Topoisomerase II- and condensin-dependent breakage of MEC1ATR-sensitive fragile sites occurs independently of spindle tension, anaphase, or cytokinesis.
Topoisomerase II- and condensin-dependent breakage of MEC1ATR-sensitive fragile sites occurs independently of spindle tension, anaphase, or cytokinesis.
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DOI：
10.1371/journal.pgen.1002978
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发表时间：
2012
期刊：
影响因子：
4.5
通讯作者：
Cha RS
中科院分区：
文献类型：
--
作者：
Hashash N;Johnson AL;Cha RS
脆性位点是基因组中经常发生染色体断裂的位点。它们存在于从细菌到人类的生物体中，并与基因组不稳定性，进化和癌症有关。在芽殖酵母中，Mec 1（哺乳动物ATR的同源物）的失活导致染色体在称为复制慢区（RSZ）的脆性位点断裂。RSZ被认为与哺乳动物常见脆性位点（CFSs）同源，其稳定性受ATR调节。在S期的扰动，导致停滞的复制叉的水平升高，是必要的，但不足以在RSZ或CFS染色体断裂。为了解决断裂形成所需的额外事件的性质，我们检查了目前已知或涉及的内源性染色体断裂机制的参与，包括复制叉重新启动错误，过早有丝分裂染色体凝聚，纺锤体张力，后期和胞质分裂。结果显示，在RSZ的染色体断裂是独立的RAD 52上位组基因和TOP 3，SGS 1，SRS 2，MMS 4，或MUS 81，表明同源重组和其他重组相关的过程与复制叉重新启动不太可能参与。我们还发现纺锤体力，后期，或胞质分裂是不稳定的。然而，RSZ断裂需要编码缩合素亚基（YCG 1，YSC 4）和拓扑异构酶II（TOP 2）的基因。我们建议，染色体断裂形成RSZs以下Mec 1失活，哺乳动物脆性位点断裂的模型，是由有丝分裂染色体凝聚过程中产生的内部染色体应力介导的。染色体断裂可发生在正常的细胞分裂过程中。当它发生时，断裂并不是在整个基因组中随机出现的，而是在被称为脆性位点的优选位置。染色体脆性位点的断裂是一种进化上保守的现象，与进化和物种形成有关。在人类中，脆性部位不稳定性也与精神发育迟滞和癌症有关。尽管其具有生物学和临床相关性，但在哺乳动物脆性部位引入断裂的机制仍未得到解决。虽然已经提出了几种合理的模型，但主要由于缺乏合适的实验系统，尚不可能确定它们的贡献。在这里，我们研究了一个酵母模型系统，密切重演的现象，染色体断裂在哺乳动物的脆性位点。我们排除了所有，但目前认为的模型之一，不完全复制的基因组在细胞分裂过程中分离的准备过早压缩。我们还发现，断裂需要参与基因组压缩的三种蛋白质的功能，这是一个从细菌到人类进化保守的重要过程。我们的研究结果表明，正常细胞分裂所需的基本染色体过程可以矛盾地导致基因组不稳定和/或细胞死亡，通过触发染色体断裂在脆弱的网站。
Fragile sites are loci of recurrent chromosome breakage in the genome. They are found in organisms ranging from bacteria to humans and are implicated in genome instability, evolution, and cancer. In budding yeast, inactivation of Mec1, a homolog of mammalian ATR, leads to chromosome breakage at fragile sites referred to as replication slow zones (RSZs). RSZs are proposed to be homologous to mammalian common fragile sites (CFSs) whose stability is regulated by ATR. Perturbation during S phase, leading to elevated levels of stalled replication forks, is necessary but not sufficient for chromosome breakage at RSZs or CFSs. To address the nature of additional event(s) required for the break formation, we examined involvement of the currently known or implicated mechanisms of endogenous chromosome breakage, including errors in replication fork restart, premature mitotic chromosome condensation, spindle tension, anaphase, and cytokinesis. Results revealed that chromosome breakage at RSZs is independent of the RAD52 epistasis group genes and of TOP3, SGS1, SRS2, MMS4, or MUS81, indicating that homologous recombination and other recombination-related processes associated with replication fork restart are unlikely to be involved. We also found spindle force, anaphase, or cytokinesis to be dispensable. RSZ breakage, however, required genes encoding condensin subunits (YCG1, YSC4) and topoisomerase II (TOP2). We propose that chromosome break formation at RSZs following Mec1 inactivation, a model for mammalian fragile site breakage, is mediated by internal chromosomal stress generated during mitotic chromosome condensation. Chromosome breakage can occur during normal cell division. When it occurs, the breaks do not arise randomly throughout the genome, but at preferred locations referred to as fragile sites. Chromosome breakage at fragile sites is an evolutionarily conserved phenomenon, implicated in evolution and speciation. In humans, fragile site instability is also implicated in mental retardation and cancer. Despite its biological and clinical relevance, the mechanism(s) by which breaks are introduced at mammalian fragile sites remains unresolved. Although several plausible models have been proposed, it has not been possible to ascertain their contribution, largely due to the lack of a suitable experimental system. Here, we study a yeast model system that closely recapitulates the phenomenon of chromosome breakage at mammalian fragile sites. We eliminate all but one of the currently considered models—premature compaction of the incompletely replicated genome in preparation for their segregation during cell division. We also find that the breakage required functions of three proteins involved in the genome compaction, an essential process that is evolutionarily conserved from bacteria to humans. Our findings suggest that a fundamental chromosomal process required for normal cell division can paradoxically cause genome instability and/or cell death, by triggering chromosome breakage at fragile sites.
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