Hosoya T, Takeuchi H, Kanesaka Y, Yamakawa H, Miyano-Kurosaki N, Takai K, Yamamoto N, Takaku H
Department of Industrial Chemistry, Chiba Institute of Technology, Tsudanuma, Narashino, Chiba, Japan.
FEBS Lett. 1999 Nov 19;461(3):136-40. doi: 10.1016/s0014-5793(99)01450-7.
The inhibition of specific transcription regulatory proteins is a new approach to control gene expression. The transcriptional activities of DNA-binding proteins can be inhibited by the use of double-stranded oligonucleotides that compete for the binding to their specific target sequences in promoters and enhancers. We used nicked (NDODN-kappaB) and circular (CDODN-kappaB) dumbbell DNA oligonucleotides containing a NF-kappaB binding site to analyze the inhibition of the NF-kappaB-dependent activation of the human immunodeficiency virus type-1 (HIV-1) enhancer. The dumbbell DNA oligonucleotides are stable, short segments of double-stranded DNA with closed nucleotide loops on each end, which confer resistance to exonucleases. The dumbbell and other oligonucleotides (decoys) with the NF-kappaB sequence were found to compete with the native strand for NF-kappaB binding. The circular dumbbell and double-stranded phosphorothioate oligonucleotides competed with the native strand for binding to the NF-kappaB binding proteins, while the nicked NF-kappaB dumbbell was a less effective competitor. In Jurkat T-cells, the dumbbell and other oligonucleotides were tested for their ability to block the activation of the plasmid HIV-NL4-3 Luc. The CDODN-kappaB strongly inhibits the specific transcriptional regulatory proteins, as compared with the NDODN-kappaB and the double stranded phosphodiester oligonucleotides. On the other hand, the double stranded phosphorothioate oligonucleotides could also block this activation, but the effect was non-specific. The circular (CDODN) dumbbell oligonucleotides may efficiently compete for the binding of specific transcription factors within cells, thus providing anti-HIV-1 or other therapeutic effects.
抑制特定转录调节蛋白是控制基因表达的一种新方法。通过使用双链寡核苷酸可抑制DNA结合蛋白的转录活性，这些双链寡核苷酸可竞争结合启动子和增强子中的特定靶序列。我们使用了含有NF-κB结合位点的带切口哑铃状DNA寡核苷酸（NDODN-κB）和环状哑铃状DNA寡核苷酸（CDODN-κB）来分析对人免疫缺陷病毒1型（HIV-1）增强子NF-κB依赖性激活的抑制作用。哑铃状DNA寡核苷酸是双链DNA的稳定短片段，两端带有封闭的核苷酸环，可抵抗核酸外切酶。发现带有NF-κB序列的哑铃状及其他寡核苷酸（诱饵）与天然链竞争NF-κB结合。环状哑铃状和双链硫代磷酸酯寡核苷酸与天然链竞争结合NF-κB结合蛋白，而带切口的NF-κB哑铃状寡核苷酸是效果较差的竞争者。在Jurkat T细胞中，测试了哑铃状及其他寡核苷酸阻断质粒HIV-NL4-3 Luc激活的能力。与NDODN-κB和双链磷酸二酯寡核苷酸相比，CDODN-κB强烈抑制特定转录调节蛋白。另一方面，双链硫代磷酸酯寡核苷酸也可阻断这种激活，但效果是非特异性的。环状（CDODN）哑铃状寡核苷酸可有效竞争细胞内特定转录因子的结合，从而提供抗HIV-1或其他治疗效果。
