Department of Life Science, Division of Life and Pharmaceutical Sciences, Ewha Womans University, Seoul 120-750, Korea.
Mol Cells. 2012 Jul;34(1):85-91. doi: 10.1007/s10059-012-0112-4. Epub 2012 Jun 12.
The formation of γ-H2AX foci after DNA double strand breaks (DSBs) is crucial for the cellular response to this lethal DNA damage. We previously have shown that BRG1, a chromatin remodeling enzyme, facilitates DSB repair by stimulating γ-H2AX formation, and this function of BRG1 requires the binding of BRGI to acetylated histone H3 on γ-H2AX-containing nucleosomes using its bromodomain (BRD), a protein module that specifically recognizes acetyl-Lys moieties. We also have shown that the BRD of BRG1, when ectopically expressed in cells, functions as a dominant negative inhibitor of the BRG1 activity to stimulate γ-H2AX and DSB repair. Here, we found that BRDs from a select group of proteins have no such activity, suggesting that the γ-H2AX inhibition activity of BRG1 BRD is specific. This finding led us to search for more BRDs that exhibit γ-H2AX inhibition activity in the hope of finding additional BRD-containing proteins involved in DNA damage responses. We screened a total of 52 individual BRDs present in 38 human BRD-containing proteins, comprising 93% of all human BRDs. We identified the BRD of cat eye syndrome chromosome region candidate 2 (Cecr2), which recently was shown to form a novel chromatin remodeling complex with unknown cellular functions, as having a strong γ-H2AX inhibition activity. This activity of Cecr2 BRD is specific because it depends on the chromatin binding affinity of Cecr2 BRD. Small interfering RNA knockdown experiments showed that Cecr2 is important for γ-H2AX formation and DSB repair. Therefore, our genomewide screen identifies Cecr2 as a novel DNA damage response protein.
γ-H2AX 焦点的形成是细胞对这种致命 DNA 损伤做出反应的关键。我们之前已经表明，染色质重塑酶 BRG1 通过刺激 γ-H2AX 的形成来促进 DSB 修复，BRG1 的这种功能需要 BRGI 利用其溴结构域（BRD）结合到含有 γ-H2AX 的核小体上的乙酰化组蛋白 H3 上，BRD 是一种特异性识别乙酰化 Lys 部分的蛋白模块。我们还表明，BRG1 的 BRD 在细胞中外源表达时，可作为 BRG1 活性的显性负抑制剂发挥作用，从而刺激 γ-H2AX 和 DSB 修复。在这里，我们发现一组特定蛋白质的 BRD 没有这种活性，这表明 BRG1 BRD 的 γ-H2AX 抑制活性是特异性的。这一发现促使我们寻找更多具有 γ-H2AX 抑制活性的 BRD，希望能找到更多参与 DNA 损伤反应的含有 BRD 的蛋白质。我们总共筛选了 38 个人类含有 BRD 的蛋白质中存在的 52 个单独的 BRD，占所有人类 BRD 的 93%。我们鉴定出猫眼综合征染色体区域候选 2（Cecr2）的 BRD，它最近被证明与一个具有未知细胞功能的新型染色质重塑复合物形成，具有很强的 γ-H2AX 抑制活性。Cecr2 BRD 的这种活性是特异性的，因为它依赖于 Cecr2 BRD 与染色质的结合亲和力。小干扰 RNA 敲低实验表明 Cecr2 对 γ-H2AX 的形成和 DSB 修复很重要。因此，我们的全基因组筛选确定 Cecr2 为一种新型的 DNA 损伤反应蛋白。
