Howard Hughes Medical Institute, Immune Disease Institute, Program in Cellular and Molecular Medicine, Children's Hospital Boston and Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
Cell. 2011 Sep 30;147(1):107-19. doi: 10.1016/j.cell.2011.07.049.
Whereas chromosomal translocations are common pathogenetic events in cancer, mechanisms that promote them are poorly understood. To elucidate translocation mechanisms in mammalian cells, we developed high-throughput, genome-wide translocation sequencing (HTGTS). We employed HTGTS to identify tens of thousands of independent translocation junctions involving fixed I-SceI meganuclease-generated DNA double-strand breaks (DSBs) within the c-myc oncogene or IgH locus of B lymphocytes induced for activation-induced cytidine deaminase (AID)-dependent IgH class switching. DSBs translocated widely across the genome but were preferentially targeted to transcribed chromosomal regions. Additionally, numerous AID-dependent and AID-independent hot spots were targeted, with the latter comprising mainly cryptic I-SceI targets. Comparison of translocation junctions with genome-wide nuclear run-ons revealed a marked association between transcription start sites and translocation targeting. The majority of translocation junctions were formed via end-joining with short microhomologies. Our findings have implications for diverse fields, including gene therapy and cancer genomics.
虽然染色体易位在癌症中是常见的致病事件，但促进它们的机制还知之甚少。为了阐明哺乳动物细胞中的易位机制，我们开发了高通量、全基因组易位测序（HTGTS）。我们利用 HTGTS 来鉴定数以万计的独立易位连接点，这些连接点涉及到固定的 I-SceI 大片段酶产生的 DNA 双链断裂（DSB），这些 DSB 位于激活诱导的胞嘧啶脱氨酶（AID）依赖性 IgH 类转换诱导的 B 淋巴细胞中的 c-myc 癌基因或 IgH 基因座内。DSB 广泛地在整个基因组中转位，但优先靶向转录的染色体区域。此外，许多 AID 依赖性和 AID 非依赖性热点也被靶向，后者主要包括隐匿的 I-SceI 靶点。将易位连接点与全基因组核运转进行比较，揭示了转录起始位点与易位靶向之间的显著关联。大多数易位连接点是通过短微同源性的末端连接形成的。我们的发现对包括基因治疗和癌症基因组学在内的多个领域都具有重要意义。
