Duke Institute for Genome Sciences and Policy, Duke University, Durham, North Carolina, USA.
PLoS Genet. 2010 Aug 12;6(8):e1001061. doi: 10.1371/journal.pgen.1001061.
Genome rearrangement often produces chromosomes with two centromeres (dicentrics) that are inherently unstable because of bridge formation and breakage during cell division. However, mammalian dicentrics, and particularly those in humans, can be quite stable, usually because one centromere is functionally silenced. Molecular mechanisms of centromere inactivation are poorly understood since there are few systems to experimentally create dicentric human chromosomes. Here, we describe a human cell culture model that enriches for de novo dicentrics. We demonstrate that transient disruption of human telomere structure non-randomly produces dicentric fusions involving acrocentric chromosomes. The induced dicentrics vary in structure near fusion breakpoints and like naturally-occurring dicentrics, exhibit various inter-centromeric distances. Many functional dicentrics persist for months after formation. Even those with distantly spaced centromeres remain functionally dicentric for 20 cell generations. Other dicentrics within the population reflect centromere inactivation. In some cases, centromere inactivation occurs by an apparently epigenetic mechanism. In other dicentrics, the size of the alpha-satellite DNA array associated with CENP-A is reduced compared to the same array before dicentric formation. Extra-chromosomal fragments that contained CENP-A often appear in the same cells as dicentrics. Some of these fragments are derived from the same alpha-satellite DNA array as inactivated centromeres. Our results indicate that dicentric human chromosomes undergo alternative fates after formation. Many retain two active centromeres and are stable through multiple cell divisions. Others undergo centromere inactivation. This event occurs within a broad temporal window and can involve deletion of chromatin that marks the locus as a site for CENP-A maintenance/replenishment.
基因组重排经常产生带有两个着丝粒（双着丝粒）的染色体，由于细胞分裂过程中桥的形成和断裂，这些染色体天生不稳定。然而，哺乳动物的双着丝粒，特别是人类的双着丝粒，可以非常稳定，通常是因为一个着丝粒的功能被沉默了。由于很少有系统可以实验性地产生人类双着丝粒染色体，因此着丝粒失活的分子机制还不太清楚。
在这里，我们描述了一种富集新形成的双着丝粒的人类细胞培养模型。我们证明，人类端粒结构的瞬时破坏会非随机地产生涉及近端着丝粒染色体的双着丝粒融合。诱导的双着丝粒在融合断点附近的结构上有所不同，并且与自然发生的双着丝粒一样，表现出不同的着丝粒间距离。许多功能性双着丝粒在形成后可以持续数月。即使那些着丝粒间距离较远的双着丝粒在 20 个细胞世代中仍然保持功能性双着丝粒状态。群体中的其他双着丝粒反映了着丝粒失活。在某些情况下，着丝粒失活是通过表观遗传机制发生的。在其他双着丝粒中，与 CENP-A 相关的α-卫星 DNA 阵列的大小与双着丝粒形成前相比减小了。含有 CENP-A 的额外染色体片段经常与双着丝粒出现在同一细胞中。这些片段中的一些来自与失活着丝粒相同的α-卫星 DNA 阵列。
我们的结果表明，形成后的双着丝粒人类染色体会经历不同的命运。许多双着丝粒保留两个活性着丝粒，并通过多次细胞分裂保持稳定。其他双着丝粒则经历着丝粒失活。这个事件发生在一个广泛的时间窗口内，并且可以涉及删除标记着丝粒作为 CENP-A 维持/补充位点的染色质。
