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首页> 外文期刊>Protoplasma: An International Journal of Cell Biology >Rho/Rho-dependent kinase affects locomotion and actin-myosin II activity of Amoeba proteus
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Rho/Rho-dependent kinase affects locomotion and actin-myosin II activity of Amoeba proteus

机译：Rho / Rho依赖激酶影响变形虫变形虫的运动和肌动蛋白-肌球蛋白II活性

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摘要

The highly motile free-living unicellular organism Amoeba proteus has been widely used as a model to study cell motility. However, the molecular mechanisms underlying its unique locomotion are still scarcely known. Recently, we have shown that blocking the amoebae's endogenous Rac- and Rho-like proteins led to distinct and irreversible changes in the appearance of these large migrating cells as well as to a significant inhibition of their locomotion. In order to elucidate the mechanism of the Rho pathway, we tested the effects of blocking the endogenous Rho-dependent kinase (ROCK) by anti-ROCK antibodies and Y-27632, (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl)cyclohexanecarboxamide dihydrochloride, a specific inhibitor of ROCK, on migrating amoebae and the effect of the Rho and ROCK inhibition on the actin-activated Mg-ATPase of the cytosolic fraction of the amoebae. Amoebae microinjected with anti-ROCK inhibitors remained contracted and strongly attached to the glass surface and exhibited an atypical locomotion. Despite protruding many pseudopodia that were advancing in various directions, the amoebae could not effectively move. Immunofluorescence studies showed that ROCK-like protein was dispersed throughout the cytoplasm and was also found in the regions of actin-myosin II interaction during both isotonic and isometric contraction. The Mg-ATPase activity was about two- to threefold enhanced, indicating that blocking the Rho/Rho-dependent kinase activated myosin. It is possible then that in contrast to the vertebrate cells, the inactivation of Rho/Rho-dependent kinase in amoebae leads to the activation of myosin II and to the observed hypercontracted cells which cannot exert effective locomotion.
机译：高度活动自由生活的单细胞生物变形虫变形杆菌已被广泛用作研究细胞运动性的模型。然而，其独特运动的分子机制仍然鲜为人知。最近，我们已经表明，阻断变形虫的内源性Rac和Rho样蛋白会导致这些大型迁移细胞的外观发生明显且不可逆的变化，并显着抑制其运动。为了阐明Rho途径的机制，我们测试了抗ROCK抗体和Y-27632，（+）-（R）-trans-4-（1）对内源性Rho依赖性激酶（ROCK）的阻断作用-氨基乙基）-N-（4-吡啶基）环己烷甲酰胺盐酸盐，一种特定的ROCK抑制剂，对阿米巴的迁移以及Rho和ROCK抑制作用对阿米巴细胞质中肌动蛋白激活的Mg-ATPase的影响。注射了抗ROCK抑制剂的变形虫保持收缩并牢固地附着在玻璃表面，并表现出非典型的运动。尽管突出了许多朝各个方向前进的假足，但变形虫无法有效地移动。免疫荧光研究表明，ROCK等蛋白分散在整个细胞质中，并且在等渗和等距收缩过程中也发现在肌动蛋白-肌球蛋白II相互作用的区域。 Mg-ATPase活性提高了约2到3倍，表明阻断Rho / Rho依赖性激酶激活了肌球蛋白。与脊椎动物细胞相反，变形虫中Rho / Rho依赖性激酶的失活可能导致肌球蛋白II的活化，并导致观察到的过度收缩的细胞无法发挥有效的运动能力。

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