Sequence-specific gene silencing in mammalian cells by alkylating pyrrole-imidazole polyamides.
Gene silencing was examined by sequence-specific alkylation of DNA by N-methylpyrrole (Py)-N-methylimidazole (Im) hairpin polyamides. Polyamides ImImPyPygammaImImPyLDu86 (A) and ImImPyPygammaImPyPyLDu86 (B) selectively alkylated the coding regions of the renilla and firefly luciferases, respectively, according to the base pair recognition rule of Py-Im polyamides. Two different plasmids, encoding renilla luciferase and firefly luciferase, were used as vectors to examine the effect of alkylation on gene silencing. Transfection of the alkylated luciferase vectors-by polyamide A or B-into HeLa, 293, and NIH3T3 cells demonstrated that these sequence-specific DNA alkylations lead to selective silencing of gene expression. Next, the vectors were cotransfected into HeLa cells and the cells were treated with polyamide A or B. Selective reduction of luciferase activities was caused by both polyamides. On the basis of this sequence-specific alkylation and gene silencing activity, these alkylating Py-Im polyamides thus have potential as antitumor drugs to target specific gene expression in human cells.
Shinohara
Ken-Ichi
K
Department of Pathological Biochemistry, Medical Research Institute, Tokyo Medical and Dental University, Chiyoda-ku, Tokyo 101-0062, Japan.
Narita
Akihiko
A
Oyoshi
Takanori
T
Bando
Toshikazu
T
Teraoka
Hirobumi
H
Sugiyama
Hiroshi
H
D000478:Alkylation; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004247:DNA / Q000737:chemistry*; D015870:Gene Expression; D020868:Gene Silencing; D006367:HeLa Cells; D006801:Humans; D007093:Imidazoles / Q000737:chemistry*; D008156:Luciferases / Q000737:chemistry / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D015394:Molecular Structure; D041681:NIH 3T3 Cells; D009690:Nucleic Acid Conformation; D009757:Nylons / Q000737:chemistry*; D011758:Pyrroles / Q000737:chemistry*
D007093:Imidazoles; D009757:Nylons; D011758:Pyrroles; D004247:DNA; D008156:Luciferases