Wölfl S, Wittig B, Dorbic T, Rich A
Massachusetts Institute of Technology, Department of Biology, Cambridge, USA.
Biochim Biophys Acta. 1997 May 30;1352(2):213-21. doi: 10.1016/s0167-4781(97)00015-8.
DNA elements with sequences suitable for Z-DNA formation are found frequently at various positions in chromatin. Z-DNA formation in these sequences depends largely on the level of local negative supercoiling. We can use binding of a Z-DNA specific antibody at low concentrations in metabolically active permeabilized nuclei to detect naturally occurring Z-DNA formation. Previously we identified three sequence elements in the human c-myc gene that adopt the Z-DNA conformation in the transcribed gene. The three elements are found far upstream (Z1), close to the main transcription start site (Z2) and in the first intron (Z3). Here we measure the persistence of Z-DNA at these three sites under the influence of various metabolic inhibitors. This provides some insight into the varying levels of negative supercoiling. alpha-Amanitin, an inhibitor of transcription, reduced the persistence of Z-DNA in all three elements. Aphidicolin, an inhibitor of replication, increased the persistence of Z-DNA in one element without significantly influencing the other two elements. When camptothecin an inhibitor of topoisomerase I was added in the presence of alpha-amanitin, the persistence of Z-DNA was extended in all three elements. However, in the presence of aphidicolin no effect of camptothecin on Z-DNA formation was observed.
具有适合形成Z-DNA序列的DNA元件在染色质的不同位置频繁出现。这些序列中Z-DNA的形成很大程度上取决于局部负超螺旋的水平。我们可以在代谢活跃的通透细胞核中使用低浓度的Z-DNA特异性抗体结合来检测天然存在的Z-DNA形成。此前我们在人类c-myc基因中鉴定出三个序列元件，它们在转录基因中采用Z-DNA构象。这三个元件分别位于远上游（Z1）、靠近主要转录起始位点（Z2）以及第一个内含子中（Z3）。在这里，我们测量了在各种代谢抑制剂影响下这三个位点Z-DNA的持续性。这为负超螺旋水平的变化提供了一些见解。α-鹅膏蕈碱是一种转录抑制剂，它降低了所有三个元件中Z-DNA的持续性。阿非迪霉素是一种复制抑制剂，它增加了一个元件中Z-DNA的持续性，而对其他两个元件没有显著影响。当在存在α-鹅膏蕈碱的情况下添加拓扑异构酶I抑制剂喜树碱时，所有三个元件中Z-DNA的持续性都延长了。然而，在存在阿非迪霉素的情况下，未观察到喜树碱对Z-DNA形成有影响。
