Subcellular localization and dynamics of MysPDZ (Myo18A) in live mammalian cells.
Subcellular localization and dynamics of MysPDZ (Myo18A) in live mammalian cells.
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DOI：
10.1016/j.bbrc.2004.11.058
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发表时间：
2005-01
影响因子：
3.1
通讯作者：
Kentaro Mori;K. Matsuda;T. Furusawa;M. Kawata;Toshiaki Inoue;M. Obinata
中科院分区：
文献类型：
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作者：
Kentaro Mori;K. Matsuda;T. Furusawa;M. Kawata;Toshiaki Inoue;M. Obinata
MysPDZ 是一种非常规肌球蛋白，属于 XVIII 类肌球蛋白，含有富含 KE（赖氨酸和谷氨酰胺）的结构域和 PDZ 结构域，其与肌动蛋白纤维部分共分布，其肌球蛋白头结构域中没有任何规范的肌动蛋白结合序列。最近，我们报道了一种新的 MysPDZ 亚型的鉴定，该亚型缺乏这些结构域，并且表现出与 MysPDZ 原始形式不同的亚细胞定位和表达谱。为了描绘指导 MysPDZ 亚细胞定位的结构域，我们使用与增强型黄色荧光蛋白融合的 MysPDZ 突变体进行了免疫共沉淀实验和图像分析。免疫共沉淀分析表明，MysPDZ 可以通过其 C 端卷曲螺旋结构域进行自缔合，而富含 KE 的结构域介导与肌动蛋白的相互作用。我们通过图像分析观察到，MysPDZ 与肌动蛋白纤维的共分布和细胞膜内表面的定位分别由富含 KE 的结构域和 PDZ 结构域控制。延时视频显微镜显示，细胞质中的MysPDZ在短范围内随机快速移动，并被分配到亚细胞区室，而没有被MysPDZ水解ATP。这表明 MysPDZ 是一种与大多数非常规肌球蛋白不同的蛋白质。我们的研究揭示了富含 KE 和 PDZ 结构域在指导亚细胞定位中的新作用，也有助于更好地理解 MysPDZ 亚型的功能差异。
MysPDZ is an unconventional myosin belonging to the class XVIII myosin containing a KE (lysine and glutamine)-rich domain and a PDZ domain, which codistributes with actin fibers partially without any canonical actin binding sequence in its myosin head domain. Recently, we reported the identification of a novel isoform of MysPDZ lacking these domains and exhibiting subcellular localization and expression profile different from the original form of MysPDZ. In order to delineate domains directing the subcellular localization of MysPDZ, we performed co-immunoprecipitation experiments and image analyses using mutants of MysPDZ fused with enhanced yellow fluorescent protein. Co-immunoprecipitation analyses showed that MysPDZ can self-associate through its C-terminus coiled-coil domain and the KE-rich domain mediates the interaction with actin. We observed by image analyses that the codistribution with actin fibers and the localization in inner surface of cell membrane of MysPDZ are controlled by the KE-rich domain and the PDZ domain, respectively. Time lapse video microscopy showed that MysPDZ in the cytoplasm moves randomly and rapidly within short range and is allocated to a subcellular compartment without ATP hydrolysis by MysPDZ. This suggests that MysPDZ is a protein which is unlike most unconventional myosins. Our study uncovers a novel role of the KE-rich and PDZ domains in directing subcellular localization and also contributes to a better understanding of functional differences in MysPDZ isoforms.