A gene delivery system containing nuclear localization signal: Increased nucleus import and transfection efficiency with the assistance of RanGAP1
A gene delivery system containing nuclear localization signal: Increased nucleus import and transfection efficiency with the assistance of RanGAP1
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DOI：
10.1016/j.actbio.2016.11.004
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发表时间：
2017-01-15
影响因子：
9.7
通讯作者：
Chen, Dawei
中科院分区：
文献类型：
--
作者：
Chen, Kang;Guo, Lingling;Chen, Dawei
制备了一种含核定位信号肽（NLS）的可降解基因传递系统（PAMS/DNA/10 NLS）。PAMS/DNA/10 NLS的琼脂糖凝胶电泳、粒径和zeta电位与PAMS/DNA相似，证明NLS不影响PAMS与DNA的相互作用。PAMS/DNA/10 NLS在酸性条件下表现出明显的细胞外和细胞内降解。这种降解被认为使NLS容易与importins接触，从而能够介导核输入。PAMS/ DNA/10 NLS在NLS的辅助下，转染能力明显高于PAMS/DNA。此外，PAMS/DNA/10 NLS的转染比PAMS/DNA更不依赖于核膜的破坏。考虑到GTP酶激活蛋白1（RanGAP 1）能够激活NLS介导的核输入所必需的内源性GTP酶，产生RanGAP 1过表达的细胞（RanGAP 1细胞）。该结果表明，RanGAP 1细胞比正常细胞具有更高的GT3活性。PAMS/DNA/10 NLS在RanGAP 1细胞中的核导入率和转染效率均明显高于正常细胞。体内转染结果也显示PAMS/DNA/10 NLS在RanGAP 1预处理小鼠中的转染效率高于正常小鼠。这些结果表明，PAMS/DNA/10 NLS是一个有前途的基因传递系统的辅助下，RanGAP 1。本报告描述了含有核定位信号（NLS）的可降解基因递送系统（PAMS/DNA/10 NLS）在GTP酶激活蛋白1（RanGAP 1）的帮助下提高的转染效率。PAMS/DNA/10 NLS的理化性质与PAMS/DNA相似。PAMS/DNA/10 NLS具有较强的细胞外和细胞内降解能力，这可能使NLS更容易与importins接触。PAMS/DNA/10 NLS在NLS的辅助下，转染能力明显高于PAMS/DNA。PAMS/DNA/10 NLS转染对核膜破裂的依赖性较小。PAMS/ DNA/10 NLS在RanGAP 1过表达细胞中的核导入率和转染效率均高于正常细胞。PAMS/DNA/lONLS转染RanGAP 1小鼠的效率高于正常小鼠。(C)2016 Acta Materialia Inc.由爱思唯尔有限公司出版。保留所有权利。
In the present report, a degradable gene delivery system (PAMS/DNA/1 0NLS) containing nucleus location signal peptide (NLS) was prepared. The agarose gel electrophoresis, particle size and zeta potential of PAMS/DNA/10NLS were similar to those of PAMS/DNA, which proved that NLS did not affect the interaction between PAMS and DNA. PAMS/DNA/10NLS exhibited marked extracellular and intracellular degradation under acidic conditions. The degradation was believed to allow NLS to come into contact with importins easily, which was able to mediate the nucleus import. With the help of NLS, PAMS/ DNA/10NLS exhibited a higher transfection capability than PAMS/DNA. Moreover, the transfection of PAMS/DNA/10NLS was less dependent on the breakdown of the nucleus envelope than PAMS/DNA. Considering that GTPase-activating protein 1 (RanGAP1) was able to activate the endogenous GTPase, which was necessary for NLS-mediated nucleus import, RanGAP1 overexpressed cells (RanGAP1 cells) were produced. This result showed that RanGAP1 cells had higher GTPase activities than normal cells. Both the nucleus import and transfection efficiency of PAMS/DNA/10NLS were markedly higher in RanGAP1 cells than that in normal cells. The in vivo transfection results also showed that the transfection efficiency of PAMS/DNA/10NLS was higher in RanGAP1 pre-treated mice than that in normal mice. These findings showed that PAMS/DNA/10NLS is a promising gene delivery system with the assistance of RanGAP1. Statement of Significance The present report describes the increased transfection efficiency of a degradable gene delivery system (PAMS/DNA/10NLS) containing nuclear location signal (NLS) with the assistance of GTPase-activating protein 1 (RanGAP1). The physicochemical properties of PAMS/DNA/10NLS were similar to those of PAMS/DNA. PAMS/DNA/10NLS exhibited great extracellular and intracellular degradations, which might allow NLS to contact with importins easily. With the help of NLS, PAMS/DNA/10NLS exhibited a higher transfection capability than PAMS/DNA. The transfection of PAMS/DNA/10NLS had less dependence on the breakdown of nuclear envelope. Both the nuclear import and transfection efficiency of PAMS/ DNA/10NLS were higher in RanGAP1 overexpressed cells than that in normal cells. Moreover, the transfection efficiency of PAMS/DNA/lONLS was higher in RanGAP1 pre-treated mice than that in normal mice. (C) 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.