Initiation of DNA Replication of Yeast Chromosomes

描述（由申请人提供）：基因组的及时和准确复制对所有真核细胞的正常增殖至关重要。因此，DNA复制的起始与细胞周期的进展密切协调。这项提议的长期目标是确定选择DNA复制位点和激活起始过程的事件。DNA复制酶的控制与复制前复合物（pre-replicative complex，pre-RC）的形成和激活紧密相关，所述复制前复合物包括起始识别复合物（origin recognition complex，ORC）、Cdc 6和Mcm 2 -7复合物（推定的复制性DNA解旋酶）。使用新的体外试验的组合前RC形成和突变体的ORC，Cdc 6和Mcm 2 -7蛋白贝尔博士将：1。确定前RC的成分和前RC的形成机制。2.确定ATP如何控制前RC的形成。3.确定细胞周期蛋白依赖性激酶和ORC如何共同作用，以防止起源在细胞周期中的不适当时间重新启动。4.开发新的体外试验，用于检测从确定的复制起点开始复制的事件。这些研究将使用酵母S。然而，真核DNA复制的高度保守性表明，酵母中的进展将迅速转化为对人类细胞中相同过程的理解。在S.酿酒酵母使其成为进行这些研究的理想生物。对这些蛋白质的新认识将为抗真菌化合物和识别癌细胞的潜在标记物提供强有力的候选靶点。此外，在这些研究中获得的酵母复制蛋白的理解将指导研究，以确定这些蛋白的人类类似物的抑制剂，这代表了新的化疗化合物的重要目标。

DESCRIPTION (provided by applicant): The timely and accurate replication of the genome is essential to the normal proliferation of all eukaryotic cells. Accordingly the initiation of DNA replication is carefully coordinated with the progress of the cell cycle. The long-term objective of this proposal is to determine the events that select the sites of DNA replication and activate the initiation process. Control of DNA replicaiton is tightly linked to the formation and activation of the pre-replicative complex (pre-RC) that includes the origin recognition complex (ORC), Cdc6 and the Mcm2-7 complex (the putative replicative DNA helicase). Using a combination of new in vitro assays for pre- RC formation and mutants in ORC, Cdc6 and the Mcm2-7 proteins Dr. Bell will: 1. Determine the components of the pre-RC and the mechanism of pre-RC formation. 2. Determine the how ATP controls the formation of the pre-RC. 3. Determine how cyclin dependent kinases and ORC work together to prevent origins from re-initiating at inappropriate times during the cell cycle. 4. Develop new in vitro assays for the events leading to replication initiation from a defined origin of replication. These studies will be performed using the yeast S. cerevisiae, however, the highly conserved nature of eukaryotic DNA replication indicates progress in yeast will rapidly translated to an understanding of the same processes in human cells. The rapid progress that can be made in S. cerevisiae make it an ideal organism to perform these studies. New understanding of these proteins will lead to strong candidate targets for antifungal compounds and potential markers to identify cancer cells. In addition, the understanding of the yeast replication proteins gained in these studies will direct studies to identify inhibitors of the human analogs of these proteins, which represent important targets for novel chemotherapeutic compounds.