Factors influencing the ability of nuclear localization sequence peptides to enhance nonviral gene delivery
Factors influencing the ability of nuclear localization sequence peptides to enhance nonviral gene delivery
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DOI：
10.1021/bc034140k
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发表时间：
2004-01-01
影响因子：
4.7
通讯作者：
Read, ML
中科院分区：
文献类型：
--
作者：
Bremner, KH;Seymour, LW;Read, ML
非病毒基因递送受到DNA从细胞质到细胞核的低效转移的限制。核定位序列（NLS）肽已被广泛用于开发细胞内转运机制和促进DNA的核摄取。然而，成功利用NLS肽特性的确切条件仍不清楚。在本研究中，比较了一组结合不同转运受体的NLS肽增强非病毒基因转移的能力。几个因素，如纳入NLS肽的方法，NLS肽的类型，DNA形态，和NLS肽/DNA缀合物的适当表征被确定为利用NLS肽增强基因转移的重要考虑因素。特别地，显示了来源于人T细胞白血病病毒1型（HTLV）的肽能够有效地将DNA浓缩成离散颗粒，并介导转基因表达水平比基于聚赖氨酸的多聚复合物高多达32倍。这是第一个研究，以证明有效的转染介导的importin β-结合肽的基础上HTLV序列。使用NLS肽/DNA缀合物也实现了有希望的结果，所述NLS肽/DNA缀合物通过肽核酸（PNA）夹将延伸的SV 40肽位点特异性连接而形成，基因表达增加7倍。总之，这项研究的结果应该有助于确定成功的NLS增强转染的要求。
Nonviral gene delivery is limited by inefficient transfer of DNA from the cytoplasm to the nucleus. Nuclear localization sequence (NLS) peptides have been widely used to exploit intracellular transport mechanisms and promote nuclear uptake of DNA. However, the exact conditions to successfully utilize the properties of NLS peptides are still unclear. In the present study a panel of NLS peptides that bind different transport receptors were compared for their ability to enhance nonviral gene transfer. Several factors such as method of incorporating the NLS peptide, type of NLS peptide, DNA morphology, and proper characterization of NLS peptide/DNA conjugates were identified as important considerations in utilizing NLS peptides to enhance gene transfer. In particular, it was shown that a peptide derived from human T cell leukaemia virus type 1 (HTLV) was able to effectively condense DNA into discrete particles and mediate levels of transgene expression up to 32-fold greater than polylysine-based polyplexes. This is the first study to demonstrate efficient transfection mediated by an importin beta-binding peptide based on the HTLV sequence. Promising results were also achieved with a 7-fold increase in gene expression using a NLS peptide/DNA conjugate formed by site-specific linkage of an extended SV40 peptide via a peptide nucleic acid (PNA) clamp. Altogether, the results from this study should help to define the requirements for successful NLS-enhanced transfection.