DNA REPLICATION-LINKED CHROMATIN ASSEMBLY IN YEAST

描述：这是一个研究染色质组装因子的建议 在酵母中，使用互补的遗传和生物化学方法。 的PI 将集中在酵母蛋白复合物称为染色质组装因子-I （CAF-I），优先将组蛋白H3和H4组装到复制的DNA上 体外 含有编码CAF-I的基因的无效等位基因的细胞 亚基是有活力的，但显示出对羟基脲的敏感性改变， 苯菌灵，破坏细胞周期的药物。 私家侦探会测试这些变种人 染色质结构、基因表达和细胞周期的缺陷 检查点控制 确定冗余装配活动是否考虑 对于缺乏CAF-I的细胞的存活，PI将筛选突变 这产生了对完整CAF-I和高拷贝抑制因子的需求 CAF-I突变体的药物敏感性表型。 表位标记酵母 CAF-I亚基将用于分离体内结合CAF-I的蛋白质。 通过这些方法鉴定的蛋白质将在生化试验中进行检测。

DESCRIPTION: This is a proposal to investigate chromatin assembly factors in yeast, using complementary genetic and biochemical approaches. The PI will focus on a yeast protein complex termed Chromatin Assembly Factor-I (CAF-I) that assembles histones H3 and H4 preferentially onto replicated DNA in vitro. Cells containing null alleles of the genes encoding CAF-I subunits are viable but display altered sensitivity to hydroxyurea and benomyl, drugs that disrupt the cell cycle. The PI will test these mutants for defects in chromatin structure, gene expression, and cell cycle checkpoint control. To determine if redundant assembly activities account for the survival of cells lacking CAF-I, the PI will screen for mutations that generate a requirement for intact CAF-I and for high copy suppressors of the drug sensitivity phenotypes of CAF-I mutants. Epitope-tagged yeast CAF-I subunits will be used to isolate proteins bound to CAF-I in vivo. Proteins identified by these methods will be tested in biochemical assays.