A novel function of Orc6 during mitosis in S. cerevisiae

描述（由申请人提供）：真核细胞在s期复制其染色体，并在有丝分裂期间将染色体分离成两个子细胞。染色体分离错误会导致染色体的获得或丢失。这是染色体不稳定的一个原因，而染色体不稳定是肿瘤发生的一个标志。我们的长期目标是了解酿酒酵母染色体精确分离的分子机制。Orc6p控制着从酵母到人类的DNA复制。最近有人提出，Orc蛋白除了在DNA复制中具有众所周知的功能外，还可能具有有丝分裂功能。人类Orc6p定位于着丝点，与染色体分离的控制有关。然而，Orc6p如何控制有丝分裂尚不清楚。我推测Orc6p在s.s cerevisiae中具有有丝分裂功能。目的：研究酿酒酵母Orc6p的有丝分裂功能；(1) orc6的条件突变体将被分离并鉴定。(2)分析蛋白定位，观察有丝分裂过程中Orc6p是否定位于着丝点。(3)利用orc6条件突变体进行抑制基因筛选，揭示orc6与有丝分裂基因之间的具体遗传相互作用。成功分离仅在有丝分裂过程中存在缺陷的orc6突变体，将有助于研究Orc6p的有丝分裂功能。Orc6p的着丝点定位将确保Orc6p在染色体分离中发挥作用。抑制基因筛选将有助于我们了解Orc6功能的分子机制。Orc6p从酵母到人类都是保守的。本研究结果可用于了解Orc6p在高等真核生物中的分子功能。准确的染色体分离是有丝分裂过程中避免染色体不稳定的关键步骤。研究真核生物染色体分离的机制对了解肿瘤发生的分子机制具有重要意义。

DESCRIPTION (provided by applicant): Eukaryotic cells duplicate their chromosomes during S-phase, and segregate the chromosomes into two daughter cells during mitosis. Errors in the chromosome segregation could lead to gain or loss of chromosome. This is a cause of chromosome instability which is a hallmark of tumorigenesis. Our long-term goal is to understand the molecular mechanism of accurate chromosome segregation in S. cerevisiae. Orc6p controls DNA replication from yeast to humans. Recently it has been proposed that Orc proteins might also have mitotic functions besides its well characterized function in DNA replication. Human Orc6p is localized to kinetochore, and has been implicated in the control of chromosome segregation. However it is not clear how Orc6p controls mitosis. I hypothesize that Orc6p has a mitotic function in S. cerevisiae. To study the mitotic function of Orc6p in S. cerevisiae; (1) a conditional mutant of orc6 will be isolated and characterized. (2) Protein localization will be analyzed to see if Orc6p is localized kinetochore during mitosis. (3) A suppressor gene screening using the orc6 conditional mutant will be conducted to reveal the specific genetic interactions between ORC6 and mitotic genes. Successful isolation of orc6 mutant which is defective only during mitosis will help us to study the mitotic function of Orc6p. Kinetochore localization of Orc6p will ensure that Orc6p has a role in chromosome segregation. Suppressor gene screening will help us to understand the molecular mechanism of Orc6 function. Orc6p is conserved from yeast to humans. The results obtained from this study can be applied to understand the molecular function of Orc6p in higher eukaryotes. The accurate chromosome segregation is a critical step during mitosis to avoid chromosome instability. It is important to study the mechanism of chromosome segregation in eukaryotes to understand the molecular mechanism of tumorigenesis.