Somatic mutation landscapes at single-molecule resolution
Somatic mutation landscapes at single-molecule resolution
复制标题
单分子分辨率的体细胞突变景观
DOI：
10.1038/s41586-021-03477-4
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发表时间：
2021-04-28
期刊：
影响因子：
64.8
通讯作者：
Martincorena, Inigo
中科院分区：
文献类型：
--
作者：
Abascal, Federico;Harvey, Luke M. R.;Martincorena, Inigo
体细胞突变驱动癌症的发展，并可能导致衰老和其他疾病（1，2）。尽管它们很重要，但检测仅存在于单个细胞或小克隆中的突变的困难限制了我们对少数组织的体细胞诱变的了解。在这里，为了克服这些限制，我们开发了纳米孔测序（NanoSeq），这是一种双链体测序方案，在来自细胞群体的单个DNA分子中，每十亿个碱基对的错误率小于5个错误。该速率比典型的体细胞突变负荷低两个数量级，使得能够独立于克隆性研究任何组织中的体细胞突变。我们使用这种单分子敏感性来研究几种组织中非分裂细胞的体细胞突变，比较干细胞和分化细胞，并研究在没有细胞分裂的情况下的诱变。血液和结肠中的分化细胞显示出与相应干细胞非常相似的突变负荷和特征，尽管成熟血细胞经历了相当多的分裂。然后，我们表征了有丝分裂后神经元和多克隆平滑肌的突变景观，证实了神经元在整个生命过程中以恒定的速率积累体细胞突变，而没有细胞分裂，与有丝分裂活性组织的速率相似。总之，我们的研究结果表明，独立于细胞分裂的突变过程是体细胞诱变的重要贡献者。我们预计，可靠地检测单个DNA分子中突变的能力可以改变我们对体细胞诱变的理解，并使大规模队列的非侵入性研究成为可能。
Somatic mutations drive the development of cancer and may contribute to ageing and other diseases(1,2). Despite their importance, the difficulty of detecting mutations that are only present in single cells or small clones has limited our knowledge of somatic mutagenesis to a minority of tissues. Here, to overcome these limitations, we developed nanorate sequencing (NanoSeq), a duplex sequencing protocol with error rates of less than five errors per billion base pairs in single DNA molecules from cell populations. This rate is two orders of magnitude lower than typical somatic mutation loads, enabling the study of somatic mutations in any tissue independently of clonality. We used this single-molecule sensitivity to study somatic mutations in non-dividing cells across several tissues, comparing stem cells to differentiated cells and studying mutagenesis in the absence of celldivision. Differentiated cells in blood and colon displayed remarkably similar mutation loads and signatures to their corresponding stem cells, despite mature blood cells having undergone considerably more divisions. We then characterized the mutational landscape of post-mitotic neurons and polyclonal smooth muscle, confirming that neurons accumulate somatic mutations at a constant rate throughout life without cell division, with similar rates to mitotically active tissues. Together, our results suggest that mutational processes that are independent of cell division are important contributors to somatic mutagenesis. We anticipate that the ability to reliably detect mutations in single DNA molecules could transform our understanding of somatic mutagenesis and enable non-invasive studies on large-scale cohorts.