Intranuclear protein transduction through a nucleoside salvage pathway
Intranuclear protein transduction through a nucleoside salvage pathway
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DOI：
10.1074/jbc.c700090200
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发表时间：
2007-07-20
影响因子：
4.8
通讯作者：
Weisbart, Richard H.
中科院分区：
文献类型：
--
作者：
Hansen, James E.;Tse, Chung-Ming;Weisbart, Richard H.
通过转录因子等大分子的核内转导来调节基因表达是治疗许多疾病的一种替代基因疗法。然而，确定一种有效的核内递送载体和治疗性大分子通过血浆和核膜运输的途径，已经提出了一个重大的挑战。抗dna抗体片段3E10 Fv作为一种新型的分子递送载体受到关注，因为它具有特异性核定位的渗透到活细胞中，没有毒性，并且在体外和体内成功递送治疗货物蛋白。阐明允许3E10 Fv穿过细胞膜的途径对于开发新的分子疗法至关重要。在这里，我们表明3E10 Fv通过核苷回收转运体穿透细胞。3E10 Fv不能渗透到缺乏平衡核苷转运体ENT2的细胞中，将ENT2重组到缺乏ENT2的细胞中可以恢复3E10 Fv转运到细胞核中。我们的研究结果首次证明了蛋白质通过核苷回收途径进行转运。我们期望我们的发现将促进多种基因调控方法在人类疾病的治疗中，为核苷挽救途径的研究开辟新的途径，并增强我们对自身免疫性疾病病理生理学的理解。
Regulation of gene expression by intranuclear transduction of macromolecules such as transcription factors is an alternative to gene therapy for the treatment of numerous diseases. The identification of an effective intranuclear delivery vehicle and pathway for the transport of therapeutic macromolecules across plasma and nuclear membranes, however, has posed a significant challenge. The anti-DNA antibody fragment 3E10 Fv has received attention as a novel molecular delivery vehicle due to its penetration into living cells with specific nuclear localization, absence of toxicity, and successful delivery of therapeutic cargo proteins in vitro and in vivo. Elucidation of the pathway that allows 3E10 Fv to cross cell membranes is critical to the development of new molecular therapies. Here we show that 3E10 Fv penetrates cells through a nucleoside salvage transporter. 3E10 Fv is unable to penetrate into cells deficient in the equilibrative nucleoside transporter ENT2, and reconstitution of ENT2 into ENT2-deficient cells restores 3E10 Fv transport into cell nuclei. Our results represent the first demonstration of protein transport through a nucleoside salvage pathway. We expect that our finding will facilitate a variety of methods of gene regulation in the treatment of human diseases, open up new avenues of research in nucleoside salvage pathways, and enhance our understanding of the pathophysiology of autoimmune diseases.