Belikov S V, Grokhovsky S L, Isaguliants M G, Surovaya A N, Gursky G V
Department of Cell and Molecular Biology, The Medical Nobel Institute, Karolinska Institute, SE 17177, Stockholm, Sweden.
J Biomol Struct Dyn. 2005 Oct;23(2):193-202. doi: 10.1080/07391102.2005.10507059.
The mouse mammary tumor virus (MMTV) promoter is induced by glucocorticoid hormone. A robust hormone- and receptor-dependent gene activation could be reproduced in Xenopus laevis oocytes. The homogeneous response in this system allowed a detailed analysis of the DNA-protein interactions following hormone activation. The strategy of artificial regulating of gene activity by sequence-specific minor groove binding ligands is very attractive. We have synthesized and studied the interaction with DNA of bis-linked netropsin derivatives in which two monomers are attached via short linkers in head-to-head and tail-to-tail manners. We have found that cis-diammine-platinum bridged bis-netropsin added to Xenopus oocytes media penetrates cellular and nuclear membrane and binds selectively to the MMTV promoter at the DNA segment that partly overlaps with the site recognized by glucocorticoid receptor. DNase I footprinting studies demonstrate that there are more stronger binding sites for cis-diammine-platinum bridged bis-netropsin on the naked MMTV DNA which are found to be inaccessible for its binding in oocytes.
小鼠乳腺肿瘤病毒（MMTV）启动子受糖皮质激素诱导。在非洲爪蟾卵母细胞中可重现强大的激素和受体依赖性基因激活。该系统中的均匀反应允许对激素激活后DNA-蛋白质相互作用进行详细分析。通过序列特异性小沟结合配体人工调节基因活性的策略非常有吸引力。我们合成并研究了双连接的纺锤菌素衍生物与DNA的相互作用，其中两个单体通过短连接子以头对头和尾对尾的方式连接。我们发现添加到非洲爪蟾卵母细胞培养基中的顺二氨铂桥连双纺锤菌素穿透细胞膜和核膜，并在与糖皮质激素受体识别位点部分重叠的DNA片段上选择性地与MMTV启动子结合。DNase I足迹研究表明，在裸露的MMTV DNA上有更多更强的顺二氨铂桥连双纺锤菌素结合位点，而这些位点在卵母细胞中被发现无法结合。
