Myosin Va mediates docking of secretory granules at the plasma membrane
Myosin Va mediates docking of secretory granules at the plasma membrane
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DOI：
10.1523/jneurosci.1228-07.2007
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发表时间：
2007-09-26
影响因子：
5.3
通讯作者：
Darchen, Francois
中科院分区：
文献类型：
--
作者：
Desnos, Claire;Huet, Sebastien;Darchen, Francois
肌球蛋白 Va (MyoVa) 是控制神经元和神经内分泌细胞中基于肌动蛋白的细胞器运动的主要候选者。通过宽视野和全内反射荧光显微镜在肠嗜铬细胞中研究其在分泌颗粒（SG）运输中的功能。内源性MyoVa的分布与SG和微管部分重叠。通过截短的构建体（MyoVa 尾部）或 RNA 干扰损害 MyoVa 功能，可以防止细胞外周富含 SG 区域的形成，并降低质膜下区域的 SG 密度。跟踪各个 SG 轨迹以分析 SG 移动性。观察到它们的扩散系数 D-xy 分布广泛。基于以下两个特性，几乎不动的 SG（Dxy < 5 x 10-4 μ m(2) 中心点 s(-1)）被认为停靠在质膜上：（1）经历胞吐作用的 SG 在融合前至少 2 s 的 Dxy 低于此阈值； (2) 在 Dxy 低于阈值的子轨迹中发现垂直运动的负自相关。使用这种对接标准，我们发现 MyoVa 抑制的主要作用是减少对接颗粒的数量，从而导致分泌反应减少。令人惊讶的是，这种减少并不是由于 SG 向释放点的运输减少所致。相比之下，MyoVa 沉默减少了长期对接期的发生，但不减少短期对接期的发生。因此，我们提出，尽管 MyoVa 具有已知的运动活性，但它直接介导 SG 在质膜上的稳定附着。
Myosin Va (MyoVa) is a prime candidate for controlling actin-based organelle motion in neurons and neuroendocrine cells. Its function in secretory granule (SG) trafficking was investigated in enterochromaffin cells by wide-field and total internal reflection fluorescence microscopy. The distribution of endogenous MyoVa partially overlapped with SGs and microtubules. Impairing MyoVa function by means of a truncated construct (MyoVa tail) or RNA interference prevented the formation of SG-rich regions at the cell periphery and reduced SG density in the subplasmalemmal region. Individual SG trajectories were tracked to analyze SG mobility. A wide distribution of their diffusion coefficient, D-xy, was observed. Almost immobile SGs (Dxy < 5 x 10-4 mu m(2) center dot s(-1)) were considered as docked at the plasma membrane based on two properties: (1) SGs that undergo exocytosis have a Dxy below this threshold value for at least 2 s before fusion; (2) a negative autocorrelation of the vertical motion was found in subtrajectories with a Dxy below the threshold. Using this criterion of docking, we found that the main effect of MyoVa inhibition was to reduce the number of docked granules, leading to reduced secretory responses. Surprisingly, this reduction was not attributable to a decreased transport of SGs toward release sites. In contrast, MyoVa silencing reduced the occurrence of long-lasting, but not short-lasting, docking periods. We thus propose that, despite its known motor activity, MyoVa directly mediates stable attachment of SGs at the plasma membrane.