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首页> 外文期刊>Molecular biology of the cell >EpsinR is an adaptor for the SNARE protein Vti1b
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EpsinR is an adaptor for the SNARE protein Vti1b

机译：EpsinR是SNARE蛋白Vti1b的适配器

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摘要

EpsinR is a clathrin-coated vesicle (CCV)-associated protein that binds to vti1b, suggesting that it may be a vti1b-selective adaptor. Depletion of epsinR to undetectable levels in HeLa cells using siRNA causes vti1b to redistribute from the perinuclear region to the cell periphery, but vti1a also redistributes in epsinR-depleted cells, and both vti isoforms redistribute in AP-1-depleted cells. As a more direct assay for epsinR function, we isolated CCVs from control and siRNA-treated cells and then looked for differences in cargo content. In clathrin-depleted cells, both coat and cargo proteins are greatly reduced in this preparation. Knocking down epsinR causes a similar to50% reduction in the amount of AP-1 copurifying with CCVs and vice versa, indicating that the two proteins are dependent on each other for maximum incorporation into the coat. In addition, vti1b, but not vti1a, is reduced by >70% in CCVs from both epsinR- and AP-1-depleted cells. Because AP-1 knockdown reduces the amount of epsinR in CCVs, it is possible that its effect on vti1b may be indirect. These findings provide in vivo evidence that epsinR is an adaptor for vti1b, and they also show that CCV isolation can be used as an assay for adaptor function.
机译：EpsinR是与网格蛋白包被的囊泡（CCV）相关的蛋白，可与vti1b结合，表明它可能是vti1b的选择性衔接子。使用siRNA在HeLa细胞中将epsinR耗竭至无法检测的水平会导致vti1b从核周区域重新分布到细胞周围，但是vti1a也在epsinR耗竭的细胞中重新分布，并且两种vti异构体都在AP-1耗竭的细胞中重新分布。为了更直接地分析epsinR功能，我们从对照和siRNA处理的细胞中分离了CCV，然后寻找货物含量的差异。在缺乏网格蛋白的细胞中，此制备过程中外壳蛋白和货物蛋白均大大降低。敲低epsinR会导致与CCV共纯化的AP-1数量减少近50％，反之亦然，这表明这两种蛋白相互依赖，从而最大程度地掺入了皮层。此外，在耗尽epsinR和AP-1的细胞中，CCV中的vti1b而不是vti1a减少了70％以上。由于AP-1敲低可减少CCV中的epsinR数量，因此它对vti1b的影响可能是间接的。这些发现提供了体内的证据，证明epsinR是vti1b的衔接子，并且它们还表明CCV分离可用作衔接子功能的检测方法。

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