily form SNARE complexes with exogenously added SNAREs. Furthermore, complexes between endogenous SNAREs were not detectable when the membranes are freshly prepared, but they slowly form upon prolonged incubation in vitro. We conclude that the activity of membrane-resident SNAREs is not downregulated by control proteins but is constitutively active even if not engaged in fusion events.