Effect of Spatial Proximity on Chromosomal Translocations

描述（由申请人提供）：空间邻近对染色体易位的影响染色体易位是一种已知与癌症发生和进展有关的基因组重排，它涉及两个非同源染色体之间遗传物质的交换。在某些癌症中，特别是白血病、淋巴瘤和肉瘤，特异性易位是复发性的，这意味着不同的患者在相同的两条染色体上有相同的两个位点易位。这种易位的重复导致了这样一种假设，即基因组的某些区域更容易重排，可能是通过核空间的紧密空间接近。尽管使用荧光原位杂交（FISH）技术等相关证据表明基因组常见易位区域的空间接近性，但空间接近有助于染色体易位形成的观点尚未得到直接或彻底的验证。我们的实验室开发了一种新的遗传系统，我们可以检测非同源末端连接（NHEJ）介导的酿酒葡萄球菌模式生物的染色体易位。该系统允许在分离的染色体上同时诱导DNA双工断裂，并实时检测互惠染色体易位。此外，酵母基因组最近在三维空间中被绘制，揭示了细胞核中非随机染色体区域的模式，着丝粒聚集在纺锤极体附近，染色体臂伸出，端粒系在核膜上。该提案的重点是直接测试空间邻近有助于形成复发性染色体易位的两种方式的想法。首先，利用三维基因组图谱作为指导，两个DNA断裂位点将被放置在基因组中的不同位置，彼此之间的空间距离不同。染色体易位形成的频率将与空间距离相关，通过染色体构象捕获（3C）测定。接下来，在这些特定的位置，染色质结构将通过改变细胞周期阶段和通过删除端粒-与核周围的连接相关的因子来修饰。空间接近度和染色体易位频率将分别使用3C测定和易位遗传菌株进行评估。这项研究将揭示导致染色体易位的分子机制，并为描述人类的等效过程奠定基础。

DESCRIPTION (provided by applicant): Effect of Spatial Proximity on Chromosomal Translocations Chromosomal translocations are a type of genomic rearrangement known to be involved in cancer development and progression, and they involve the exchange of genetic material between two non- homologous chromosomes. In certain cancers, especially leukemias, lymphomas, and sarcomas, specific translocations are recurrent, meaning different patients present with translocations involving the same two loci on the same two chromosomes. This recurrence of certain translocations leads to the hypothesis that certain regions of the genome are more susceptible to rearrangement, likely through a close spatial proximity in the nuclear space. Despite correlative evidence demonstrating close spatial proximity of commonly translocated regions of the genome using, for instance, Fluorescent In-Situ Hybridization (FISH) techniques, the idea that spatial proximity contributes to the formation of chromosomal translocations has not be directly or thoroughly tested. Our laboratory developed a novel genetic system with which we can detect Non-Homologous End- Joining (NHEJ)-mediated reciprocal chromosomal translocations in the S. cerevisiae model organism. This system allows for the simultaneous induction of DNA duplex breaks on separate chromosomes, and the real time detection of reciprocal chromosomal translocations. Additionally, the yeast genome has recently been mapped in three-dimensional space, revealing the patterns of non-random chromosomal territories in the nucleus, with the centromeres clustered near the spindle pole body, the chromosome arms outstretched, and the telomeres tethered to the nuclear membrane. The focus of this proposal is to directly test the idea that spatial proximity contributes to the formation of recurrent chromosomal translocations in two ways. First, using the three-dimensional genome map as a guide, two DNA break-sites will be placed at different locations in the genome, at different spatial distances from each other. The frequency of chromosomal translocation formation will be correlated to the spatial distance, as measure by the Chromosome Conformation Capture (3C) assay. Next, at these specified locations, the chromatin architecture will be modified by changing the cell cycle phase and by deleting factors involved in the telomere-tethering to the nuclear periphery. Spatial proximity and chromosomal translocation frequency will be assessed using the 3C assay and the translocation genetic strain, respectively. This study will shed light on the molecular mechanism leading to chromosomal translocations and set the stage for characterizing the equivalent processes in humans. PUBLIC HEALTH RELEVANCE: The long-term goal of our study is to dissect a molecular mechanism that contributes to chromosomal translocations. Chromosomal translocations are a type of genomic rearrangement involved in cancer development and progression, and the proposed research will characterize how the chromatin architecture may cause susceptibility to this mutagenic process. This research will identify and characterize the mechanisms involved in translocation formation to potentially uncover novel therapeutic strategies aimed at preventing or hindering the formation of genomic rearrangements, such as translocations.