Getting loaded—make mine a double! Chromosomal DNA replication initiates bidirectionally by loading two ring-shaped helicases onto DNA in opposite orientations. How this symmetry is achieved has been puzzling because replication initiation sites contain only one essential binding site for the initiator, the origin recognition complex (ORC). Coster and Diffley now show that both helicases are loaded by a similar mechanism. Efficient loading requires binding of two ORC complexes to two ORC binding sites in opposite orientations. Natural origins were found to be partially symmetrical, containing functionally relevant secondary ORC sites. Sites can be flexibly spaced, but introducing an intervening “roadblock” prevented loading, suggesting that individual helicases translocate toward each other on DNA to form a stable double ring. Science, this issue p. 314 The minichromosome maintenance double hexamer, a precursor of the replicative DNA helicase, results from a symmetrical loading reaction. Bidirectional replication from eukaryotic DNA replication origins requires the loading of two ring-shaped minichromosome maintenance (MCM) helicases around DNA in opposite orientations. MCM loading is orchestrated by binding of the origin recognition complex (ORC) to DNA, but how ORC coordinates symmetrical MCM loading is unclear. We used natural budding yeast DNA replication origins and synthetic DNA sequences to show that efficient MCM loading requires binding of two ORC molecules to two ORC binding sites. The relative orientation of these sites, but not the distance between them, was found to be critical for MCM loading in vitro and origin function in vivo. We propose that quasi-symmetrical loading of individual MCM hexamers by ORC and directed MCM translocation into double hexamers acts as a unifying mechanism for the establishment of bidirectional replication in archaea and eukaryotes.

中文翻译：

---

通过准对称解旋酶加载建立双向真核 DNA 复制


装满东西——让我的变成双份！染色体 DNA 复制通过将两个环形解旋酶以相反方向加载到 DNA 上来启动双向复制。这种对称性是如何实现的一直令人困惑，因为复制起始位点仅包含起始子的一个重要结合位点，即起点识别复合物（ORC）。科斯特和迪夫利现在表明，两种解旋酶都是通过类似的机制加载的。有效加载需要两个 ORC 复合物以相反方向结合到两个 ORC 结合位点。发现天然起源是部分对称的，包含功能相关的次级 ORC 位点。位点可以灵活地间隔，但引入中间“路障”阻止了加载，这表明各个解旋酶在 DNA 上相互易位，形成稳定的双环。科学，本期第 14 页。 314 微型染色体维持双六聚体是复制 DNA 解旋酶的前体，由对称加载反应产生。从真核 DNA 复制起点进行的双向复制需要在 DNA 周围以相反方向加载两个环形微型染色体维持 (MCM) 解旋酶。 MCM 加载是通过起源识别复合物 (ORC) 与 DNA 的结合来协调的，但 ORC 如何协调对称 MCM 加载尚不清楚。我们使用天然芽殖酵母 DNA 复制起点和合成 DNA 序列来表明，有效的 MCM 加载需要两个 ORC 分子与两个 ORC 结合位点的结合。这些位点的相对方向，而不是它们之间的距离，被发现对于体外 MCM 负载和体内起源功能至关重要。 我们提出，ORC 对单个 MCM 六聚体的准对称负载和定向 MCM 易位成双六聚体可作为在古细菌和真核生物中建立双向复制的统一机制。