Rec114 ChIP-seq

DNA duplication is intimately connected to setting up post-replicative chromosome structures and events, but molecular details of this coordination are not well understood. A striking example occurs during yeast meiosis, where replication locally influences timing of the DNA double-strand breaks (DSBs) that initiate recombination. We show here that replication-DSB coordination is eliminated by overexpressing Dbf4-dependent Cdc7 kinase (DDK) or removing Tof1 or Csm3, components of the replication fork protection complex (FPC). DDK physically associates with Tof1, and Tof1 is dispensable for replication-DSB coordination if DDK is artificially tethered to replisomes. Furthermore, DDK phosphorylation of the DSB-promoting factor Mer2 is locally coordinated with replication, dependent on Tof1. These findings indicate that DDK recruited by FPC to replisomes phosphorylates chromatin-bound Mer2 in the wake of the replication fork, thus synchronizing replication with an early prerequisite for DSB formation. This may be a general mechanism to ensure spatial and temporal coordination of replication with other chromosomal processes. Ninety-six samples total: 12 time points (each time points contains ChIP and input samples) from Rec114-myc ARS+, Rec114-myc ars∆ strains, Rec114-myc tof1∆ARS+ and Rec114-myc tof1∆ ars∆ strains

DNA复制与复制后染色体结构及相关事件的建立紧密关联，但二者协调作用的分子细节尚未被充分解析。一个极具代表性的例证出现在酵母减数分裂过程中，此时DNA复制会局部调控启动同源重组的DNA双链断裂（double-strand breaks, DSBs）的发生时序。本研究发现，过表达依赖Dbf4的Cdc7激酶（Dbf4-dependent Cdc7 kinase, DDK），或缺失复制叉保护复合物（replication fork protection complex, FPC）的组分Tof1或Csm3，均可消除复制与DSB之间的协调调控。DDK可与Tof1发生物理结合；若将DDK人工锚定至复制体，则Tof1对于复制-DSB的协调调控不再是必需的。进一步研究显示，在依赖Tof1的前提下，DDK对促进DSB形成的因子Mer2的磷酸化过程会与DNA复制发生局部协同。上述结果表明，由FPC招募至复制体的DDK，会在复制叉后方对结合于染色质的Mer2进行磷酸化修饰，从而使DNA复制与DSB形成的早期先决条件实现同步。这一机制可能是一种通用策略，用以确保DNA复制与其他染色体生物学过程在时空维度上的协调统一。本次实验共计96个样本：涵盖Rec114-myc ARS+、Rec114-myc ars∆、Rec114-myc tof1∆ARS+以及Rec114-myc tof1∆ ars∆四种菌株的12个时间点样本，每个时间点均包含染色质免疫沉淀（ChIP）样本与输入对照样本。