Role of DNA Double Strand Break Response in Suppression of Thymic Lymphoma

未成熟T细胞淋巴瘤是人类淋巴系统恶性肿瘤的重要组成部分。这些肿瘤中的许多具有复发性染色体易位和相关畸变，其激活原癌基因、抑制肿瘤抑制基因或产生新的致癌融合基因。人类未成熟T细胞淋巴瘤的大多数致癌性易位被认为是通过V（D）J重组期间在T细胞受体（TCR）基因座处引入的DNA双链断裂（DSB）的修复错误和/或在其他基因组位置处的一般DSB而发生的。我们建议阐明DSB反应的功能， 抑制与T细胞淋巴瘤相关的易位，并产生人T细胞淋巴瘤的新型小鼠模型。我们还建议阐明T细胞淋巴瘤复发易位的分子机制，包括空间邻近性，DSB频率和DNA修复途径的可用性如何影响易位模式。在ATM缺陷小鼠胸腺淋巴瘤中经常发现TCR a/d基因座附近的复发性14号染色体易位，并且在具有突变的ATM基因的人T细胞淋巴瘤中发现类似的易位。在这方面，我们发现，一个区域10 Mb的TCR α/d基因座的上游是高度扩增的14号染色体在大多数ATM缺陷型小鼠胸腺淋巴瘤。我们建议充分研究这种在ATM缺陷型T细胞淋巴瘤中反复发生的易位/扩增，i）阐明其机制方面，包括TCR α/d位点V（D）J重组和TCR α/d增强子的潜在作用（与项目2，Harald von Boehmer合作），ii）鉴定靶癌基因（与项目5，Rick Young合作），和iii）确定与人T细胞淋巴瘤的相关性（与项目1 Tom Look合作）。对于易位，不同染色体上的参与基因座必须被破坏，并且必须非常接近以进行连接。因此，我们建议测试的假设，频繁激活某些原癌基因通过易位到TCR基因座在人类，但不是小鼠，T细胞淋巴瘤可能反映了相对频率的DNA DSB和空间接近的目标基因座。为了解决这个问题，我们将采用各种方法，包括3D FISH和新的细胞培养物和小鼠模型的产生，其中DNA断裂在T细胞发育过程中引入靶T细胞癌基因。我们还将使用这些模型来检验我们的假设，即ATM及其底物（例如，H2 AX）通过稳定在V（D）J重组期间引入的TCR基因座DSB来防止由异常V（D）J重组引起的易位。总之，这些研究应该使我们能够解决长期存在的关于T-ALL和其他癌症中染色体易位靶向机制的问题。

Immature T cell lymphomas comprise a significant portion of human lymphoid malignancies. Many of these tumors harbor recurrent chromosomal translocations and related aberrations that either activate proto-oncogenes, inactivate tumor suppressor genes, or create novel oncogenic fusion genes. Most oncogenic translocations of human immature T cell lymphomas are thought to occur via errors in the repair of DNA double strand breaks (DSBs) introduced at T cell receptor (TCR) loci during V(D)J recombination and/or general DSBs at other genomic locations. We propose to elucidate functions of the DSB response in suppression of translocations associated with T cell lymphomas and to generate novel mouse models for human T cell lymphoma. We also propose to elucidate molecular mechanisms that underlie recurrent translocations in T cell lymphomas, including how spatial proximity, DSB frequency and DNA repair pathway availability affect translocation patterns. Recurrent chromosome 14 translocations in the vicinity of the TCRa/d locus are found frequently in ATM-deficient mouse thymic lymphomas and similar translocations are found in human T cell lymphomas that have mutated ATM genes. In this regard, we find that a region 10 Mb upstream of the TCRa/d locus is highly amplified on chromosome 14 in most ATM-deficient mouse thymic lymphomas. We propose to fully investigate this recurrent translocation/amplification in ATM-deficient T cell lymphomas i) to elucidate mechanistic aspects, including potential roles of TCRa/d locus V(D)J recombination and TCRa/d enhancers (with Project 2, Harald von Boehmer), ii) to identify target oncogene(s) (with Project 5, Rick Young), and iii) to determine relevance to human T cell lymphomas (with Project 1 Tom Look). For translocations, participating loci on different chromosomes must be broken and must be in close proximity for joining. Thus we propose to test the hypothesis that frequent activation of certain proto-oncogenes via translocation to TCR loci in human, but not mouse, T cell lymphomas may reflect the relative frequency of DNA DSBs and the spatial proximity of target loci. To address this question, we will employ various approaches including 3D FISH and the generation of novel cell culture and mouse models in which DNA breaks are introduced into target T-cell oncogenes during T-cell development. We will also employ these models to test our hypothesis that ATM and its substrates (e.g., H2AX) prevent translocations resulting from aberrant V(D)J recombination by stabilizing TCR locus DSBs introduced during V(D)J recombination. Together, these studies should allow us to address long-standing questions regarding the mechanisms underlying chromosomal translocation targeting in T-ALL and other cancers.