Role of ATM and RAG in maintaining genome stability during Tcra/d rearrangement.

描述（由申请人提供）：B和T细胞表达识别几乎无限种抗原的受体。这些受体的多样性是通过称为V（D）J重组的过程产生的。这涉及抗原受体基因的重复切割和修复，这些基因分散在几个不同的染色体上。突变受到严格的调控，发生在特定序列的不同染色体位点。这通常是顺利进行的，但有时也会发生错误。已知非同源末端连接（NHEJ）和DNA损伤反应途径的组分中的缺陷易诱发异常V（D）J重排事件，其可导致白血病和淋巴瘤。在很大程度上，损伤谱将取决于每个因子对RAG介导的双链断裂修复的贡献。在这种情况下，个别修复和损伤反应因子的影响已被广泛研究，但除此之外，很少有人知道这些因素是否有调节RAG酶活性或抗原受体基因座和周围基因的可及性的作用，以停止切割后，RAG介导的断裂被引入一个等位基因。这项资助中概述的工作将导致对如何以受控方式引入程序化DNA断裂以保护V（D）J重组期间基因组稳定性的更详细和机械的理解。我们的目标是在发育T细胞的过程中观察基因组的3D组织，以识别与Tcra/d在细胞中重组密切相关的“风险”候选人。此外，我们将确定亲密关系是否是通过活跃转录基因的循环和聚集在转录工厂中来培养的，这些转录工厂可以富集RAG蛋白。核组织和基因活性（已知是RAG靶向的重要决定因素）的联合分析将为我们提供一个起点，从这个起点来解开单个修复因子的影响。重要的是，这种分析将使我们能够将已知的致癌基因易位事件置于全球背景下，以便我们可以开始理解为什么特定基因被靶向并与染色体重排中的Tcra/d结合。)

DESCRIPTION (provided by applicant): B and T cells express receptors that recognize an almost infinite variety of antigens. The diversity of these receptors is generated through a process known as V(D)J recombination. This involves repeated cleavage and repair of antigen receptor genes, which are scattered across several different chromosomes. Recombination is carefully regulated and occurs at different chromosomal loci in a specific sequence. This normally proceeds without mishap, but mistakes do sometimes occur. Deficiencies in components of the nonhomologous end joining (NHEJ) and DNA damage response pathways are known to predispose to aberrant V(D)J rearrangement events that can lead to leukemias and lymphomas. In large part, the spectrum of damage will depend on the contribution of each factor to the repair of RAG-mediated double strand breaks. The effects of individual repair and damage response factors in this context have been extensively studied, but beyond this, little is known about whether any of these factors have a role in regulating RAG enzymatic activity or the accessibility of the antigen receptor loci and surrounding genes to halt cleavage after RAG-mediated breaks are introduced on one allele. The work outlined in this grant will lead to a more detailed and mechanistic understanding of how programmed DNA breaks are introduced in a controlled manner to protect genome stability during V(D)J recombination. We aim to look at the 3D organization of the genome in developing T cells to identify 'at risk' candidates that are closely associated with Tcra/d in cells where they are being recombined. Further, we will determine whether intimate relationships are fostered by looping out of actively transcribed genes and congregation in transcription factories that could be enriched for RAG proteins. The combined analyses of nuclear organization and gene activity (which is known to be an important determinant of RAG targeting) will provide us with a starting point from which to unravel the effects of individual repair factors. Importantly, this analysis will allow us to put known oncogenc translocation events into a global context so that we can begin to understand why particular genes are targeted and join with Tcra/d in chromosomal rearrangements. )