The actin cytoskeleton and active membrane trafficking machinery are essential for polarized cell growth. To understand the interactions between myosin XI, vesicles and actin filaments in vivo, we performed fluorescence recovery after photobleaching and showed that the dynamics of myosin XIa at the tip of the spreading earthmoss Physcomitrella patens caulonemal cells are actin-dependent and that 50% of myosin XI is bound to vesicles. To obtain single-particle information, we used variable-angle epifluorescence microscopy in protoplasts to demonstrate that protein myosin XIa and VAMP72-labeled vesicles localize in time and space over periods lasting only a few seconds. By tracking data with Hidden Markov modeling, we showed that myosin XIa and VAMP72-labeled vesicles exhibit short runs of actin-dependent directed transport. We also found that the interaction of myosin XI with vesicles is short-lived. Together, this vesicle-bound fraction, fast off-rate and short average distance traveled seem be crucial for the dynamic oscillations observed at the tip, and might be vital for regulation and recycling of the exocytosis machinery, while simultaneously promoting vesicle focusing and vesicle secretion at the tip, necessary for cell wall expansion.

中文翻译：

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肌球蛋白十一，囊泡和丝状肌动蛋白之间的体内相互作用是快速和短暂的。

肌动蛋白细胞骨架和活性膜运输机制对于极化细胞的生长至关重要。为了了解肌球蛋白XI，囊泡和肌动蛋白丝在体内的相互作用，我们进行了光漂白后的荧光恢复，并显示了在蔓延的大地藓Physcomitrella patens caulonemal细胞尖端的肌球蛋白XIa的动力学是肌动蛋白依赖性的，并且有50％的肌球蛋白十一绑定到囊泡。为了获得单颗粒信息，我们在原生质体中使用了可变角度的落射荧光显微镜来证明蛋白肌球蛋白XIa和VAMP72标记的囊泡仅持续几秒钟就可以在时间和空间上定位。通过使用隐式马尔可夫模型跟踪数据，我们表明肌球蛋白XIa和VAMP72标记的囊泡表现出短期的肌动蛋白依赖性定向转运。我们还发现，肌球蛋白XI与囊泡的相互作用是短暂的。总之，这种囊泡结合部分，快速的失速速率和短的平均距离行进似乎对于在尖端观察到的动态振荡至关重要，并且对于调节和回收胞吐作用机制至关重要，同时促进囊泡聚焦和囊泡分泌在尖端，是细胞壁扩张所必需的。