Cell-free DNA (cf.DNA) is a powerful noninvasive biomarker for cancer and prenatal testing, and it circulates in plasma as short fragments. To elucidate the biology of cf.DNA fragmentation, we explored the roles of deoxyribonuclease 1 (DNASE1), deoxyribonuclease 1 like 3 (DNASE1L3), and DNA fragmentation factor subunit beta (DFFB) with mice deficient in each of these nucleases. By analyzing the ends of cf.DNA fragments in each type of nuclease-deficient mice with those in wild-type mice, we show that each nuclease has a specific cutting preference that reveals the stepwise process of cf.DNA fragmentation. Essentially, we demonstrate that cf.DNA is generated first intracellularly with DFFB, intracellular DNASE1L3, and other nucleases. Then, cf.DNA fragmentation continues extracellularly with circulating DNASE1L3 and DNASE1. With the use of heparin to disrupt the nucleosomal structure, we also show that the 10 bp periodicity originates from the cutting of DNA within an intact nucleosomal structure. Altogether, this work establishes a model of cf.DNA fragmentation.

中文翻译：

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无细胞DNA片段化的生物学以及DNASE1，DNASE1L3和DFFB的作用。

无细胞DNA（cf.DNA）是用于癌症和产前检测的强大的非侵入性生物标志物，它以短片段的形式在血浆中循环。为了阐明cf.DNA片段断裂的生物学特性，我们探索了脱氧核糖核酸酶1（DNASE1），像3一样的脱氧核糖核酸酶1（DNASE1L3）和DNA断裂因子亚基β（DFFB）与每种核酸酶缺乏的小鼠的作用。通过分析每种核酸酶缺陷型小鼠与野生型小鼠中cf.DNA片段的末端，我们表明每种核酸酶具有特定的切割偏好，揭示了cf.DNA片段化的逐步过程。从本质上讲，我们证明cf.DNA首先与DFFB，细胞内DNASE1L3和其他核酸酶一起在细胞内生成。然后，cf.DNA片段继续在细胞外循环DNASE1L3和DNASE1。通过使用肝素破坏核小体结构，我们还显示10 bp的周期性起源于完整核小体结构内DNA的切割。总之，这项工作建立了cf.DNA片段化的模型。