High density of octaarginine stimulates macropinocytosis leading to efficient intracellular trafficking for gene expression
High density of octaarginine stimulates macropinocytosis leading to efficient intracellular trafficking for gene expression
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DOI：
10.1074/jbc.m503202200
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发表时间：
2006-02-10
影响因子：
4.8
通讯作者：
Harashima, H
中科院分区：
文献类型：
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作者：
Khalil, IA;Kogure, K;Harashima, H
富精氨酸肽介导的细胞摄取机制目前是一个有争议的问题。包括肽的类型、货物的性质以及它们之间的连接体在内的几个因素似乎会影响吸收。可能影响摄取机制的较少研究的因素之一是肽密度对货物表面的影响。在这里，我们研究了细胞摄取和细胞内贩运的脂质体修饰的不同密度的八精氨酸（R8）肽的机制。用低R8密度修饰的脂质体主要通过网格蛋白介导的内吞作用被摄取，导致广泛的溶酶体降解，而用高R8密度修饰的脂质体主要通过巨胞饮作用被摄取，并且较少受到溶酶体降解。此外，高密度R8-脂质体能够刺激巨胞饮介导的其他颗粒的摄取。当质粒DNA浓缩并封装在R8-脂质体中时，基因表达水平比高密度脂质体高三个数量级。高密度R8-脂质体增强的基因表达通过阻断巨胞饮摄取而高度受损。来自脂质体上不同密度的R8肽的基因表达的不同程度可以主要由细胞内运输途径的存在来解释，而不是由内化脂质体的摄取量来解释。这些结果表明，脂质体上R8肽的密度决定了摄取机制，并且这与细胞内运输直接相关，导致不同水平的基因表达。
The mechanism of the arginine-rich peptide-mediated cellular uptake is currently a controversial issue. Several factors, including the type of peptide, the nature of the cargo, and the linker between them, appear to affect uptake. One of the less studied factors, which may affect the uptake mechanism, is the effect of peptide density on the surface of the cargo. Here, we examined the mechanism of cellular uptake and intracellular trafficking of liposomes modified with different densities of the octaarginine (R8) peptide. Liposomes modified with a low R8 density were taken up mainly through clathrin- mediated endocytosis, leading to extensive lysosomal degradation, whereas those modified with a high R8 density were taken up mainly through macropinocytosis and were less subject to lysosomal degradation. Furthermore, the high density R8-liposomes were able to stimulate the macropinocytosis- mediated uptake of other particles. When plasmid DNA was condensed and encapsulated in R8-liposomes, the levels of gene expression were three orders of magnitude higher for the high density liposomes. The enhanced gene expression by the high density R8- liposomes was highly impaired by blocking uptake through macropinocytosis. The different extents of gene expression from different densities of the R8 peptide on the liposomes could be explained principally by the existence of an intracellular trafficking route, but not by the uptake amount, of internalized liposomes. These results show that the density of the R8 peptide on liposomes determines the uptake mechanism and that this is directly linked to intracellular trafficking, resulting in different levels of gene expression.