Cis-and trans-acting determinants of replication timing

描述(由申请人提供)： 癌症发生所需的细胞变化之一是“基因组”。 不稳定“，这个术语描述的是一系列通向 基因突变和基因表达模式改变的频率增加。 导致基因组不稳定的机制包括染色体断裂、基因 扩增和基因表达水平的变化。后者是 经常与基因甲基化状态的变化有关。所有这些都是 机制一直与复制时间的变化相关，而且它是 在某些情况下，复制时间的更改可能是主要原因 造成这些现象的原因。到目前为止，很难评估 癌症发展过程中复制时间的变化，因为几乎没有 了解控制复制计时的机制。幸运的是 芽殖酵母和裂殖酵母(Saccharmyces cerevisiae和 在我们的实验室和其他实验室中，裂殖酵母(Schizosaccharmyces Pombe)提供了入口 了解复制时间。在这两种酵母菌中我们都鉴定出 包含内部顺式作用序列的复制起始点 复制时间延迟。在这里，我们提出一些简单的策略，使我们能够 为了精确定位这些序列并使用它们来提取 相应的反式作用蛋白。此外，在其他方面的研究 实验室和我们的实验室已经确定了几种可能发挥作用的蛋白质 在复制计时中的角色。我们打算调查一下它的潜在作用 通过研究这些蛋白质在复制计时中的作用 相应基因的突变，并通过研究二进制 这些蛋白质之间的相互作用。建议的实验结果 应该允许我们开发控制机制的详细模型 芽酵母和分裂酵母的复制时间。保守的方面 这些模型很可能被证明适用于其他真核细胞， 包括人类细胞。这一信息可能会使药物的开发 这将增强正常复制定时稳定性，从而抑制或 逆转癌症的发展。

DESCRIPTION (provided by applicant): One of the cellular alterations required for development of cancer is "genomic instability," a term that describes a collection of pathways all leading to increased frequency of gene mutation and altered patterns of gene expression. Among the mechanisms causing genomic instability are chromosome breakage, gene amplification, and changes in levels of gene expression. The latter are frequently associated with changes in gene methylation status. All of these mechanisms have been correlated with changes in replication timing, and it is possible that in some cases changes in replication timing may be the primary cause of these phenomena. So far it has been difficult to evaluate the role of changes in replication timing in cancer development, because so little is known about the mechanisms controlling replication timing. Fortunately previous studies in budding and fission yeasts (Saccharomyces cerevisiae and Schizosaccharomyces pombe), in our laboratory and others, provide entry points into understanding replication timing. In both yeasts we have identified replication origins containing internal cis-acting sequences that determine late replication timing. Here we propose simple strategies that will permit us to precisely localize those sequences and use them to pull out the corresponding trans-acting proteins. In addition, studies in other laboratories and ours have already identified several proteins likely to play roles in replication timing. We intend to investigate the potential roles of all of these proteins in replication timing by studying the effects on timing of mutations in the corresponding genes and by studying the binary interactions between these proteins. The results of the proposed experiments should permit us to develop detailed models of the mechanisms controlling replication timing in both budding and fission yeasts. The conserved aspects of these models are likely to prove applicable to other eukaryotic cells, including human cells. This information may permit the development of drugs that will enhance normal replication timing stability and thus inhibit or reverse the development of cancer.