Analysis of nuclear actin by overexpression of wild-type and actin mutant proteins
Analysis of nuclear actin by overexpression of wild-type and actin mutant proteins
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DOI：
10.1007/s00418-013-1151-4
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发表时间：
2014-02-01
影响因子：
2.3
通讯作者：
Knoell, Bernd
中科院分区：
文献类型：
--
作者：
Kokai, Enikoe;Beck, Henning;Knoell, Bernd
与细胞质中f -肌动蛋白的丰度相比，细胞核中的f -肌动蛋白水平通常很低。然而，核肌动蛋白存在于某些细胞类型中，包括卵母细胞，以及在某些细胞条件下，包括应激或血清刺激。目前，对核肌动蛋白网络的结构和聚合状况还没有详细的分析。在这项研究中，我们研究了这种核肌动蛋白网络的结构和功能。我们通过融合核定位信号（NLS）的肌动蛋白过表达生成核肌动蛋白聚合物。提高NLS野生型肌动蛋白的核丰度，我们观察到phalloidin和lifeact阳性肌动蛋白束形成由弯曲的f -肌动蛋白组成的核细胞骨架网络。相反，聚合物稳定肌动蛋白突变体（NLS-G15S-actin）缺乏与肌动蛋白结合蛋白cofilin的相互作用，产生核肌动蛋白网络，使人联想到细胞质中的直应力纤维样微丝。我们提供了这种核肌动蛋白聚合物的第一个电子显微镜描述，表明肌动蛋白细丝的捆绑。利用包括神经元在内的不同种类的细胞，我们发现核肌动蛋白的形态和产生潜能是保守的。最后，我们证明核肌动蛋白影响细胞功能，包括形态、血清反应因子介导的基因表达和单纯疱疹病毒感染。我们的数据表明，肌动蛋白能够在细胞核内形成丝状结构，其结构和功能与胞质f -肌动蛋白相似。
Compared to the cytoplasmic F-actin abundance in cells, nuclear F-actin levels are generally quite low. However, nuclear actin is present in certain cell types including oocytes and under certain cellular conditions including stress or serum stimulation. Currently, the architecture and polymerization status of nuclear actin networks has not been analyzed in great detail. In this study, we investigated the architecture and functions of such nuclear actin networks. We generated nuclear actin polymers by overexpression of actin proteins fused to a nuclear localization signal (NLS). Raising nuclear abundance of a NLS wild-type actin, we observed phalloidin- and LifeAct-positive actin bundles forming a nuclear cytoskeletal network consisting of curved F-actin. In contrast, a polymer-stabilizing actin mutant (NLS-G15S-actin) deficient in interacting with the actin-binding protein cofilin generated a nuclear actin network reminiscent of straight stress fiber-like microfilaments in the cytoplasm. We provide a first electron microscopic description of such nuclear actin polymers suggesting bundling of actin filaments. Employing different cell types from various species including neurons, we show that the morphology of and potential to generate nuclear actin are conserved. Finally, we demonstrate that nuclear actin affects cell function including morphology, serum response factor-mediated gene expression, and herpes simplex virus infection. Our data suggest that actin is able to form filamentous structures inside the nucleus, which share architectural and functional similarities with the cytoplasmic F-actin.