Microtubule involvement in the intracellular dynamics for gene transfection mediated by cationic liposomes
Microtubule involvement in the intracellular dynamics for gene transfection mediated by cationic liposomes
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DOI：
10.1038/sj.gt.3301573
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发表时间：
2001-11-01
期刊：
影响因子：
5.1
通讯作者：
Nakanishi, M
中科院分区：
文献类型：
--
作者：
Hasegawa, S;Hirashima, N;Nakanishi, M
用激光共聚焦显微镜观察微管聚合对脂质体介导的基因转染的影响。诺考达唑和紫杉醇均能明显提高基因转染效率。用荧光标记的阳离子脂质体在表达黄色荧光蛋白（YFP）标记的微管蛋白的COS-7细胞中进行脂质体转染，揭示脂质体沿着微管转运到与微管组织中心（MTOC）共定位的溶酶体。诺考达唑破坏微管并在细胞质中产生均匀分布的YFP标记的微管蛋白。在这些条件下，脂质体和溶酶体都分散在整个细胞质中，并且它们不共定位。在紫杉醇的存在下，微管被稳定化，并形成了几个焦点区域，如MTOC。溶酶体位于细胞核周围，而脂质体则被困在微管中。在这些条件下，脂质体和DNA都不与溶酶体共定位。这些结果表明，脂质体-DNA复合物通过微管介导的途径转运到溶酶体，诺考达唑和紫杉醇对转染效率的影响可以通过脂质体-DNA复合物转运到溶酶体的失败来解释，其中DNA被降解。
The effects of microtubule polymerization on liposome-mediated gene transfection were investigated by confocal laser scanning microscopy in target living cells. Both nocodazole and taxol apparently increased the efficiency of gene transfection. Lipofection with fluorescence-labeled cationic liposomes in a COS-7 cell expressing yellow fluorescent protein (YFP)-tagged tubulin revealed that the liposomes were transported along microtubules to lysosomes which are colocalized with the microtubule organizing center (MTOC). Nocodazole disrupted microtubules and produced a uniform distribution of YFP-tagged tubulin in the cytoplasm. Under these conditions, both liposomes and lysosomes were scattered throughout the cytoplasm and they did not colocalize. In the presence of taxol, microtubules were stabilized and several focal regions, like the MTOC, were formed. Lysosomes resided around the nucleus, while liposomes were trapped in microtubules. Under these conditions, neither liposomes nor DNA colocalized with lysosomes. These results demonstrated that the liposome-DNA complexes are transported to lysosomes by a microtubule-mediated pathway, and the effects of nocodazole and taxol on transfection efficiency can be explained by failure of the transport of the liposome-DNA complexes to lysosomes where DNAs are degraded.