Mechanisms of dynein regulation

项目总结 微管相关的马达蛋白将来自三磷酸腺苷水解的能量耦合到机械功，这 然后用来为与细胞分裂、迁移和细胞运输相关的细胞内运动提供动力 货物。有两类微管相关的马达，细胞质动力蛋白-1(Dynein)和动蛋白。 这些马达在微管轨道上以相反的方向运动：动力蛋白朝向微管减去- 末端，通常位于细胞核附近，运动蛋白向微管+末端移动。虽然有 在超过40种动蛋白中，只有一种动力蛋白能促进细胞质货物的运输。动力蛋白含量很高 受调控并与调节其功能和活性的巨大蛋白质网络相互作用。然而，分子 有多少动力蛋白结合蛋白调节动力蛋白活性的机制在很大程度上是未知的。理解 动力蛋白如何被调控是一个迫切需要的问题，因为动力蛋白或其调节因子的突变与许多 神经发育和神经退行性疾病。这项提案试图破译管理 结合蛋白质组学，生物化学， 结构生物学、单分子荧光显微镜、固定和活细胞成像。我们将重点关注 两个最重要的目标。第一个目标是确定一类新的动力蛋白调节蛋白的功能。 是我们实验室发现的。这些新的调节因子与激动素、动力蛋白和已知的动力蛋白调节因子有关。 促进局灶性粘连的形成。通过确定这些新监管机构的职能，我们将 确定基于微管的转运在焦点粘连组装和动力学中的作用。第二个目标 是确定动力蛋白调节蛋白Nudel如何发挥作用的分子机制。在这里，我们将 重点确定Nudel如何影响动力蛋白激活的动力学以及动力蛋白结合和移动的能力 在微管轨道上。完成这一目标是当务之急，因为努德尔功能障碍与 小头症、癫痫和精神分裂症。综上所述，这项工作的结果将揭示 动力蛋白调节，这对人类健康和疾病具有广泛的影响。

PROJECT SUMMARY Microtubule-associated motor proteins couple the energy derived from ATP hydrolysis to mechanical work, which is then used to power intracellular movements associated with cell division, migration, and transport of cellular cargoes. There are two classes of microtubule-associated motors, cytoplasmic dynein-1 (dynein) and kinesins. These motors move in opposite directions on the microtubule track: dynein moves toward the microtubule minus- end, which is typically near the nucleus and kinesin moves toward the microtubule plus-end. While there are over forty kinesins, there is only one dynein that promotes cytoplasmic cargo trafficking. Dynein is highly regulated and interacts with a vast network of proteins that modulate its function and activity. However, molecular mechanisms of how many dynein binding proteins regulate dynein activity is largely unknown. Understanding how dynein is regulated is a pressing need as mutation in dynein or its regulators is associated with many neurodevelopmental and neurodegenerative diseases. This proposal seeks to decipher the rules that govern dynein-mediated cellular trafficking using an interdisciplinary approach combining proteomics, biochemistry, structural biology, single molecule fluorescence microscopy, and fixed and live cell imaging. We will focus on two overarching goals. The first goal is to determine the function of a new class of dynein regulatory proteins that our lab has discovered. These novel regulators associate with kinesin, dynein, and known dynein regulators and promote the formation of focal adhesions. By determining the function of these new regulators, we will determine the role of microtubule-based transport in focal adhesion assembly and dynamics. The second goal is to determine the molecular mechanism of how the dynein regulatory protein, Nudel functions. Here, we will focus on determining how Nudel effects the kinetics of dynein activation and dynein’s ability to bind to and move on the microtubule track. Completion of this goal is a pressing need, as Nudel dysfunction is associated with microcephaly, epilepsy, and schizophrenia. Together, the results of this work will reveal novel mechanisms of dynein regulation, which has broad implications in human health and disease.