Yeh Christina, Coyaud Étienne, Bashkurov Mikhail, van der Lelij Petra, Cheung Sally W T, Peters Jan Michael, Raught Brian, Pelletier Laurence
Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto, ON M5G 1X5, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada.
Princess Margaret Cancer Centre, University Health Network, 101 College Street, Toronto, ON M5G 1L7, Canada.
Curr Biol. 2015 Aug 31;25(17):2290-9. doi: 10.1016/j.cub.2015.07.025. Epub 2015 Aug 20.
A bipolar mitotic spindle facilitates the equal segregation of chromosomes to two daughter cells. To achieve bipolar attachment of microtubules to kinetochores of sister chromatids, chromatids must remain paired after replication. This cohesion is mediated by the conserved cohesin complex comprised of SMC1, SMC3, SCC1, and either SA1 or SA2 in humans. Because defects in spindle assembly or sister chromatid cohesion can lead to aneuploidy in daughter cells, proper regulation of these processes is essential for fidelity in chromosome segregation. In an RNAi screen for regulators of spindle assembly, we identify the deubiquitinase USP37 as a regulator of mitotic progression, centrosome integrity, and chromosome alignment. USP37 associates with cohesin and contributes to sister chromatid resolution. Cohesion defects are rescued by expression of an RNAi-resistant USP37, but not the catalytically impaired USP37(C350A) mutant. Further, USP37 associates with WAPL, a negative regulator of cohesion necessary for cohesin release in prophase, in a manner dependent on USP37's second and third ubiquitin-interacting motifs. Depletion of USP37 reduces the stability of chromatin-associated WAPL and increases the fraction of WAPL that is more heavily ubiquitylated in mitosis. Consistently, overexpression of USP37(C350A) results in increased modification of WAPL, and addition of purified USP37(WT), but not USP37(C350A), to WAPL immunoprecipitates results in a reduction of ubiquitylated products. Taken together, our results ascribe a novel function for USP37 in mitotic progression and further suggest that USP37 positively regulates the stability of chromatin-associated WAPL to facilitate sister chromatid resolution.
双极有丝分裂纺锤体有助于将染色体平均分配到两个子细胞中。为了实现微管与姐妹染色单体动粒的双极附着，染色单体在复制后必须保持配对。这种黏连由保守的黏连蛋白复合体介导，在人类中该复合体由SMC1、SMC3、SCC1以及SA1或SA2组成。由于纺锤体组装或姐妹染色单体黏连缺陷可导致子细胞出现非整倍体，因此对这些过程进行适当调控对于染色体分离的准确性至关重要。在一项针对纺锤体组装调节因子的RNA干扰筛选中，我们确定去泛素化酶USP37是有丝分裂进程、中心体完整性和染色体排列的调节因子。USP37与黏连蛋白相关，并有助于姐妹染色单体的分离。通过表达抗RNA干扰的USP37可挽救黏连缺陷，但催化受损的USP37（C350A）突变体则无法挽救。此外，USP37与WAPL相关，WAPL是前期黏连蛋白释放所必需的黏连负调节因子，其结合方式依赖于USP37的第二个和第三个泛素相互作用基序。USP37的缺失降低了与染色质相关的WAPL的稳定性，并增加了在有丝分裂中被更大量泛素化的WAPL的比例。一致的是，USP37（C350A）的过表达导致WAPL的修饰增加，并且向WAPL免疫沉淀复合物中添加纯化的USP37（野生型）而非USP37（C350A）会导致泛素化产物减少。综上所述，我们的结果赋予了USP37在有丝分裂进程中的新功能，并进一步表明USP37正向调节与染色质相关的WAPL的稳定性，以促进姐妹染色单体的分离。
