The localization of myosin VI at the golgi complex and leading edge of fibroblasts and its phosphorylation and recruitment into membrane ruffles of A431 cells after growth factor stimulation.
The localization of myosin VI at the golgi complex and leading edge of fibroblasts and its phosphorylation and recruitment into membrane ruffles of A431 cells after growth factor stimulation.
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在生长因子刺激后，肌球蛋白VI在高尔基体复合物和成纤维细胞的前缘及其磷酸化和募集到A431细胞的膜褶皱中的定位。
DOI：
10.1083/jcb.143.6.1535
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发表时间：
1998-12-14
影响因子：
7.8
通讯作者：
Paul Luzio, J
中科院分区：
文献类型：
--
作者：
Buss, F;Kendrick-Jones, J;Lionne, C;Knight, A E;Cote, G P;Paul Luzio, J
肌球蛋白VI是一种非传统的肌球蛋白，可能在真核细胞中通过细胞质肌动蛋白丰富区域的囊泡膜运输中发挥作用。在本研究中，我们克隆并测序了一个编码鸡肠道刷状缘肌球蛋白VI的基因，用多克隆抗血清获得了该肌球蛋白VI的细菌表达片段。通过免疫印迹和免疫沉淀，纯化的抗体对肌球蛋白VI具有高度的特异性，并用免疫荧光和免疫电子显微镜对该蛋白进行了定位研究。研究发现，在NRK和A431细胞中，肌球蛋白VI既与高尔基复合体有关，也与细胞的前缘和褶皱边缘有关，并存在于胞浆池中。在用EGF刺激细胞表面皱折的A431细胞中，肌球蛋白VI被磷酸化，并与Ezrin和Myosin V一起被募集到新形成的褶皱中。体外实验表明，p21激活的激酶(PAK)可能是负责运动区磷酸化的激酶。这些结果有力地支持了肌球蛋白VI在分泌和内吞途径的膜运输中的作用。
Myosin VI is an unconventional myosin that may play a role in vesicular membrane traffic through actin rich regions of the cytoplasm in eukaryotic cells. In this study we have cloned and sequenced a cDNA encoding a chicken intestinal brush border myosin VI. Polyclonal antisera were raised to bacterially expressed fragments of this myosin VI. The affinity purified antibodies were highly specific for myosin VI by immunoblotting and immunoprecipitation and were used to study the localization of the protein by immunofluorescence and immunoelectron microscopy. It was found that in NRK and A431 cells, myosin VI was associated with both the Golgi complex and the leading, ruffling edge of the cell as well as being present in a cytosolic pool. In A431 cells in which cell surface ruffling was stimulated by EGF, myosin VI was phosphorylated and recruited into the newly formed ruffles along with ezrin and myosin V. In vitro experiments suggested that a p21-activated kinase (PAK) might be the kinase responsible for phosphorylation in the motor domain. These results strongly support a role for myosin VI in membrane traffic on secretory and endocytic pathways.