Cell cycle regulation by PP1 and Cdc7

PP1 和 Cdc7 的细胞周期调节

• 批准号: RGPIN-2018-04577
• 负责人: Lee, Hoyun
• 金额: $ 3.06万
• 依托单位: Laurentian University of Sudbury
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=713101
• 关键词: Cell; cycle; regulation; PP1; Cdc7

My long-term research goal is unravelling the mechanism of how human cells maintain genetic stability in the context of DNA replication and cell cycle control. To maintain genetic stability, the entire genome must replicate faithfully and only once per cell cycle. The latter is achieved by strictly regulating the stepwise formation of the pre-replication complex (pre-RC; “licensing”) and the orderly activation of replication initiation. Available data indicate that Cdc7 plays critical roles for both licensing and replication initiation processes. Cdc7 itself is activated when it is bound by the Dbf4 regulatory subunit. However, the exact mechanism when Dbf4 binds to Cdc7 to activate it is poorly understood. A prevalent model has been that Cdc7 is activated by Cdk-mediated phosphorylation in the context of high levels of Dbf4. Contrarily to this model, however, we have recently found that non-phosphorylated Cdc7 has high affinity for the origin of DNA replication and activates DNA replication. Furthermore, Cdk1/cyclin B-mediated Cdc7 phosphorylation leads to its dissociation from chromatin, preventing DNA (re)replication and, thus, ensuring once-per-cell cycle DNA replication. We also found that Cdc7 as the replication activator is “restored” by PP1-mediated dephosphorylation as cells exit mitosis. PP1 has also been found to activate a replication checkpoint after pre-RC is already formed. Thus, PP1 may function as a replication promoter as well as an inhibitor: for the former by restoring Cdc7 function and the latter by activating replication checkpoint (i.e., inhibiting replication initiation). Thus, human cells need to have intimate crosstalk between Cdc7, Cdk1 and PP1 to maintain genetic stability. Based on these observations, we postulate that the PP1-mediated Cdc7 dephosphorylation is an active regulation mechanism to promote replication at two different steps: the licensing and initiation activation steps. To test this hypothesis, we propose to: (i) systematically examine the role of PP1-mediated Cdc7 dephosphorylation in the replication licensing and activation process in normal, immortalized, and cancerous human cells; and (ii) determine the potential role of Cdc7 dephosphorylation in the regulation of cytokinesis. Data from this study will shed new light on the mechanism of how human cells maintain or lose genetic stability in the context of DNA replication and cell cycle control.

我的长期研究目标是揭示人类细胞如何在DNA复制和细胞周期控制的背景下维持遗传稳定性的机制，整个基因组必须忠实地复制，并且每个细胞周期只能实现一次。通过严格调控复制前复合体（pre-RC；“许可”）的逐步形成和复制启动的有序激活，现有数据表明，Cdc7 在许可和复制启动过程中发挥着关键作用。这是然而，人们对 Dbf4 与 Cdc7 结合并激活它的确切机制知之甚少。与此相反，Cdc7 是由 Cdk 介导的磷酸化激活的。然而，我们最近发现非磷酸化的Cdc7对DNA复制起点具有高亲和力并激活DNA复制。此外，Cdk1/cyclin。 B 介导的 Cdc7 磷酸化导致其与染色质解离，从而阻止 DNA 复制，从而确保每个细胞周期 DNA 复制一次。我们还发现，Cdc7 作为复制激活剂可通过 PP1 介导的去磷酸化“恢复”。当细胞退出有丝分裂时，PP1 也被发现在前 RC 形成后激活复制检查点，因此，PP1 可能充当复制启动子。抑制剂：前者通过恢复 Cdc7 功能，后者通过激活复制检查点（即抑制复制起始）。因此，人类细胞需要 Cdc7、Cdk1 和 PP1 之间的密切串扰来维持遗传稳定性。假设 PP1 介导的 Cdc7 去磷酸化是一种主动调节机制，可在两个不同的步骤中促进复制：许可步骤和起始激活步骤。为了检验这一假设，我们建议： (i) 系统地研究 PP1 介导的 Cdc7 去磷酸化在正常、永生化和癌性人类细胞的复制许可和激活过程中的作用；以及 (ii) 确定 Cdc7 去磷酸化在胞质分裂调节中的潜在作用。这项研究将为人类细胞在 DNA 复制和细胞周期控制的背景下如何维持或失去遗传稳定性的机制提供新的线索。