Myosin-X is a molecular motor that functions in filopodia formation
Myosin-X is a molecular motor that functions in filopodia formation
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DOI：
10.1073/pnas.0602443103
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发表时间：
2006-08-15
影响因子：
11.1
通讯作者：
Cheney, Richard E.
中科院分区：
文献类型：
--
作者：
Bohil, Aparna B.;Robertson, Brian W.;Cheney, Richard E.
尽管最近在了解片状伪足延伸方面取得了进展，但调节丝状伪足形成的分子机制在很大程度上仍然未知。Myo 10是一种MyTH 4-FERM肌球蛋白，定位于丝状伪足的尖端，并被假设在丝状伪足形成中起作用。为了确定丝状伪足的形成是否需要内源性MyolO，我们使用扫描EM来测定通常存在于HeLa细胞背表面的大量丝状伪足。我们在这里表明，siRNA介导的敲低HeLa细胞中的Myo 1 O导致背丝状伪足的戏剧性损失。MyolO的卷曲螺旋区作为显性负调控因子过度表达也会导致背侧丝状伪足的缺失，从而为Myo 10在丝状伪足形成中发挥作用提供了独立的证据。我们还表明，在COS-7细胞，一个细胞系，通常缺乏背丝状伪足表达Myo 10，导致大量诱导背丝状伪足。由于Myo 10诱导的背侧丝状伪足不附着于基底，Myo 10可以通过独立于基底附着的机制促进丝状伪足。与这一观察结果一致，缺乏FERM结构域（与整联蛋白结合的区域）的Myo 10构建体保留了诱导背丝状伪足的能力。然而，MyTH 4-FERM区域的缺失完全废除了Myo 10的丝状伪足促进活性，就像运动结构域的缺失一样。关于Myo 10作用机制的其他实验表明，它作用于Cdc 42的下游，并且可以在不存在VASP蛋白的情况下促进丝状伪足。总之，这些数据表明，Myo 10是一个分子马达，在丝状伪足形成的功能。
Despite recent progress in understanding lamellipodia extension, the molecular mechanisms regulating filopodia formation remain largely unknown. Myo10 is a MyTH4-FERM myosin that localizes to the tips of filopodia and is hypothesized to function in filopodia formation. To determine whether endogenous MyolO is required for filopodia formation, we have used scanning EM to assay the numerous filopodia normally present on the dorsal surfaces of HeLa cells. We show here that siRNA-mediated knockdown of Myo1O in HeLa cells leads to a dramatic loss of dorsal filopodia. Overexpressing the coiled coil region from MyolO as a dominantnegative also leads to a loss of dorsal filopodia, thus providing independent evidence that Myo10 functions in filopodia formation. We also show that expressing Myo10 in COS-7 cells, a cell line that normally lacks dorsal filopodia, leads to a massive induction of dorsal filopodia. Because the dorsal filopodia induced by Myo10 are not attached to the substrate, Myo10 can promote filopodia by a mechanism that is independent of substrate attachment. Consistent with this observation, a Myo10 construct that lacks the FERM domain, the region that binds to integrin, retains the ability to induce dorsal filopodia. Deletion of the MyTH4-FERM region, however, completely abolishes Myo10's filopodia-promoting activity, as does deletion of the motor domain. Additional experiments on the mechanism of Myo10 action indicate that it acts downstream of Cdc42 and can promote filopodia in the absence of VASP proteins. Together, these data demonstrate that Myo10 is a molecular motor that functions in filopodia formation.