Centromeric chromatin in mitosis reorganizes into a functional structure stabilized by cohesin [CENPA-ChIP-seq]

Three-dimensional organization of mitotic chromosomes is established by cohesin and condensin complexes. In the centromere, cohesin maintains sister chromatid pairing until anaphase onset, while condensin provides elastic resistance to spindle forces. However, how condensin and cohesin structure vertebrate centromeres remains unclear. By super-resolution imaging, Capture-C analysis and polymer modeling we show that vertebrate centromeres are partitioned into two condensin-dependent subdomains during mitosis. This bipartite sub-structure is found in human, mouse and chicken centromeres, and is therefore a fundamental feature of vertebrate centromere. Super-resolution imaging and electron tomography reveal that bipartite centromeres assemble bipartite kinetochores with each subdomain capable of binding a distinct microtubule bundle. Cohesin helps to link the centromere subdomains, limiting their separation in response to spindle forces and preventing merotelic attachments. The two-domain structure described here may have implications for avoiding chromosomal instability as uncoupling of centromere subdomains is a common feature of lagging chromosomes in cancer cells. Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for Histone veriant CENP-A to identify centromere region.

有丝分裂染色体的三维组织由黏连蛋白（cohesin）复合体与凝缩蛋白（condensin）复合体共同建立。在着丝粒中，黏连蛋白可维持姐妹染色单体配对直至有丝分裂后期起始，而凝缩蛋白则为纺锤体拉力提供弹性阻力。然而，目前学界尚不清楚凝缩蛋白与黏连蛋白如何塑造脊椎动物着丝粒的结构。
本研究通过超分辨率成像、Capture-C分析与聚合物建模，证实脊椎动物着丝粒在有丝分裂过程中被划分为两个依赖于凝缩蛋白的亚结构域。这种二分性亚结构在人类、小鼠及鸡的着丝粒中均存在，因此是脊椎动物着丝粒的基本特征。
超分辨率成像与电子断层扫描技术显示，二分性着丝粒可组装出二分动粒，每个亚结构域均能结合特定的微管束。黏连蛋白可介导着丝粒亚结构域的连接，限制其在纺锤体拉力作用下的分离，并防止动粒错附（merotelic attachments）。
本文所述的双结构域结构或可为规避染色体不稳定性提供思路——着丝粒亚结构域的解耦是癌细胞中滞后染色体的常见特征。
本研究通过针对组蛋白变体CENP-A（histone variant CENP-A）的染色质免疫沉淀测序（ChIP-seq）来鉴定着丝粒区域。