McHugh M M, Beerman T A, Burhans W C
Department of Experimental Therapeutics, Roswell Park Cancer Institute, Buffalo, New York 14263, USA.
Biochemistry. 1997 Feb 4;36(5):1003-9. doi: 10.1021/bi962121a.
This study used 2-D agarose gel techniques to examine the effects of the DNA-strand scission enediyne C-1027 on DNA replication in SV40-infected BSC-1 cells. Replication of SV40 DNA was inhibited by C-1027 to a greater extent than was BSC-1 genomic DNA replication in infected cells. Low nanomolar concentrations (0.2-10 nM) of C-1027 affected a rapid, progressive decrease in SV40 replication activity and replication intermediates (RIs) within 15 min after drug addition. A concurrent decrease in the signal of both the SV40 bubble arc and replication activity with increasing concentrations of C-1027 suggested that C-1027 inhibited initiation of new RIs. Additionally, the reduction in bubble arc signal observed with C-1027 was prevented when elongation of nascent chains was blocked by aphidicolin. Thus, the C-1027-induced disappearance of RIs probably is related to the maturation of preformed replication molecules in the absence of initiation of new RIs. Strand damage to SV40 DNA was barely detectable at concentrations where inhibition of replication activity was nearly complete, indicating that C-1027 replication inhibition occurs in trans.
本研究采用二维琼脂糖凝胶技术，检测DNA链断裂烯二炔C-1027对感染SV40的BSC-1细胞中DNA复制的影响。与感染细胞中BSC-1基因组DNA复制相比，C-1027对SV40 DNA复制的抑制作用更强。低纳摩尔浓度（0.2 - 10 nM）的C-1027在添加药物后15分钟内，使SV40复制活性和复制中间体（RI）迅速、逐渐降低。随着C-1027浓度增加，SV40泡状弧信号和复制活性同时降低，这表明C-1027抑制了新RI的起始。此外，当新生链延伸被阿非迪霉素阻断时，C-1027所观察到的泡状弧信号降低被阻止。因此，C-1027诱导的RI消失可能与在无新RI起始情况下预先形成的复制分子的成熟有关。在复制活性抑制几乎完全的浓度下，几乎检测不到SV40 DNA的链损伤，这表明C-1027的复制抑制作用是反式发生的。
