PATHWAYS THAT PRESERVE GENOME STABILITY DURING ANTIGEN RECEPTOR GENE ASSEMBLY

抗原受体基因组装过程中保持基因组稳定性的途径

• 批准号: 8649628
• 负责人: Bo-Ruei Chen
• 金额: $ 5.51万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-02-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8649628
• 关键词: Antibodies; Antigen Receptors; Apoptosis; Ataxia-Telangiectasia-Mutated protein kinase; B-Lymphocytes; Binding; Biological Assay; Cell Cycle Arrest; Cell physiology; Cells; Chromatin; Chromatin Structure; Chromosomal translocation; Chromosome Deletion; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Repair Pathway; DNA Sequence; DNA Sequence Rearrangement; DNA lesion; DNA repair protein; Double Strand Break Repair; Enzymes; Event; Excision; Exons; G1 Phase; G2 Phase; Genome Stability; Genomic Instability; Genotoxic Stress; Histones; Human; In Vitro; Infection; Ionizing radiation; Label; Laboratories; Lead; Lesion; Lymphocyte; Lymphocyte antigen; Lymphoid; Malignant Neoplasms; Malignant lymphoid neoplasm; Mediating; Modification; Molecular; Monitor; Nonhomologous DNA End Joining; Oncogenic; Pathway interactions; Phosphorylation; Process; Proteins; Receptor Gene; Recruitment Activity; Research Proposals; Resolution; Role; Signal Transduction; Single-Stranded DNA; Structure; T-Cell Receptor; Testing; Variant; base; chromatin immunoprecipitation; endonuclease; fighting; histone modification; homologous recombination; in vitro Assay; in vivo; insight; leukemia/lymphoma; novel; nuclease; physical separation; prevent; public health relevance; receptor; repair enzyme; repaired; response; sensor; tumorigenesis

DESCRIPTION (provided by applicant): DNA double strand breaks (DSBs) are perhaps the most harmful among all types of DNA damage because they involve physical separation of DNA molecules. If un-repaired or repaired aberrantly, DSBs can lead to loss (i.e. deletion) or abnormal rearrangement (i.e. translocation) of chromosomes, which can drive the genesis of a wide variety of cancers. DSBs can be generated by genotoxic stress, such as ionizing radiation, or as intermediates of normal cellular activities, such as the assembly of antigen receptor genes in lymphocytes that encode antibodies and T cell receptors that are key to fighting infection. During the assembly process, DNA DSBs are introduced at antigen receptor gene segments, and then repaired to generate the final DNA sequence encoding antigen receptor proteins. However, under certain circumstances, such DSBs cannot be properly repaired and eventually lead to the oncogenic deletions or translocations commonly seen in leukemias and lymphomas. Our laboratory previously identified a novel pathway that has an important role in protecting DSBs at antigen receptor genes from undergoing inappropriate processing by cellular enzymes and aberrant resolution. This pathway depends on the modification of a histone protein, H2AX, that is closely associated with cellular DNA. In response to DNA double strand breaks (DSBs), H2AX is phosphorylated by DNA damage sensor kinase ATM to form ?-H2AX that functions to recruit DNA damage response and repair proteins, and mediates additional modifications on other histone proteins. We have found that the phosphorylation of H2AX associated with broken DNA ends prevents processing of these DNA ends in ways that could lead to their aberrant repair. Here we propose to elucidate the mechanisms by which H2AX functions to prevent aberrant DSB repair. We hypothesize that ?-H2AX maintains DSB end structure through mechanisms that include modulating chromatin structure and DNA end accessibility through directing additional histone PTMs. In addition, given that ?-H2AX is shown to regulate protein association at DSBs, we predict that ?-H2AX can also impact the association of nucleases at DSB ends. To test our hypothesis, we will first reveal which histone modifications are generated in response to DSBs in a ?-H2AX-dependent manner and then determine whether ?-H2AX and ?-H2AX-dependent histone modifications modulate DNA end structures in a way that renders them less susceptible to degradation by nucleases. We will then determine whether ?-H2AX may prevent aberrant DNA end processing by limiting the association of nucleases with DSB ends. These studies will further our understanding of the genesis of chromosomal lesions that lead to lymphoid transformation and have broader implications for our understanding of the basis for genome instability in a variety of cancers.

描述（由申请人提供）：DNA双链断裂（DSB）可能是所有类型的DNA损伤中最有害的，因为它们涉及DNA分子的物理分离。如果不理或修复不正当，DSB会导致染色体的损失（即缺失）或异常重排（即易位），这可以驱动各种癌症的起源。 DSB可以通过遗传毒性应激（例如电离辐射）或正常细胞活性的中间体产生，例如淋巴细胞中抗原受体基因的组装，这些抗原受体基因编码抗体和T细胞受体，这些受体和T细胞受体是对抗感染的关键。在组装过程中，在抗原受体基因段上引入DNA DSB，然后修复以生成编码抗原受体蛋白的最终DNA序列。但是，在某些情况下，这种DSB无法正确修复，并最终导致白血病和淋巴瘤中常见的致癌缺失或易位。我们的实验室先前确定了一种新的途径，该途径在保护抗原受体基因的DSB中具有重要作用，以免通过细胞酶和异常分辨率进行不适当的处理。该途径取决于与细胞DNA密切相关的组蛋白蛋白H2AX的修饰。 为了响应DNA双链断裂（DSB），H2AX被DNA损伤传感器激酶ATM磷酸化以形成？-H2AX，其功能可募集DNA损伤反应和修复蛋白，并介导其他组蛋白的其他修饰。 我们发现，与断裂的DNA末端相关的H2AX的磷酸化阻止了这些DNA末端的处理，从而导致其异常修复。 在这里，我们建议阐明H2AX功能防止异常DSB修复的机制。我们假设？-H2AX通过机制维持DSB末端结构，包括通过指导其他组蛋白PTM来调节染色质结构和DNA最终可及性。另外，鉴于显示的？-H2AX被证明可以调节DSB的蛋白质关联，我们预测？-H2AX也会影响DSB末端的核酸酶的缔合。为了检验我们的假设，我们将首先揭示以？-H2AX依赖性方式对DSB产生哪些组蛋白修饰，然后确定？-H2AX和？-H2AX依赖性组蛋白修饰是否以一种使它们不太容易通过核裂解的方式调节DNA终端结构。然后，我们将通过限制核酸酶与DSB末端的关联来确定？-H2AX是否可以防止异常的DNA末端处理。 这些研究将进一步了解导致淋巴样转化的染色体病变的起源，并对我们对各种癌症基因组不稳定性基础的理解具有更广泛的意义。