Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, CH-4058 Basel, Switzerland.
Nat Cell Biol. 2012 Apr 8;14(5):502-9. doi: 10.1038/ncb2465.
Chromatin mobility is thought to facilitate homology search during homologous recombination and to shift damage either towards or away from specialized repair compartments. However, unconstrained mobility of double-strand breaks could also promote deleterious chromosomal translocations. Here we use live time-lapse fluorescence microscopy to track the mobility of damaged DNA in budding yeast. We found that a Rad52-YFP focus formed at an irreparable double-strand break moves in a larger subnuclear volume than the undamaged locus. In contrast, Rad52-YFP bound at damage arising from a protein-DNA adduct shows no increase in movement. Mutant analysis shows that enhanced double-strand-break mobility requires Rad51, the ATPase activity of Rad54, the ATR homologue Mec1 and the DNA-damage-response mediator Rad9. Consistent with a role for movement in the homology-search step of homologous recombination, we show that recombination intermediates take longer to form in cells lacking Rad9.
染色质的流动性被认为有助于同源重组过程中的同源搜索，并将损伤转移到专门的修复隔室中或远离这些隔室。然而，双链断裂的不受限制的流动性也可能促进有害的染色体易位。在这里，我们使用活细胞延时荧光显微镜来跟踪芽殖酵母中受损 DNA 的流动性。我们发现，在不可修复的双链断裂处形成的 Rad52-YFP 焦点在核内的移动范围比未受损的基因座大。相比之下，在由蛋白质-DNA 加合物引起的损伤处结合的 Rad52-YFP 没有显示出运动的增加。突变分析表明，增强的双链断裂流动性需要 Rad51、Rad54 的 ATP 酶活性、ATR 同源物 Mec1 和 DNA 损伤反应介质 Rad9。与运动在同源重组的同源搜索步骤中的作用一致，我们表明，缺乏 Rad9 的细胞中重组中间体的形成需要更长的时间。
