Ayoub Nabieh, Jeyasekharan Anand D, Venkitaraman Ashok R
The Medical Research Council Cancer Cell Unit, Cambridge, UK.
Cell Cycle. 2009 Sep 15;8(18):2945-50.
The pathways that signal double-strand DNA breaks (DSBs) in mammalian cells are central to the maintenance of genome integrity. We have reported (Ayoub et al., Nature 2008; 453: 682-6) that the rapid mobilization of the heterochromatin protein, HP1beta, within seconds from DSB sites promotes chromatin changes like H2AX phosphorylation that trigger this response. Notably, this paper and a subsequent report (Ayoub et al., Cell Cycle 2009; 8: 1494-500), demonstrate that transient HP1beta mobilization is followed by its accumulation over time at DSB sites. Indeed, two recent papers (Luijsterburg et al., J Cell Biol 2009; 185:577-86 and Zarebski et al., Cytometry A May 2009) suggest that HP1 recruitment to damage sites, rather than its rapid mobilization, is the predominant behaviour exhibited by this protein. Here, we present new experimental analyses which corroborate that fluorophore-tagged HP1beta exhibits two distinct behaviours at DSB sites in living cells - rapid, transient mobilization, most evident in heterochromatic regions, followed by slower recruitment. Experimental methods allowing visualization of these behaviours are described. Interestingly, chemical inhibition of the DNA-damage responsive enzyme, casein kinase 2 (CK2), suppresses HP1beta mobilization while permitting recruitment. Our findings reconcile recent findings in a new model, wherein rapid HP1beta mobilization from DSBs mediated by its phosphorylation on Thr51 by CK2, is followed by, and may overlap with, its accumulation at these sites via the chromoshadow domain, independent of Thr51. Our analyses provide fresh insight into the earliest events that trigger the DNA damage response in mammalian cells.
在哺乳动物细胞中，发出双链DNA断裂（DSB）信号的通路对于维持基因组完整性至关重要。我们曾报道过（阿尤布等人，《自然》，2008年；453卷：682 - 6页），异染色质蛋白HP1β在数秒内从DSB位点快速移动，会促进染色质变化，如H2AX磷酸化，从而触发这种反应。值得注意的是，本文以及随后的一篇报道（阿尤布等人，《细胞周期》，2009年；8卷：1494 - 500页）表明，HP1β短暂移动后会随着时间在DSB位点积累。实际上，最近的两篇论文（路易斯特伯格等人，《细胞生物学杂志》，2009年；185卷：577 - 86页和扎雷布斯基等人，《细胞分析A》，2009年5月）指出，HP1募集到损伤位点，而非其快速移动，是该蛋白表现出的主要行为。在此，我们展示了新的实验分析，证实荧光团标记的HP1β在活细胞的DSB位点呈现出两种不同行为——快速、短暂的移动，在异染色质区域最为明显，随后是较慢的募集。文中描述了能够观察到这些行为的实验方法。有趣的是，对DNA损伤反应酶酪蛋白激酶2（CK2）的化学抑制会抑制HP1β的移动，同时允许其募集。我们的研究结果在一个新模型中协调了近期的研究发现，即由CK2在苏氨酸51位点磷酸化介导的HP1β从DSB的快速移动之后，且可能与其通过染色体阴影结构域在这些位点的积累重叠，这一过程独立于苏氨酸51。我们的分析为触发哺乳动物细胞DNA损伤反应的最早事件提供了新的见解。
