High-throughput sequence-based epigenomic analysis of Alu repeats in human cerebellum.
High-throughput sequence-based epigenomic analysis of Alu repeats in human cerebellum.
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DOI：
10.1093/nar/gkp393
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发表时间：
2009-07
影响因子：
14.9
通讯作者：
Soares MB
中科院分区：
文献类型：
--
作者：
Xie H;Wang M;Bonaldo Mde F;Smith C;Rajaram V;Goldman S;Tomita T;Soares MB
DNA甲基化是哺乳动物DNA唯一已知的共价修饰，主要发生在CpG二核苷酸中。人类基因组中51%的CpG位于重复序列中，25%位于Alu元件中。尽管如此，还没有报道用于大规模确定重复序列中CpG甲基化的方法。在这里，我们描述了一种基于测序的策略，用于平行测定数千个Alu重复序列的CpG甲基化状态，以及一种计算算法来设计引物，使其能够从亚硫酸氢盐转化的基因组DNA中特异性扩增。使用单一引物对，我们产生了高序列复杂性的扩增子，并从31 178个Alu元件及其5′侧翼序列中获得CpG甲基化数据，总共代表了超过4 Mb的人类小脑表观基因组。Alu甲基化组分析表明，Alu元件的甲基化水平在内含子和基因间区域较高，但在靠近转录起始位点的区域较低。鉴定了几种低甲基化的Alu元素，并通过焦磷酸测序验证了它们的低甲基化状态。有趣的是，一些Alu元件表现出惊人的组织特异性甲基化模式。我们预期本文所述的扩增子证明作为表观基因组表征是无价的，以监测正常发育期间、衰老和疾病（如癌症）中的表观基因组改变。
DNA methylation, the only known covalent modification of mammalian DNA, occurs primarily in CpG dinucleotides. 51% of CpGs in the human genome reside within repeats, and 25% within Alu elements. Despite that, no method has been reported for large-scale ascertainment of CpG methylation in repeats. Here we describe a sequencing-based strategy for parallel determination of the CpG-methylation status of thousands of Alu repeats, and a computation algorithm to design primers that enable their specific amplification from bisulfite converted genomic DNA. Using a single primer pair, we generated amplicons of high sequence complexity, and derived CpG-methylation data from 31 178 Alu elements and their 5′ flanking sequences, altogether representing over 4 Mb of a human cerebellum epigenome. The analysis of the Alu methylome revealed that the methylation level of Alu elements is high in the intronic and intergenic regions, but low in the regions close to transcription start sites. Several hypomethylated Alu elements were identified and their hypomethylated status verified by pyrosequencing. Interestingly, some Alu elements exhibited a strikingly tissue-specific pattern of methylation. We anticipate the amplicons herein described to prove invaluable as epigenome representations, to monitor epigenomic alterations during normal development, in aging and in diseases such as cancer.
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