Myosin-Va regulates exocytosis through the submicromolar Ca2+-dependent binding of syntaxin-1A.
Myosin-Va regulates exocytosis through the submicromolar Ca2+-dependent binding of syntaxin-1A.
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DOI：
10.1091/mbc.e05-03-0252
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发表时间：
2005-10
影响因子：
3.3
通讯作者：
Michitoshi Watanabe;Kazushige Nomura;A. Ohyama;R. Ishikawa;Y. Komiya;K. Hosaka;E. Yamauchi;H. Taniguchi;N. Sasakawa;K. Kumakura;T. Ushiki;O. Sato;M. Ikebe;M. Igarashi
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文献类型：
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作者：
Michitoshi Watanabe;Kazushige Nomura;A. Ohyama;R. Ishikawa;Y. Komiya;K. Hosaka;E. Yamauchi;H. Taniguchi;N. Sasakawa;K. Kumakura;T. Ushiki;O. Sato;M. Ikebe;M. Igarashi
肌球蛋白-Va是一种以肌动蛋白为基础的过程性马达，负责运输细胞内的货物。突触小泡是肌球蛋白-Va最重要的运输工具之一，但哺乳动物的肌球蛋白-Va在分泌中的作用不如它的酵母同源物Myo2p那么清楚。在目前的研究中，我们发现突触小泡上的肌球蛋白-Va在0.3微米或更高的钙离子浓度下，与参与胞吐作用的t-陷阱Synaxin-1A相互作用。干扰合成素-1A-肌球蛋白-Va复合体的形成会降低嗜铬细胞的胞吐频率。令人惊讶的是，Synaxin-1A结合位点不在myosin-Va的尾部，而是在颈部，这是一个包含钙调蛋白结合IQ基序的区域。此外，我们还发现，在有钙离子存在的情况下，肌球蛋白-Va与Synaxin-1A的结合依赖于肌球蛋白-Va颈部钙调蛋白的释放，从而允许Synaxin-1A占据空缺的IQ基序。使用抗肌球蛋白-Va颈部抗体，阻断这种结合，我们证明了抗体抑制活性最重要的步骤是后期持续阶段，这涉及到提供容易释放的囊泡。我们的结果表明，肌球蛋白-Va和合成素-1A之间的相互作用参与了胞吐作用，并提示肌球蛋白-Va颈不仅参与了大步长，而且还参与了钙离子对胞吐作用的调节。
Myosin-Va is an actin-based processive motor that conveys intracellular cargoes. Synaptic vesicles are one of the most important cargoes for myosin-Va, but the role of mammalian myosin-Va in secretion is less clear than for its yeast homologue, Myo2p. In the current studies, we show that myosin-Va on synaptic vesicles interacts with syntaxin-1A, a t-SNARE involved in exocytosis, at or above 0.3 microM Ca2+. Interference with formation of the syntaxin-1A-myosin-Va complex reduces the exocytotic frequency in chromaffin cells. Surprisingly, the syntaxin-1A-binding site was not in the tail of myosin-Va but rather in the neck, a region that contains calmodulin-binding IQ-motifs. Furthermore, we found that syntaxin-1A binding by myosin-Va in the presence of Ca2+ depends on the release of calmodulin from the myosin-Va neck, allowing syntaxin-1A to occupy the vacant IQ-motif. Using an anti-myosin-Va neck antibody, which blocks this binding, we demonstrated that the step most important for the antibody's inhibitory activity is the late sustained phase, which is involved in supplying readily releasable vesicles. Our results demonstrate that the interaction between myosin-Va and syntaxin-1A is involved in exocytosis and suggest that the myosin-Va neck contributes not only to the large step size but also to the regulation of exocytosis by Ca2+.