Epiplakin modifies the motility of the HeLa cells and accumulates at the outer surfaces of 3-D cell clusters
Epiplakin modifies the motility of the HeLa cells and accumulates at the outer surfaces of 3-D cell clusters
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DOI：
10.1111/1346-8138.12076
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发表时间：
2013-04-01
影响因子：
3.1
通讯作者：
Fujiwara, Sakuhei
中科院分区：
文献类型：
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作者：
Shimada, Hiromitsu;Nambu-Niibori, Akiko;Fujiwara, Sakuhei
在创伤愈合过程中，通过基因打靶消除表皮样蛋白(EPPK)可加速角质形成细胞的迁移，提示上皮细胞EPPK可能在细胞运动调节中起重要作用。为了研究EPPK的功能，我们建立了EPPK敲除(KD)和EPPK高表达的HeLa细胞，并利用荧光/差分干涉对比时间推移显微镜和肌动蛋白和波形蛋白的免疫定位分析了细胞的表型和运动能力。EPPK-KD细胞活力显著升高，而EPPK过表达细胞活力明显降低。EPPK-KD细胞上有大量的棘状突起，高表达细胞上的棘状突起较少。在EPPK-KD细胞中，E-钙粘附素的表达与对照组相比没有变化，但波形蛋白纤维较对照组更细、更稀疏，并且更集中在核周围，正如EPPK/小鼠创伤边缘的迁移角质形成细胞所观察到的那样。在Matrigel 3D培养中，EPPK与紧密连接的标志ZO-1共定位于细胞团的外表面。我们的结果表明，EPPK与细胞运动机制有关，并通过中间细丝的重排对组织结构做出贡献。
Elimination of epiplakin (EPPK) by gene targeting in mice results in acceleration of keratinocyte migration during wound healing, suggesting that epithelial cellular EPPK may be important for the regulation of cellular motility. To study the function of EPPK, we developed EPPK knock-down (KD) and EPPK-overexpressing HeLa cells and analyzed cellular phenotypes and motility by fluorescence/differential interference contrast time-lapse microscopy and immunolocalization of actin and vimentin. Cellular motility of EPPK-KD cells was significantly elevated, but that of EPPK-overexpressing cells was obviously depressed. Many spike-like projections were observed on EPPK-KD cells, with fewer such structures on overexpressing cells. By contrast, in EPPK-KD cells, expression of E-cadherin was unchanged but vimentin fibers were thinner and sparser than in controls, and they were more concentrated at the peri-nucleus, as observed in migrating keratinocytes at wound edges in EPPK/ mice. In Matrigel 3-D cultures, EPPK co-localized on the outer surface of cell clusters with zonula occludens-1 (ZO-1), a marker of tight junctions. Our results suggest that EPPK is associated with the machinery for cellular motility and contributes to tissue architecture via the rearrangement of intermediate filaments.