Disparate pathways for extrachromosomal DNA biogenesis and genomic DNA repair.
Disparate pathways for extrachromosomal DNA biogenesis and genomic DNA repair.
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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。
DOI：
10.1101/2023.10.22.563489
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发表时间：
2023
期刊：
影响因子：
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通讯作者：
Chang,HowardY
中科院分区：
文献类型：
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作者：
Rose,JohnC;Wong,IvyTsz-Lo;Daniel,Bence;Jones,MatthewG;Yost,KathrynE;Hung,KingL;Curtis,EllisJ;Mischel,PaulS;Chang,HowardY
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如何形成的理解有限。在这项研究中，我们将基于CRISPR的ecDNA诱导方法与新形成的ecDNA的广泛表征结合起来，以研究它们的生物起源。我们发现，DNA环化是有效的，无论3D基因组背景下，800 kb，1 Mb，和1.8 Mb的ecDNAs的形成达到或超过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-PK催化亚基和ATM的抑制对ecDNA的形成有相反的影响。ecDNA和相应的染色体切除疤痕可以以显著不同的速率形成，并且对DNA-PK催化亚基和ATM抑制的反应不同。总而言之，我们的结果支持了ecDNA形成的模型，其中双链断裂末端在连接非法末端之前与其合法的连接伴侣解离，形成ecDNA和切除疤痕。重要意义我们的研究利用基于CRISPR的方法来检查ecDNA生物发生，揭示了双链断裂之间的有效环化。ecDNA及其相应的染色体瘢痕可以通过非同源末端连接或微同源介导的末端连接形成，但ecDNA和瘢痕形成过程是不同的。基于我们的研究结果，我们建立了切除ecDNA形成的机制模型。
Oncogene amplification on extrachromosomal DNA (ecDNA) is a pervasive driver event in cancer, yet our understanding of how ecDNA forms is limited. In this study, we couple a CRISPR-based method for ecDNA induction with extensive characterization of newly formed ecDNAs to examine their biogenesis. We find that DNA circularization is efficient, irrespective of 3D genome context, with the formation of 800 kb, 1 Mb, and 1.8 Mb ecDNAs reaching or exceeding 15%. We show nonhomologous end joining and microhomology-mediated end joining both contribute to ecDNA formation, whereas inhibition of DNA-PK catalytic subunit 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-PK catalytic subunit 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.SignificanceOur study harnesses a CRISPR-based method to examine ecDNA biogenesis, uncovering efficient circularization between double-strand breaks. ecDNAs and their corresponding chromosomal scars can form via nonhomologous end joining or microhomology-mediated end joining, but the ecDNA and scar formation processes are distinct. Based on our findings, we establish a mechanistic model of excisional ecDNA formation.