Yao Lin-Lin, Shen Mei, Lu Zekuan, Ikebe Mitsuo, Li Xiang-dong
From the Group of Cell Motility and Muscle Contraction, State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China and.
the Department of Cellular and Molecular Biology, University of Texas Health Science Center at Tyler, Tyler, Texas 75708.
J Biol Chem. 2016 Apr 8;291(15):8241-50. doi: 10.1074/jbc.M115.693762. Epub 2016 Feb 24.
Vertebrates have three isoforms of class V myosin (Myo5), Myo5a, Myo5b, and Myo5c, which are involved in transport of multiple cargoes. It is well established that the motor functions of Myo5a and Myo5b are regulated by a tail inhibition mechanism. Here we found that the motor function of Myo5c was also inhibited by its globular tail domain (GTD), and this inhibition was abolished by high Ca(2+), indicating that the tail inhibition mechanism is conserved in vertebrate Myo5. Interestingly, we found that Myo5a-GTD and Myo5c-GTD were not interchangeable in terms of inhibition of motor function, indicating isoform-specific interactions between the GTD and the head of Myo5. To identify the isoform-specific interactions, we produced a number of Myo5 chimeras by swapping the corresponding regions of Myo5a and Myo5c. We found that Myo5a-GTD, with its H11-H12 loop being substituted with that of Myo5c, was able to inhibit the ATPase activity of Myo5c and that Myo5a-GTD was able to inhibit the ATPase activity of Myo5c-S1 and Myo5c-HMM only when their IQ1 motif was substituted with that of Myo5a. Those results indicate that the H11-H12 loop in the GTD and the IQ1 motif in the head dictate the isoform-specific interactions between the GTD and head of Myo5. Because the IQ1 motif is wrapped by calmodulin, whose conformation is influenced by the sequence of the IQ1 motif, we proposed that the calmodulin bound to the IQ1 motif interacts with the H11-H12 loop of the GTD in the inhibited state of Myo5.
脊椎动物有三种V类肌球蛋白（Myo5）亚型，即Myo5a、Myo5b和Myo5c，它们参与多种货物的运输。众所周知，Myo5a和Myo5b的运动功能受尾部抑制机制调控。在此，我们发现Myo5c的运动功能也受到其球状尾部结构域（GTD）的抑制，而高钙（Ca²⁺）可消除这种抑制作用，这表明尾部抑制机制在脊椎动物Myo5中是保守的。有趣的是，我们发现就运动功能抑制而言，Myo5a - GTD和Myo5c - GTD不可互换，这表明GTD与Myo5头部之间存在亚型特异性相互作用。为了确定亚型特异性相互作用，我们通过交换Myo5a和Myo5c的相应区域构建了许多Myo5嵌合体。我们发现，Myo5a - GTD的H11 - H12环被Myo5c的替换后，能够抑制Myo5c的ATP酶活性，并且只有当Myo5c - S1和Myo5c - HMM的IQ1基序被Myo5a的替换时，Myo5a - GTD才能抑制它们的ATP酶活性。这些结果表明，GTD中的H11 - H12环和头部中的IQ1基序决定了Myo5的GTD与头部之间的亚型特异性相互作用。由于IQ1基序被钙调蛋白包裹，其构象受IQ1基序序列影响，我们推测在Myo5的抑制状态下，与IQ1基序结合的钙调蛋白与GTD的H11 - H12环相互作用。
