Bewersdorf Jörg, Bennett Brian T, Knight Kendall L
Institute for Molecular Biophysics, The Jackson Laboratory, Bar Harbor, ME 04609, USA.
Proc Natl Acad Sci U S A. 2006 Nov 28;103(48):18137-42. doi: 10.1073/pnas.0608709103. Epub 2006 Nov 16.
DNA double-strand breaks (DSBs) caused by cellular exposure to genotoxic agents or produced by inherent metabolic processes initiate a rapid and highly coordinated series of molecular events resulting in DNA damage signaling and repair. Phosphorylation of histone H2AX to form gamma-H2AX is one of the earliest of these events and is important for coordination of signaling and repair activities. An intriguing aspect of H2AX phosphorylation is that gamma-H2AX spreads a limited distance up to 1-2 Mbp from the site of a DNA break in mammalian cells. However, neither the distribution of H2AX throughout the genome nor the mechanism that defines the boundary of gamma-H2AX spreading have yet been described. Here, we report the identification of previously undescribed H2AX chromatin structures by successfully applying 4Pi microscopy to visualize endogenous nuclear proteins. Our observations suggest that H2AX is not distributed randomly throughout bulk chromatin, rather it exists in distinct clusters that themselves are uniformly distributed within the nuclear volume. These data support a model in which the size and distribution of H2AX clusters define the boundaries of gamma-H2AX spreading and also may provide a platform for the immediate and robust response observed after DNA damage.
细胞暴露于基因毒性剂所导致的或由内在代谢过程产生的DNA双链断裂（DSB）会引发一系列快速且高度协调的分子事件，从而导致DNA损伤信号传导和修复。组蛋白H2AX磷酸化形成γ-H2AX是这些事件中最早发生的事件之一，对于信号传导和修复活动的协调至关重要。H2AX磷酸化的一个有趣之处在于，在哺乳动物细胞中，γ-H2AX从DNA断裂位点向上扩散的距离有限，可达1-2兆碱基对。然而，H2AX在整个基因组中的分布以及定义γ-H2AX扩散边界的机制尚未得到描述。在此，我们报告通过成功应用4Pi显微镜观察内源性核蛋白，鉴定出了先前未描述的H2AX染色质结构。我们的观察结果表明，H2AX并非随机分布于整个块状染色质中，而是存在于不同的簇中，这些簇本身在核体积内均匀分布。这些数据支持这样一个模型，即H2AX簇的大小和分布定义了γ-H2AX扩散的边界，并且还可能为DNA损伤后观察到的即时且强烈的反应提供一个平台。
