PNAS：MPE序列，对染色质结构的全基因组分析的一种新方法

摘要 : 日前，加州大学的研究团队开发了对染色质结构进行全基因组分析的新方法，MPE-seq（methidiumpropyl-EDTA sequencing）。这一成果发表在六月十五日的美国国家科学院院刊PNAS杂志上，文章的通讯作者是加州大学的任兵（Bing Ren）教授和James T. Kadonaga。

高等生物的细胞核负责储存基因组DNA，这些DNA环绕着组蛋白形成碟状的核小体结构。基因组DNA以这样的形式包装成为染色质，使DNA受到良好的保护。染色质结构对于DNA转录、复制和修复非常关键，决定着基因的表达和细胞的生理状态。人们通常使用核酸酶MNase和DNase I进行染色质结构分析，但在某些情况下这样的酶切分析并不能很好的反映真实情况。

日前，加州大学的研究团队开发了对染色质结构进行全基因组分析的新方法，MPE-seq(methidiumpropyl-EDTA sequencing)。这一成果发表在六月十五日的美国国家科学院院刊PNAS杂志上，文章的通讯作者是加州大学的任兵(Bing Ren)教授和James T. Kadonaga。

任兵教授早年毕业于中国科技大学，现为加州大学圣地亚哥分校Ludwig癌症研究所基因调控实验室主任，主要从事哺乳动物细胞基因调控网络分析及细胞表观遗传学调控机制的研究。近年来在science、Nature,、Cell国际权威杂志上发表了一系列重要的研究成果。

MPE-seq主要是用MPE-Fe(II)消化染色质，然后进行大规模平行测序。跟MNase一样，MPE-Fe(II)也优先切割核小体之间的连接DNA，不过 MPE-Fe(II)基本没有序列偏好。

研究显示，MPE-Fe(II)和MNase切割细胞核染色质是有差异的，在活跃的启动子区域尤为明显。MPE-seq会检测到核小体大小(141–190 bp)和亚核小体大小(比如101–140 bp)的染色质片段，但MNase-seq检测不到。

进一步研究表明，这些片段与核心组蛋白的出现存在一一对应的关系，应该和非经典的染色质结构有关。值得注意的是，亚核小体大小的片段与活跃启动子存在特别明显的关联。

研究指出，MPE-Fe(II)切割染色质时基本没有序列偏好，而且MPE-seq可以揭示活跃启动子区域的非经典染色质结构。这是一种直观的染色质结构分析法，可以成为MNase-seq的重要补充，揭示在基因表达调控中具有重要意义的染色质结构。

原文标题：MPE-seq, a new method for the genome-wide analysis of chromatin structure

原文摘要：The analysis of chromatin structure is essential for the understanding of transcriptional regulation in eukaryotes. Here we describe methidiumpropyl-EDTA sequencing (MPE-seq), a method for the genome-wide characterization of chromatin that involves the digestion of nuclei withMPE-Fe(II) followed by massively parallel sequencing. Like micrococcal nuclease (MNase), MPE-Fe(II) preferentially cleaves the linker DNA between nucleosomes. However, there are differences in the cleavage of nuclear chromatin by MPE-Fe(II) relative to MNase. Most notably, immediately upstream of the transcription start site of active promoters, we frequently observed nucleosome-sized (141–190 bp) and subnucleosome-sized (such as 101–140 bp) peaks of digested chromatin fragments with MPE-seq but not with MNase-seq. These peaks also correlate with the presence of core histones and could thus be due, at least in part, to noncanonical chromatin structures such as labile nucleosome-like particles that have been observed in other contexts. The subnucleosome-sized MPE-seq peaks exhibit a particularly distinct association with active promoters. In addition, unlike MNase, MPE-Fe(II) cleaves nuclear DNA with little sequence bias. In this regard, we found that DNA sequences at RNA splice sites are hypersensitive to digestion by MNase but not by MPE-Fe(II). This phenomenon may have affected the analysis of nucleosome occupancy over exons. These findings collectively indicate that MPE-seq provides a unique and straightforward means for the genome-wide analysis of chromatin structure with minimal DNA sequence bias. In particular, the combined use of MPE-seq and MNase-seq enables the identification of noncanonical chromatin structures that are likely to be important for the regulation of gene expression.

作者：秩名

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