Molecular Pathways in Suppression and Development of T Lineage Lymphomas

描述（由申请人提供）：染色体DMA双链断裂（DSB）由多种因素诱导，并在每个细胞中持续发生。DSB的正确修复对于维持基因组稳定性和防止细胞转化至关重要。我们最近证明，共济失调毛细血管扩张突变（ATM）蛋白，这是细胞DSB反应的主调节器，需要在染色体DSB修复，以维持修复复合物中的断裂DNA末端。ATM的直接底物是H2 AX，其是在DSB周围的染色质中磷酸化的核心组蛋白变体。我们已经证明H2 ax的两个等位基因拷贝的表达对于无错误的DSB修复、基因组稳定性的维持和癌症的抑制是必不可少的。值得注意的是，H2 AX基因的杂合性缺失（洛）通常出现在各种各样的人类肿瘤中，这表明类似的剂量依赖性H2 AX在man. In此应用程序中的功能，我们建议阐明机制，H2 AX的ATM和相关激酶的下游功能，以促进无错误的DSB修复，保持基因组的稳定性，并抑制恶性转化。有待研究的具体假设是，H2 AX的一个关键功能是将断裂的DNA链保持在一起，并确保DNA末端得到正确修复。在具体目标1中，我们将测试我们的假设，即H2 AX在V（D）J重组过程中发挥作用，在G1到S转换时将断裂的DNA链保持在一起，从而防止抗原受体基因座易位。我们将采用新的细胞系和原代胸腺细胞在G1期细胞的特定基因组位置诱导DSB中间体，并在细胞周期的持续进展中监测它们的修复。在具体目标2中，我们将测试我们的假设，即其他激酶下游的ATM非依赖性H2 AX功能对于正常DSB修复、预防基因组不稳定性和抑制癌症是必不可少的。我们将利用H2 AX和ATM缺陷的细胞系和胸腺细胞来阐明ATM独立的H2 AX功能的作用。在具体目标3中，我们将测试我们的假设，即H2 AX在正常和癌基因驱动的增殖过程中诱导的复制相关DSB中发挥作用，以防止p53缺失导致恶性转化。我们将直接评估H2 AX抑制p53缺失肿瘤的能力。相关性：我们的研究将使我们更好地理解基因组完整性得以维持的分子机制，以及这些缺陷如何导致恶性转化。这些知识和产生的动物模型将有助于开发更有效的人类癌症诊断和/或治疗工具。因此，我们提出的研究将对人类疾病产生广泛的影响。

DESCRIPTION (provided by applicant): Chromosomal DMA double strand breaks (DSBs) are induced by a variety of factors and occur constantly in every cell. The proper repair of DSBs is essential for maintaining genomic stability and preventing cellular transformation. We recently demonstrated that the Ataxia Telangiectasia Mutated (ATM) protein, which is the master regulator of the cellular DSB response, is required during chromosomal DSB repair to maintain broken DNA ends in repair complexes. An immediate substrate of ATM is H2AX, a core histone variant that is phosphorylated in chromatin around DSBs. We have demonstrated that expression of both allelic copies of H2ax is essential for error-free DSB repair, maintanence of genomic stability, and suppression of cancer. Notably, deletion or loss of heterozygosity (LOH) of the H2AX gene appears commonly in a wide variety of human tumors, suggesting similar dosage-dependent H2AX functions in man. In this application, we propose to elucidate mechanisms by which H2AX functions downstream of ATM and related kinases to promote error-free DSB repair, maintain genomic stability, and suppress malignant transformation. The specific hypothesis to be investigated is that one critical function of H2AX is to hold broken DNA strands together and ensure that DNA ends are properly repaired. In Specific Aim 1, we will test our hypothesis that H2AX functions during V(D)J recombination to hold together broken DNA strands upon G1 to S transition and, thereby, prevent antigen receptor locus translocations. We will employ novel cell lines and primary thymocytes to induce DSB intermediates at specific genomic locations in G1 phase cells and monitor their repair during continued cell cycle progression. In Specific Aim 2, we will test our hypothesis that ATM independent H2AX functions downstream of other kinases are essential for normal DSB repair, prevention of genomic instability, and suppression of cancer. We will utilize cell lines and thymocyes deficient for H2AX and ATM to elucidate the roles of ATM independent H2AX functions. In Specific Aim 3, we will test our hypothesis that H2AX functions at replication-associated DSBs induced during normal and oncogene-driven proliferation to prevent p53 deletions that drive malignant transformation. We will directly evaluate the ability of H2AX to suppress tumors with p53 deletions. Relevance: Our studies will lead to a greater understanding of the molecular mechanisms through which genomic integrity is maintained and how defects in these can lead to malignant transformation. This knowledge and the animal models generated will contribute to the development of more effective diagnostic and/or therapeutic tools for human cancers. Consequently, our proposed studies will have broad implications for human disease.