More Than Just a Cargo Adapter, Melanophilin Prolongs and Slows Processive Runs of Myosin Va
More Than Just a Cargo Adapter, Melanophilin Prolongs and Slows Processive Runs of Myosin Va
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DOI：
10.1074/jbc.m113.476929
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发表时间：
2013-10-11
影响因子：
4.8
通讯作者：
Trybus, Kathleen M.
中科院分区：
文献类型：
--
作者：
Sckolnick, Maria;Krementsova, Elena B.;Trybus, Kathleen M.
肌球蛋白Va(myosin Va，myoVa)是一种沿着肌动蛋白轨迹连续运输货物的分子马达。体内一个被研究得很好的载体是黑素小体，这是一种色素细胞器，首先由微管上的动蛋白移动，然后被移交给myoVa，在黑素细胞富含肌动蛋白的树突外围运输。亲黑素(MLPH)是连接Rab27a-黑素小体和myoVa的适配蛋白。利用全内反射荧光显微镜和量子点标记的全长myoVa，我们在单分子水平上表明，MLPH使连续运动的myoVa马达的数量增加了17倍。令人惊讶的是，myoVa-MLPH的移动速度类似于myoVa的4倍，而且运行长度是myoVa的两倍。这两个变化大大增加了在肌动蛋白上花费的时间，这一特性可能会促进黑素小体向邻近的角质形成细胞转移。与全长myoVa的可变步长模式相反，myoVa-MLPH复合体在头部之间显示出正常的门控模式，这是完全活跃的电机的典型特征，并与货物依赖的激活机制一致。MLPH依赖的myoVa的变化依赖于MLPH的肌动蛋白结合域中的一个带正电的氨基酸簇，这表明MLPH起到了将运动与轨迹联系起来的“系绳”的作用。我们的结果为肌动蛋白在体内黑素小体移动的异常缓慢速度提供了一个分子解释。更广泛地说，这些数据表明，将马达与货物联系起来的蛋白质可以改变马达的特性，以增强它们的生物学作用。
Myosin Va (myoVa) is a molecular motor that processively transports cargo along actin tracks. One well studied cargo in vivo is the melanosome, a pigment organelle that is moved first by kinesin on microtubules and then handed off to myoVa for transport in the actin-rich dendritic periphery of melanocytes. Melanophilin (Mlph) is the adapter protein that links Rab27a-melanosomes to myoVa. Using total internal reflection fluorescence microscopy and quantum dot-labeled full-length myoVa, we show at the single-molecule level that Mlph increases the number of processively moving myoVa motors by 17-fold. Surprisingly, myoVa-Mlph moves similar to 4-fold slower than myoVa alone and with twice the run length. These two changes greatly increase the time spent on actin, a property likely to enhance the transfer of melanosomes to the adjacent keratinocyte. In contrast to the variable stepping pattern of full-length myoVa, the myoVa-Mlph complex shows a normal gating pattern between the heads typical of a fully active motor and consistent with a cargo-dependent activation mechanism. The Mlph-dependent changes in myoVa depend on a positively charged cluster of amino acids in the actin binding domain of Mlph, suggesting that Mlph acts as a "tether" that links the motor to the track. Our results provide a molecular explanation for the uncharacteristically slow speed of melanosome movement by myoVa in vivo. More generally, these data show that proteins that link motors to cargo can modify motor properties to enhance their biological role.