Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China; Department of Chemical Pathology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China.
Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China; Department of Chemical Pathology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China.
Am J Hum Genet. 2020 Feb 6;106(2):202-214. doi: 10.1016/j.ajhg.2020.01.008. Epub 2020 Jan 30.
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.
无细胞游离 DNA（cf.DNA）是一种强大的非侵入性癌症和产前检测生物标志物，它以短片段的形式在血浆中循环。为了阐明 cf.DNA 片段化的生物学特性，我们利用缺乏这些核酸内切酶的小鼠，探索了脱氧核糖核酸酶 1（DNASE1）、脱氧核糖核酸酶 1 样 3（DNASE1L3）和 DNA 片段化因子亚基 β（DFFB）的作用。通过分析每种核酸内切酶缺乏型小鼠 cf.DNA 片段的末端与野生型小鼠的末端，我们表明每种核酸内切酶都有特定的切割偏好，揭示了 cf.DNA 片段化的逐步过程。从本质上讲，我们证明 cf.DNA 首先是在 DFFB、细胞内 DNASE1L3 和其他核酸内切酶的作用下在细胞内产生的。然后，cf.DNA 片段化继续在循环的 DNASE1L3 和 DNASE1 的作用下在细胞外进行。使用肝素破坏核小体结构，我们还表明，10 bp 周期性源自完整核小体结构内 DNA 的切割。总的来说，这项工作建立了 cf.DNA 片段化的模型。
