Oncogene amplification on extrachromosomal DNA (ecDNA) is a pervasive driver event in cancer, yet our understanding of how ecDNA forms is limited. Here, we couple a CRISPR-based method for ecDNA induction with extensive characterization of newly formed ecDNA to examine their biogenesis. We find that DNA circularization is efficient, irrespective of 3D genome context, with formation of 800kb, 1 Mb, and 1.8 Mb ecDNAs reaching or exceeding 15%. We show non-homologous end joining and microhomology-mediated end joining both contribute to ecDNA formation, while inhibition of DNA-PKcs and ATM have opposing impacts on ecDNA formation. EcDNA and the corresponding chromosomal excision scar can form at significantly different rates and respond differently to DNA-PKcs and ATM inhibition. Taken together, our results support a model of ecDNA formation in which double strand break ends dissociate from their legitimate ligation partners prior to joining of illegitimate ends to form the ecDNA and excision scar.

中文翻译：

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染色体外 DNA 生物发生和基因组 DNA 修复的不同途径


染色体外 DNA (ecDNA) 上的癌基因扩增是癌症中普遍存在的驱动事件，但我们对 ecDNA 如何形成的了解有限。在这里，我们将基于 CRISPR 的 ecDNA 诱导方法与新形成的 ecDNA 的广泛表征相结合，以检查其生物发生。我们发现，无论 3D 基因组背景如何，DNA 环化都是有效的，800kb、1 Mb 和 1.8 Mb ecDNA 的形成率达到或超过 15%。我们发现非同源末端连接和微同源介导的末端连接都有助于 ecDNA 的形成，而 DNA-PKcs 和 ATM 的抑制对 ecDNA 的形成具有相反的影响。 EcDNA 和相应的染色体切除疤痕可以以显着不同的速率形成，并且对 DNA-PKcs 和 ATM 抑制的反应不同。综上所述，我们的结果支持了一种 ecDNA 形成模型，其中双链断裂末端在连接非法末端以形成 ecDNA 和切除疤痕之前与其合法连接伙伴分离。