The human ribosomal DNA array is composed of highly homogenized tandem clusters.
The human ribosomal DNA array is composed of highly homogenized tandem clusters.
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DOI：
10.1101/gr.275838.121
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发表时间：
2021-11
期刊：
影响因子：
7
通讯作者：
Kobayashi T
中科院分区：
文献类型：
--
作者：
Hori Y;Shimamoto A;Kobayashi T
人类核糖体DNA （rDNA）簇的结构由于其高度重复性而难以分析。然而，最近长读测序技术的发展，如牛津纳米孔测序，使我们能够研究基因组的大规模结构。利用这项技术，我们发现人类细胞具有相当规则的rDNA结构。尽管每个人类rDNA拷贝在其非编码区都有一些差异，但连续的rDNA拷贝是相似的，这表明通过基因转换的均质化经常发生在拷贝之间。通过纳米孔测序进一步分析rDNA的甲基化，发现所有非编码区都严重甲基化，而大约一半的编码区明显未甲基化。在rDNA拷贝数较高的个体中，推测具有转录活性的未甲基化拷贝的比例较低，这表明存在一种保持活性拷贝数稳定的机制。此外，与对照正常细胞相比，DNA修复活性降低的类早衰综合征患者细胞中的rDNA具有更多不稳定拷贝，尽管这一比率远低于先前使用纤维- fish方法报道的比率。总的来说，我们的研究结果阐明了人类细胞中rDNA稳定性和转录调控的观点，表明存在均质化机制，以确保序列质量和维持细胞功能的活性拷贝。
The structure of the human ribosomal DNA (rDNA) cluster has traditionally been hard to analyze owing to its highly repetitive nature. However, the recent development of long-read sequencing technology, such as Oxford Nanopore sequencing, has enabled us to study the large-scale structure of the genome. Using this technology, we found that human cells have a quite regular rDNA structure. Although each human rDNA copy has some variations in its noncoding region, contiguous copies of rDNA are similar, suggesting that homogenization through gene conversion frequently occurs between copies. Analysis of rDNA methylation by Nanopore sequencing further showed that all the noncoding regions are heavily methylated, whereas about half of the coding regions are clearly unmethylated. The ratio of unmethylated copies, which are speculated to be transcriptionally active, was lower in individuals with a higher rDNA copy number, suggesting that there is a mechanism that keeps the active copy number stable. In addition, the rDNA in progeroid syndrome patient cells with reduced DNA repair activity had more unstable copies compared with control normal cells, although the rate was much lower than previously reported using a fiber-FISH method. Collectively, our results clarify the view of rDNA stability and transcription regulation in human cells, indicating the presence of mechanisms for both homogenizations to ensure sequence quality and maintenance of active copies for cellular functions.
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