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首页> 外文期刊>Journal of Molecular Biology >Initiation of DNA replication in a eukaryotic rolling-circle replicon: identification of multiple DNA-protein complexes at the geminivirus origin.
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Initiation of DNA replication in a eukaryotic rolling-circle replicon: identification of multiple DNA-protein complexes at the geminivirus origin.

机译：在真核滚环复制子中DNA复制的启动：在双生病毒起源处鉴定多个DNA-蛋白质复合物。

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摘要

The mechanism of initiation of DNA replication in eukaryotic rolling- circle replicons is still poorly understood in molecular terms. Geminiviruses, a family of plant DNA viruses, which use this strategy during part of their replicative cycle, replicate in the nucleus and are amenable to molecular studies. Except for the virally encoded initiator protein (Rep), geminivirus DNA replication relies on cellular factors, likely interfering with cell cycle regulation of the infected cell. Here, we report the identification of three distinct DNA-protein complexes of the DNA replication initiator protein encoded by wheat dwarf geminivirus (WDV) within viral regulatory sequences controlling DNA replication and transcription. We have mapped the WDV Rep binding sites by combining gel-shift assays, electron microscopy and DNase I footprinting. Two of the Rep-DNA complexes (C and the V) are high-affinity complexes, located in the proximity of the two divergent TATA boxes, at 150 and 90 bp, respectively, from the DNA replication initiation site. The third one, the O-complex, is a low-affinity complex, which can assemble under conditions supporting the DNA cleavage reaction. This suggests that it might be responsible for initiation of rolling-circle DNA replication in WDV and other members of the Mastrevirus genus. Copyright 1999 Academic Press.
机译：真核生物转环复制子中DNA复制的起始机制在分子学上仍知之甚少。双子病毒是一种植物DNA病毒家族，在其复制周期的一部分过程中使用此策略，可在细胞核中复制并适合分子研究。除了病毒编码的启动子蛋白（Rep）外，双生病毒DNA复制还依赖于细胞因子，可能会干扰受感染细胞的细胞周期调控。在这里，我们报告鉴定了由小麦矮生双生病毒（WDV）编码的控制DNA复制和转录的病毒调控序列中的DNA复制起始蛋白的三个不同的DNA-蛋白质复合物。我们通过结合凝胶位移测定，电子显微镜和DNase I足迹绘制了WDV Rep结合位点。 Rep-DNA复合物中的两个（C和V）是高亲和力复合物，位于两个不同的TATA盒附近，分别距DNA复制起始位点150和90 bp。第三个是O-复合物，是一种低亲和力的复合物，可以在支持DNA裂解反应的条件下组装。这表明它可能是导致WDV和Mastrevirus属的其他成员中滚环DNA复制的开始。版权所有1999，学术出版社。

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