Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL, 32306-4300, USA.
Yerkes National Primate Research Center, Emory Vaccine Center, 954 Gatewood Rd NE, Atlanta, GA, 30329, USA.
Sci Rep. 2017 Sep 19;7(1):11880. doi: 10.1038/s41598-017-12329-z.
The interaction between chromosomes and spindle microtubules is essential for chromosome segregation. The kinetochore complex mediates this interaction. Previous studies indicate that the stability of kinetochore attachment is regulated by Aurora B/Ipl1 kinase and this regulation is conserved from yeast to mammalian cells. In budding yeast Saccharomyces cerevisiae, the ten-subunit Dam1/DASH complex bridges the interaction between kinetochores and microtubules, and some in vitro evidence indicates that the phosphorylation of Dam1 protein by Ipl1 kinase destabilizes this interaction. However, it is not clear if Dam1 phosphorylation is sufficient to regulate the stability of kinetochore attachment in vivo. Also, the significance of this regulation in response to chromosome detachment has not been fully investigated. Here we report that phospho-deficient dam1-3A mutants show stabilized kinetochore-microtubule attachment in vivo. This significantly delays the establishment of chromosome bipolar attachment after the disruption of kinetochore-microtubule interaction by a microtubule depolymerizing drug nocodazole. Moreover, dam1-3A cells show dramatic chromosome mis-segregation after treatment with nocodazole, presumably due to the combination of compromised bipolar attachment and premature spindle assembly checkpoint silencing in the mutant cells. Therefore, the regulation of Dam1 phosphorylation imposed by Ipl1 kinase is critical for faithful chromosome segregation.
染色体与纺锤体微管之间的相互作用对于染色体分离至关重要。着丝粒复合物介导这种相互作用。先前的研究表明，动粒附着的稳定性受 Aurora B/Ipl1 激酶调节，这种调节在从酵母到哺乳动物细胞中是保守的。在芽殖酵母酿酒酵母中，十个亚基的 Dam1/DASH 复合物桥接着丝粒和微管之间的相互作用，一些体外证据表明，Ipl1 激酶对 Dam1 蛋白的磷酸化使这种相互作用不稳定。然而，Dam1 磷酸化是否足以调节体内动粒附着的稳定性尚不清楚。此外，这种调节对染色体脱离的反应的意义尚未得到充分研究。在这里，我们报告说，磷酸化缺陷的 dam1-3A 突变体在体内显示出稳定的动粒-微管附着。这显著延迟了微管去聚合药物 nocodazole 破坏动粒-微管相互作用后染色体双极附着的建立。此外，dam1-3A 细胞在用 nocodazole 处理后表现出明显的染色体错误分离，这可能是由于突变细胞中受损的双极附着和过早的纺锤体组装检查点沉默的组合所致。因此，Ipl1 激酶对 Dam1 磷酸化的调节对于忠实的染色体分离至关重要。
