Dynamic multimerization of Dab2-Myosin VI complexes regulates cargo processivity while minimizing cortical actin reorganization.
Dynamic multimerization of Dab2-Myosin VI complexes regulates cargo processivity while minimizing cortical actin reorganization.
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DOI：
10.1074/jbc.ra120.012703
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发表时间：
2021-01
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影响因子：
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通讯作者：
Sivaramakrishnan S
中科院分区：
文献类型：
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作者：
Rai A;Vang D;Ritt M;Sivaramakrishnan S
内吞货物上的肌球蛋白VI有助于通过致密的皮质肌动蛋白网络进行定向运输。肌球蛋白VI通过货物衔接子Dab 2募集到网格蛋白包被的内体。规范上，它已被假定之间的相互作用马达和其货物适配器是稳定的。然而，已经证明，由多个稳定连接的马达产生的力破坏局部细胞骨架结构，可能损害运输。在这项研究中，我们表明，动态多聚化的肌球蛋白VI-Dab 2复合物有利于货物的持续合成能力没有显着重组的皮质肌动蛋白网络。具体来说，我们发现Dab 2肌球蛋白相互作用区（MIR）以中等亲和力（184 nM）结合肌球蛋白VI，单分子动力学测量表明Dab 2 MIR-肌球蛋白VI相互作用的高周转率（1 s−1）。单分子运动性表明，饱和Dab 2-MIR浓度（2 μM）促进肌球蛋白VI同源二聚化和持续合成能力，其运行长度与组成型肌球蛋白VI二聚体相当。用Dab 2 MIR-肌球蛋白VI复合物图案化的货物模拟DNA折纸支架是弱进行性的，在单个肌动蛋白丝上显示稀疏的运动性，在细胞肌动蛋白网络上显示“走走停停”的运动。在最小的肌动蛋白皮质上组装的脂质双层，不受管制的进行性运动，无论是组成型肌球蛋白V或VI二聚体的结果在肌动蛋白重塑和病灶的形成。与此相反，Dab 2 MIR-myosin VI相互作用保持了最小皮质肌动蛋白网络的完整性。总之，我们的研究表明，在不破坏细胞骨架组织的情况下，动态马达-货物协会在使货物运输方面的重要性。
Myosin VI ensembles on endocytic cargo facilitate directed transport through a dense cortical actin network. Myosin VI is recruited to clathrin-coated endosomes via the cargo adaptor Dab2. Canonically, it has been assumed that the interactions between a motor and its cargo adaptor are stable. However, it has been demonstrated that the force generated by multiple stably attached motors disrupts local cytoskeletal architecture, potentially compromising transport. In this study, we demonstrate that dynamic multimerization of myosin VI-Dab2 complexes facilitates cargo processivity without significant reorganization of cortical actin networks. Specifically, we find that Dab2 myosin interacting region (MIR) binds myosin VI with a moderate affinity (184 nM) and single-molecule kinetic measurements demonstrate a high rate of turnover (1 s−1) of the Dab2 MIR–myosin VI interaction. Single-molecule motility shows that saturating Dab2-MIR concentration (2 μM) promotes myosin VI homodimerization and processivity with run lengths comparable with constitutive myosin VI dimers. Cargo-mimetic DNA origami scaffolds patterned with Dab2 MIR-myosin VI complexes are weakly processive, displaying sparse motility on single actin filaments and “stop-and-go” motion on a cellular actin network. On a minimal actin cortex assembled on lipid bilayers, unregulated processive movement by either constitutive myosin V or VI dimers results in actin remodeling and foci formation. In contrast, Dab2 MIR–myosin VI interactions preserve the integrity of a minimal cortical actin network. Taken together, our study demonstrates the importance of dynamic motor–cargo association in enabling cargo transportation without disrupting cytoskeletal organization.