Holloway S L, Glotzer M, King R W, Murray A W
Department of Physiology, University of California, San Francisco 94143-0444.
Cell. 1993 Jul 2;73(7):1393-402. doi: 10.1016/0092-8674(93)90364-v.
We have used frog egg extracts that assemble mitotic spindles to identify the event that triggers sister chromatid separation. Adding a nondegradable form of cyclin B prevents maturation-promoting factor (MPF) inactivation but does not block sister chromatid separation, showing that MPF inactivation is not needed to initiate anaphase. In contrast, adding an N-terminal fragment of cyclin, which acts as a specific competitor for cyclin degradation, produces a dose-dependent delay in MPF inactivation and sister chromatid separation. Methylated ubiquitin, which inhibits ubiquitin-mediated proteolysis, also delays sister chromatid separation, suggesting that ubiquitin-mediated proteolysis is necessary to initiate anaphase. The N-terminal cyclin fragment inhibits chromosome separation even in extracts that contain only nondegradable forms of cyclin, suggesting that proteins other than the known cyclins must be degraded to dissolve the linkage between sister chromatids.
我们使用了能组装有丝分裂纺锤体的蛙卵提取物来确定触发姐妹染色单体分离的事件。添加一种不可降解形式的细胞周期蛋白B可防止成熟促进因子（MPF）失活，但不会阻止姐妹染色单体分离，这表明启动后期并不需要MPF失活。相比之下，添加细胞周期蛋白的N端片段，它作为细胞周期蛋白降解的特异性竞争物，会在MPF失活和姐妹染色单体分离过程中产生剂量依赖性延迟。甲基化泛素可抑制泛素介导的蛋白水解作用，也会延迟姐妹染色单体分离，这表明泛素介导的蛋白水解作用是启动后期所必需的。即使在仅含有不可降解形式细胞周期蛋白的提取物中，N端细胞周期蛋白片段也会抑制染色体分离，这表明除了已知的细胞周期蛋白外，其他蛋白质也必须被降解才能溶解姐妹染色单体之间的连接。
