Massively multiplex single-molecule oligonucleosome footprinting.
Massively multiplex single-molecule oligonucleosome footprinting.
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DOI：
10.7554/elife.59404
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发表时间：
2020-12-02
期刊：
影响因子：
7.7
通讯作者：
Ramani V
中科院分区：
文献类型：
--
作者：
Abdulhay NJ;McNally CP;Hsieh LJ;Kasinathan S;Keith A;Estes LS;Karimzadeh M;Underwood JG;Goodarzi H;Narlikar GJ;Ramani V
我们对核小体串珠状排列的理解主要建立在高分辨率的序列不可知成像方法和序列分辨的批量生物化学技术上。为了弥合这些方法之间的鸿沟，我们提出了单分子腺嘌呤甲基化核糖体测序分析（SAMOSA）。SAMOSA是一种高通量单分子测序方法，它结合了腺嘌呤甲基转移酶足迹和单分子实时DNA测序，以天然和非破坏性地测量单个染色质纤维上的核小体位置。SAMOSA数据允许无偏分类的单个染色质纤维上的核小体占用的单分子“状态”。我们利用这一点来估计核小体的规律性和间距单染色质纤维全基因组，在预测的转录因子结合基序，并在整个人类表观基因组域。我们的分析表明，染色质由规则和不规则的单分子染色体模式组成，这些模式在表观基因组结构域的相对丰度上存在细微差异。这种不规则性在组成型异染色质中特别引人注目，它通常被视为构象静态实体。我们的概念验证研究提供了一种强大的新方法，用于以以前难以处理的分辨率研究核小体组织，并为建模和可视化高阶染色质结构提供了新的途径。
Our understanding of the beads-on-a-string arrangement of nucleosomes has been built largely on high-resolution sequence-agnostic imaging methods and sequence-resolved bulk biochemical techniques. To bridge the divide between these approaches, we present the single-molecule adenine methylated oligonucleosome sequencing assay (SAMOSA). SAMOSA is a high-throughput single-molecule sequencing method that combines adenine methyltransferase footprinting and single-molecule real-time DNA sequencing to natively and nondestructively measure nucleosome positions on individual chromatin fibres. SAMOSA data allows unbiased classification of single-molecular 'states' of nucleosome occupancy on individual chromatin fibres. We leverage this to estimate nucleosome regularity and spacing on single chromatin fibres genome-wide, at predicted transcription factor binding motifs, and across human epigenomic domains. Our analyses suggest that chromatin is comprised of both regular and irregular single-molecular oligonucleosome patterns that differ subtly in their relative abundance across epigenomic domains. This irregularity is particularly striking in constitutive heterochromatin, which has typically been viewed as a conformationally static entity. Our proof-of-concept study provides a powerful new methodology for studying nucleosome organization at a previously intractable resolution and offers up new avenues for modeling and visualizing higher order chromatin structure.