Gene delivery: A single nuclear localization signal peptide is sufficient to carry DNA to the cell nucleus

被引:617
作者
Zanta, MA [1 ]
Belguise-Valladier, P [1 ]
Behr, JP [1 ]
机构
[1] Univ Strasbourg, Fac Pharm, Lab Chim Genet, CNRS,UMR 7514, F-67070 Strasbourg, France
关键词
D O I
10.1073/pnas.96.1.91
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Translocation of exogenous DNA through the nuclear membrane is a major concern of gene delivery technologies. To take advantage of the cellular import machinery, we have synthesized a capped 3.3-kbp CMVLuciferase-NLS gene containing a single nuclear localization signal peptide (PKKKRKVEDPYC). Transfection of cells with the tagged gene remained effective down to nanogram amounts of DNA. Transfection enhancement (10- to 1,000-fold) as a result of the signal peptide was observed irrespective of the cationic vector or the cell type used. A lysine to threonine mutation of the third NLS amino acid completely abolished these remarkable features, suggesting importin-mediated translocation. Our hypothesis is that the 3-nm-wide DNA present in the cytoplasm is initially docked to and translocated through a nuclear pore by the nuclear import machinery. As DNA enters the nucleus, it is quickly condensed into a chromatin-like structure, which provides a mechanism for threading the remaining worm-like molecule through the pore. A single NLS signal is thus sufficient, whereas many signals on a gene would actually inhibit entry, the same DNA molecule being threaded through adjacent pores.
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页码:91 / 96
页数:6
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