Gandhi Rita, Gillespie Peter J, Hirano Tatsuya
Cold Spring Harbor Laboratory, One Bungtown Road, P.O. Box 100, Cold Spring Harbor, New York 11724, USA.
Curr Biol. 2006 Dec 19;16(24):2406-17. doi: 10.1016/j.cub.2006.10.061. Epub 2006 Nov 16.
The linkage between duplicated chromosomes (sister chromatids) is established during S phase by the action of cohesin, a multisubunit complex conserved from yeast to humans. Most cohesin dissociates from chromosome arms when the cell enters mitotic prophase, leading to the formation of metaphase chromosomes with two cytologically discernible chromatids. This process is known as sister-chromatid resolution. Although two mitotic kinases have been implicated in this process, it remains unknown exactly how the cohesin-mediated linkage is destabilized at a mechanistic level.
The wings apart-like (Wapl) protein was originally identified as a gene product that potentially regulates heterochromatin organization in Drosophila melanogaster. We show that the human ortholog of Wapl is a cohesin-binding protein that facilitates cohesin's timely release from chromosome arms during prophase. Depletion of Wapl from HeLa cells causes transient accumulation of prometaphase-like cells with chromosomes that display poorly resolved sister chromatids with a high level of cohesin. Reduction of cohesin relieves the Wapl-depletion phenotype, and depletion of Wapl rescues premature sister separation observed in Sgo1-depleted or Esco2-depleted cells. Conversely, overexpression of Wapl causes premature separation of sister chromatids. Wapl physically associates with cohesin in HeLa-cell nuclear extracts. Remarkably, in vitro reconstitution experiments demonstrate that Wapl forms a stoichiometric, ternary complex with two regulatory subunits of cohesin, implicating its noncatalytic function in inactivating cohesin's ability to interact with chromatin.
Wapl is a new regulator of sister chromatid resolution and promotes release of cohesin from chromosomes by directly interacting with its regulatory subunits.
复制后的染色体（姐妹染色单体）之间的连接是在S期通过黏连蛋白的作用建立的，黏连蛋白是一种从酵母到人类都保守的多亚基复合体。当细胞进入有丝分裂前期时，大多数黏连蛋白从染色体臂上解离，导致形成具有两条在细胞学上可分辨的染色单体的中期染色体。这个过程被称为姐妹染色单体分离。尽管有两种有丝分裂激酶参与了这个过程，但在机制层面上，黏连蛋白介导的连接究竟是如何不稳定的仍不清楚。
类翅膀分离蛋白（Wapl）最初被鉴定为一种可能调节黑腹果蝇异染色质组织的基因产物。我们发现Wapl的人类同源物是一种黏连蛋白结合蛋白，它在前期促进黏连蛋白从染色体臂上及时释放。从HeLa细胞中耗尽Wapl会导致前中期样细胞短暂积累，其染色体显示姐妹染色单体分离不佳且黏连蛋白水平较高。黏连蛋白的减少缓解了Wapl耗尽的表型，而Wapl的耗尽挽救了在Sgo1耗尽或Esco2耗尽细胞中观察到的姐妹染色单体过早分离。相反，Wapl的过表达导致姐妹染色单体过早分离。Wapl在HeLa细胞核提取物中与黏连蛋白发生物理结合。值得注意的是，体外重组实验表明，Wapl与黏连蛋白的两个调节亚基形成化学计量的三元复合物，暗示其在使黏连蛋白与染色质相互作用的能力失活方面的非催化功能。
Wapl是姐妹染色单体分离的新调节因子，通过直接与其调节亚基相互作用促进黏连蛋白从染色体上释放。
