Control of Programmed Replication fork Arrest by Chromosome Kissing

描述（由申请人提供）：据报道，癌细胞中主要的DNA交易，如转录、发育、x染色体失活和染色体易位，是由称为Reb1的myb样终止蛋白介导的染色体间接触（称为“染色体接吻”）控制的。我们最近在《细胞》杂志上发表的一篇论文中表明，裂变酵母中的染色体接吻也通过控制序列特异性、生理程序化的叉停来控制复制叉的运动。这一发现提出了一些主要问题：(1)哪些蛋白质参与控制染色体接吻，它们的作用机制是什么？（ii）程序化的分叉停止机制是什么？在解决第一个问题的背景下，我们发现裂变酵母的两种染色质重塑蛋白调节程序性叉阻滞。这意味着这些重塑子通过调节染色体亲和和核小体在复制末端的配置来做到这一点。这项提议的目标之一是在全基因组搜索中发现调节染色体接吻的蛋白质，并试图了解它们的作用机制。该建议的另一个重要目标是通过测试MCM2-7解旋酶单向阻滞发生的假设来揭示程序化叉阻滞的作用机制。程序叉捕获控制着其他染色体交易，如重组、基因沉默和转录通道，并处于复制和其他过程的间期。最后，提出了定位reb1依赖的全基因组复制末端的实验。这个目标的目的是测试两个假设：(i)自然发生的，非规范弱位点通过染色体亲和和/或DNA环依赖于强规范位点的相互作用而呈现功能，这是这些远距离蛋白质- DNA相互作用的一个功能；（ii）环依赖（或独立）叉阻滞的功能是防止复制和转录之间的干扰。

DESCRIPTION (provided by applicant): Major DNA transactions such as transcription, development, X-chromosome inactivation and chromosomal translocations in cancer cells were reported to be controlled by chromosome to chromosome contacts called "chromosome kissing" mediated by a myb-like terminator protein called Reb1. We have recently shown in a paper to be published in Cell that chromosome kissing in fission yeast also controls replication fork movement by controlling sequence-specific, physiologically programmed fork arrest. This finding raises some major questions: (i) which proteins are involved in controlling chromosome kissing and what are their mechanisms of action?; (ii) what is the mechanism of programmed fork arrest? In the context of addressing the first question, we have discovered that two chromatin remodeling proteins of fission yeast modulate programmed fork arrest. The implication is that these remodelers do this by modulating chromosome kissing and nucleosome disposition about the replication termini. One of the goals of this proposal is to discover in a genome-wide search the proteins that modulate chromosome kissing and try to understand their mechanism of action. Another important goal of this proposal is to uncover the mechanism of action of programmed fork arrest by testing the hypothesis that this happens by unidirectional arrest of the MCM2-7 helicase. Programmed fork arrest controls other chromosome transactions such as recombination, gene silencing and transcriptional passage and is at the interphase of replication and other processes. Finally, experiments are proposed to localize genome-wide Reb1-dependent replication termini. The objectives of this goal is to test two hypotheses: (i) naturally occurring, noncanonical weak sites are rendered functional by chromosome kissing and/or DNA looping -dependent interaction with strong canonical sites and that is one function of these long range protein- DNA interactions; (ii) a function of looping-dependent (or independent) fork arrest is to prevent interference between replication and transcription.