Regulation of MCM helicase loading and activation in the model system Saccharomyc

在每个细胞周期的s期，确保基因组的完整和忠实复制是细胞分裂的关键。了解染色体DNA复制的过程，以及协调DNA复制与细胞周期的机制，将为诊断、预后和治疗水平的广泛临床研究工作提供基础。本提案的长期目标是利用酿酒酵母模型系统了解MCM解旋酶功能对真核生物染色体DNA复制的调节。本应用程序提出以下具体目标，以实现长期目标。

Ensuring the complete and faithful duplication of the genome during S-phase of each cell cycle is key for cellular division. Understanding the process of chromosomal DNA replication, and the mechanisms that coordinate DNA replication with the cell cycle, will provide the basis for a wide range of clinical research efforts at the diagnostic, prognostic, and therapeutic levels. The long-term objective of this proposal is to understand the regulation of MCM helicase function for eukaryotic chromosomal DNA replication using the model system Saccharomyces cerevisiae. This application proposes the following specific aims to achieve the long-term objective. Specific Aim 1: To determine the importance of MCM double hexamer complex formation for in vivo DNA replication. Using both molecular and genetic techniques, the importance of MCM double hexamer complex formation for DNA replication will be tested. The results of this aim will provide important information on the mechanical mechanism of MCM helicase function, which is a primary target of cell cycle regulation controlling eukaryotic DNA replication. Furthermore, this aim will be able to directly address conflicting data and differentiate between multiple proposed models for helicase function. Specific Aim 2: To determine if the primary functional role of DDK activity for DNA replication is to load Cdc45 onto origin chromatin. A mutant of MCM5, mcm5-bob1 (P83L), results in constitutive Cdc45 chromatin loading in early G1 and bypass of required Dbf4 dependent kinase (DDK) for G1-S transition and initiation of DNA replication. This aim will determine if Cdc45 chromatin loading is the final downstream culmination of DDK activity and sufficient to achieve DDK bypass. An enhanced understanding of DNA replication control will provide additional insight into disease states where the failure of DNA replication control mechanisms leads to chromosomal instability, mutations, and aneuploidy, all of which are hallmarks of human disease including birth defects, premature ageing, and cancer.