Membrane mediated motor kinetics in microtubule gliding assays
Membrane mediated motor kinetics in microtubule gliding assays
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DOI：
10.1038/s41598-019-45847-z
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发表时间：
2019-07-03
影响因子：
4.6
通讯作者：
Hirst, Linda S.
中科院分区：
文献类型：
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作者：
Lopes, Joseph;Quint, David A.;Hirst, Linda S.
基于运动的运输机制对真核细胞的多种功能至关重要，包括囊泡货物的长距离运输。然而，我们对与脂质底物结合时控制和调节马达的因素的理解是不完整的。我们在脂质双层底物上使用微管滑动实验来研究膜扩散在激酶-1开/关结合动力学中的作用，从而研究转运速度。荧光成像实验表明，在没有ATP的情况下，由于膜扩散，单个微管上的运动聚集，随后在滑翔过程中ATP诱导的快速解离。我们的实验数据结合分析模型表明，马达的开/关结合动力学受到扩散的影响，因此，马达的有效结合和解除结合率都远低于预期的裸速率。我们的研究结果表明，膜中的运动扩散可以通过影响运动动力学在运输中发挥重要作用，因此可以作为细胞内运输动力学的调节剂。
Motor-based transport mechanisms are critical for a wide range of eukaryotic cell functions, including the transport of vesicle cargos over long distances. Our understanding of the factors that control and regulate motors when bound to a lipid substrate is however incomplete. We used microtubule gliding assays on a lipid bilayer substrate to investigate the role of membrane diffusion in kinesin-1 on/off binding kinetics and thereby transport velocity. Fluorescence imaging experiments demonstrate motor clustering on single microtubules due to membrane diffusion in the absence of ATP, followed by rapid ATP-induced dissociation during gliding. Our experimental data combined with analytical modeling show that the on/off binding kinetics of the motors are impacted by diffusion and, as a consequence, both the effective binding and unbinding rates for motors are much lower than the expected bare rates. Our results suggest that motor diffusion in the membrane can play a significant role in transport by impacting motor kinetics and can therefore function as a regulator of intracellular transport dynamics.