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: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=688435
• 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解开人类细胞如何在DNA复制和细胞周期控制的背景下保持遗传稳定性的机制。为了保持遗传稳定，整个基因组必须忠实地复制，并且每个细胞周期只复制一次。后者是通过严格调控复制前复合体(Pre-RC；“许可”)的逐步形成和复制起始的有序激活来实现的。现有数据表明，CDC7在许可和复制启动过程中都发挥着关键作用。当CDC7被Dbf4调节亚基结合时，它本身就被激活。然而，Dbf4与CDC7结合以激活它的确切机制尚不清楚。一种流行的模型是CDC7在高水平的Dbf4的背景下被CDK介导的磷酸化激活。然而，与这个模型相反的是，我们最近发现非磷酸化的CDC7对DNA复制的起始点具有很高的亲和力，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，阻止DNA(Re)复制，从而确保每个细胞周期一次的DNA复制。我们还发现，当细胞退出有丝分裂时，作为复制激活剂的CDC7可以通过PP1介导的去磷酸化得到恢复。PP1还被发现在已经形成Pre-RC之后激活复制检查点。因此，PP1既可以作为复制的促进因子，也可以作为复制的抑制因子：前者通过恢复CDC7的功能，后者通过激活复制检查点(即，抑制复制起始)。因此，人类细胞需要在CDC7、CDK1和PP1之间有密切的串扰，以维持遗传稳定性。基于这些观察，我们推测PP1介导的CDC7去磷酸化是一种积极的调控机制，通过两个不同的步骤促进复制：许可和启动激活步骤。为了验证这一假设，我们建议：(I)系统地研究PP1介导的CDC7去磷酸化在正常、永生化和癌变的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。这项研究的数据将为人类细胞如何在DNA复制和细胞周期控制的背景下保持或失去遗传稳定性的机制提供新的线索。