Synthesis of ADP-Ribosylated Histones Reveals Site-Specific Impacts on Chromatin Structure and Function
Synthesis of ADP-Ribosylated Histones Reveals Site-Specific Impacts on Chromatin Structure and Function
复制标题
DOI：
10.1021/jacs.1c05429
复制
发表时间：
2021-07-15
影响因子：
15
通讯作者：
Muir, Tom W.
中科院分区：
文献类型：
--
作者：
Hananya, Nir;Daley, Sara K.;Muir, Tom W.
核蛋白的ADP-核糖基化是各种DNA损伤修复途径的关键特征。组蛋白，特别是H3和H2 B，是ADP-核糖基化的主要靶标，并且在DNA损伤后主要在丝氨酸上用单个ADP-核糖单元修饰。虽然PARP 1依赖的多聚ADP核糖基化的整体影响进行了大量研究，但对组蛋白ADP核糖基化的具体作用知之甚少。在这里，我们报告了一个有效的和模块化的半合成路线全长ADP-核糖基化组蛋白H3和H2 B，化学安装在特定的丝氨酸残基的发展。修饰的组蛋白用于产生各种化学定义的ADP-核糖基化染色质底物，其用于生物物理测定。这些研究揭示了组蛋白H2 B的丝氨酸-6（H2 BS 6ADPr）的ADP-核糖基化抑制染色质折叠和高级组织;值得注意的是，H3 S10的ADP-核糖基化增强了这种效果。此外，ADP-核糖基化的核小体被用于采用一组赖氨酸甲基转移酶的生物化学实验中，揭示了组蛋白H3 K9甲基化的上下文依赖性抑制。设计师ADP-核糖基化染色质的可用性在这里描述，预计将促进进一步的生化和结构研究组蛋白ADP-核糖基化的DNA损伤反应中的作用。
ADP-ribosylation of nuclear proteins is a critical feature of various DNA damage repair pathways. Histones, particularly H3 and H2B, are major targets of ADP-ribosylation and are primarily modified on serine with a single ADP-ribose unit following DNA damage. While the overall impact of PARP1-dependent poly-ADP-ribosylation is heavily investigated, very little is known about the specific roles of histone ADP-ribosylation. Here, we report the development of an efficient and modular semisynthetic route to full-length ADP-ribosylated histones H3 and H2B, chemically installed at specific serine residues. The modified histones were used to generate various chemically defined ADP-ribosylated chromatin substrates, which were employed in biophysical assays. These studies revealed that ADP-ribosylation of serine-6 of histone H2B (H2BS6ADPr) inhibits chromatin folding and higher-order organization; notably, this effect was enhanced by ADP-ribosylation of H3S10. In addition, ADP-ribosylated nucleosomes were utilized in biochemical experiments employing a panel of lysine methyltransferase enzymes, revealing a context-dependent inhibition of histone H3K9 methylation. The availability of designer ADP-ribosylated chromatin described here is expected to facilitate further biochemical and structural studies regarding the roles of histone ADP-ribosylation in the DNA damage response.