Abstract：
The numerous enzymes and cofactors involved in eukaryotic DNA replication are conserved from yeast to human, and the budding yeast Saccharomyces cerevisiae (S.c.) has been a useful model organism for these studies. However, there is a gap in our knowledge of why replication origins in higher eukaryotes do not use a consensus DNA sequence as found in S.c. Using in vitro reconstitution and single-molecule visualization, we show here that S.c. origin recognition complex (ORC) stably binds nucleosomes and that ORC-nucleosome complexes have the intrinsic ability to load the replicative helicase MCM double hexamers onto adjacent nucleosome-free DNA regardless of sequence. Furthermore, we find that Xenopus laevis nucleosomes can substitute for yeast ones in engaging with ORC. Combined with re-analyses of genome-wide ORC binding data, our results lead us to propose that the yeast origin recognition machinery contains the cryptic capacity to bind nucleosomes near a nucleosome-free region and license origins, and that this nucleosome-directed origin licensing paradigm generalizes to all eukaryotes.
摘要：
参与真核DNA复制的许多酶和辅因子从酵母到人都是保守的，出芽酵母酿酒酵母（S.c.）一直是这些研究的有用模型生物。然而,我们对高等真核生物中的复制起点为何不使用S.C.中发现的共有DNA序列的认识存在差距。使用体外重建和单分子可视化，我们在这里显示S.c.起源识别复合物（ORC）稳定地结合核小体，并且ORC-核小体复合物具有将复制性解旋酶MCM双六聚体加载到相邻的无核小体DNA上的内在能力，无论序列如何。此外，我们发现非洲爪狼核小体可以替代酵母核小体参与ORC。结合全基因组ORC结合数据的重新分析，我们的研究结果引导我们提出，酵母起源识别机制包含在无核小体区域和许可起源附近结合核小体的隐秘能力，并且这种核小体定向起源许可范式适用于所有真核生物。
