Basis for genotoxic stress relief by ATP-dependent chromatin remodelers

ATP依赖性染色质重塑剂缓解基因毒性应激的基础

• 批准号: 9115652
• 负责人: HUA-YING FAN
• 金额: $ 31.18万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9115652
• 关键词: ATP Hydrolysis; Amino Acids; Apoptosis; Binding; Binding Sites; Biochemical; Biological Assay; CCCTC-binding factor; Cell Culture Techniques; Cells; Chromatin; Chromatin Structure; Chromatin Structure Alteration; Cockayne Syndrome; Collection; Coupled; DNA; DNA Repair; DNA lesion; Defect; Development; Disease; Enzymes; Epigenetic Process; Foundations; Gene Expression; Genetic Transcription; Genomic Segment; Genotoxic Stress; Goals; Health; Higher Order Chromatin Structure; Histones; Human; ISWI; In Vitro; Kinetics; Lead; Learning; Lesion; Light; Maintenance; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Modification; Molecular Chaperones; Molecular Conformation; Monitor; Mutation; Neurologic; Nuclear; Nucleosomes; Oxidants; Oxidative Stress; Play; Positioning Attribute; Premature aging syndrome; Process; Property; Public Health; Regulation; Site; Southern Blotting; Stable Isotope Labeling; Syndrome; Time; Ultraviolet Rays; Work; base; chromatin remodeling; developmental disease; genome integrity; insight; reconstitution; repaired; research study; response; ultraviolet irradiation

 DESCRIPTION (provided by applicant): The overall goals of this proposal are to understand how the ATP-dependent chromatin remodeler CSB is distinct from other chromatin remodelers and how its unique features enable this enzyme to protect cells from genotoxic stress. ATP-dependent chromatin remodelers use energy from ATP hydrolysis to alter DNA- histone contacts, leading to changes in nucleosome position, composition or conformation. Thus, they have critical functions in regulating fundamental processes, such as transcription and DNA repair. The importance of these enzymes to health is exemplified by the variety of diseases that result from defects in their activities, which include developmental syndromes and cancer. Mutations in the CSB remodeler lead to Cockayne syndrome: a devastating premature aging disorder associated with numerous developmental and neurological defects. There are more than 30 known and predicted ATP-dependent chromatin remodelers in humans; CSB, however, is the only remodeler that is essential for transcription-coupled DNA repair (TCR), a process that rapidly removes transcription-stalling DNA lesions. Moreover, CSB is critical for relieving oxidative stress. We have made significant advances in understanding how this chromatin remodeler is uniquely equipped to carry out its functions. (1) The association of CSB with chromatin is tightly regulated. In general, only 10% of CSB is chromatin bound, but upon UV irradiation or oxidative stress, more than 90% of CSB becomes chromatin associated; this is in stark contrast to other, well-studied remodelers such as SWI/SNF and ISWI that are constitutively bound to chromatin. (2) Using separation-of-function derivatives, we have demonstrated that the chromatin remodeling activity of CSB is essential for its function in transcription-coupled DNA repair (TCR). (3) The chromatin remodeling activity of CSB is tunable: CSB, on its own, displays weak remodeling activity (~20-fold less than SWI/SNF or ACF). Strikingly, together with NAP1-like histone chaperones, CSB remodels chromatin robustly, to a level similar to SWI/SNF or ACF. (4) CSB interacts with the CTCF protein and through this interaction becomes significantly enriched at CTCF binding sites upon oxidative stress. Given that CTCF is a key player in regulating higher- order chromatin-structure, CSB may collaborate with CTCF to reorganize chromatin structure in response to oxidative stress to facilitate coordinated gene expression or efficient DNA repair. The experiments outlined in this proposal will exploit these unique properties of CSB to understand how this ATP-dependent chromatin remodeler is uniquely equipped to protect cells from genotoxic stress. Moreover, this work may shed new light on the causes and mechanism of Cockayne syndrome.

 描述（由适用提供）：该提案的总体目标是了解ATP依赖性染色质重塑剂CSB如何与其他染色质重塑剂不同，以及其独特特征如何使该酶保护细胞免受遗传毒性应激。依赖ATP的染色质重塑剂使用ATP水解的能量来改变DNA - 荷酮接触，从而导致核小体位置，组成或构象的变化。这，它们在控制基本过程（例如转录和DNA修复）方面具有关键功能。这些酶对健康的重要性是由其活性缺陷引起的各种疾病（包括发育综合征和癌症）所产生的。 CSB重塑器中的突变导致Cockayne综合征：一种与大量发育和神经系统缺陷相关的毁灭性早衰疾病。人类中有30多个已知和预测的ATP依赖性染色质重塑剂。但是，CSB是转录耦合DNA修复（TCR）必不可少的唯一重塑器，该过程迅速消除转录粘贴DNA病变。此外，CSB对于缓解氧化应激至关重要。我们已经在理解这种染色质改造的方式方面取得了重大进步，其功能是独特的。 （1）CSB与染色质的关联受到严格调节。通常，只有10％的CSB是染色质结合的，但是在紫外线照射或氧化应激后，超过90％的CSB成为染色质。这与其他经过良好研究的重塑剂（例如SWI/SNF和ISWI）形成鲜明对比，后者始终与染色质结合。 （2）使用功能分离衍生物，我们证明了CSB的染色质重塑活性对于其在转录耦合DNA修复（TCR）中的功能至关重要。 （3）CSB的染色质重塑活性是可调的：CSB本身显示弱重塑活性（比SWI/SNF或ACF小约20倍）。令人惊讶的是，与NAP1样组蛋白链酮一起，CSB稳健地重塑了与SWI/SNF或ACF相似的水平。 （4）CSB与CTCF蛋白相互作用，并且通过这种相互作用在氧化应激时在CTCF结合位点显着富集。鉴于CTCF是控制高阶染色质结构的关键参与者，因此CSB可以与CTCF合作，以响应氧化应激来重组染色质结构，以促进配位基因表达或有效的DNA修复。该提案中概述的实验将探讨CSB的这些独特特性，以了解该ATP依赖性染色质重塑剂如何独特地等效，以保护细胞免受遗传毒性应激。此外，这项工作可能对Cockayne综合征的原因和机制有了新的启示。