Biology of Topoisomerase 1 in S Cerevisiae

描述（由申请人提供）：本提案的目标是了解 一种新的DNA聚合酶Pol K如何发挥作用以偶联复制叉通道 新形成的姐妹染色单体之间的凝聚力的建立。我们有 最近确定，po 1 K（由TRF 4和TRF 5编码）对这两种细胞都是必不可少的。 复制和内聚（Wang等人（2000）Science 289，774-779）。这些 观察打开了大门，进一步机械洞察力如何凝聚力 建立完成，它是如何耦合到复制叉通道 也许还与姐妹染色单体之间的结合有关。机制 在复制过程中形成内聚的方式对于维持 但仍然是一个基本的未解之谜 在细胞生物学中。我们假设POI K是 所有真核生物中的复制叉，其功能是复制凝聚力 位点特异性地结合，从而在染色单体之间建立起结合状态。 我们进一步假设po 1 K和非聚合酶之间的相互作用 pol E的C-末端结构域大约发生，并且这种相互作用是必要的 用于pol K作为S阶段检查点传感器。实验旨在测试 我们的假设概述如下。

DESCRIPTION (provided by applicant): The goal of this proposal is to understand how a novel DNA polymerase, Pol K, functions to couple replication fork passage to establishment of cohesion between newly formed sister chromatids. We have recently determined that po1 K (encoded by TRF4 and TRF5) is essential for both replication and cohesion (Wang et al. (2000) Science 289, 774-779). These observations open the door to further mechanistic insight into how cohesion establishment is accomplished, how it is coupled to replication fork passage and perhaps to the nature of the bond between sister chromatids. The mechanism by which cohesion is formed during replication is crucial for the maintenance of genomic integrity, but nonetheless remains a fundamental unanswered question in cell biology. We hypothesize that POI K is an essential component of the replication fork in all eukaryotes whose function is to replicate cohesion sites specifically and, thereby, set up the cohesed state between chromatids. We further hypothesize that an interaction between po1 K and the non-polymerase C-terminal domain of pol E about occurs and that this interactions is necessary forpol K to act as an S-phase checkpoint sensor. Experiments designed to test our hypothesis are outlined below.