Sil phosphorylation in a Pin1 binding domain affects the duration of the spindle checkpoint
Sil phosphorylation in a Pin1 binding domain affects the duration of the spindle checkpoint
复制标题
DOI：
10.1128/mcb.25.15.6660-6672.2005
复制
发表时间：
2005-08-01
影响因子：
5.3
通讯作者：
Kirsch, IR
中科院分区：
文献类型：
--
作者：
Campaner, S;Kaldis, P;Kirsch, IR
SIL是一种即时早期基因，对胚胎发育至关重要，并与t细胞白血病相关易位有关。我们现在表明，在有丝分裂期间或在被微管抑制剂阻断的细胞中，Sil蛋白被过度磷酸化。Sil的细胞周期依赖性磷酸化是其与Pin1相互作用所必需的，Pin1是有丝分裂的调节因子。氨基酸567和760之间的7个（S/T）P位点的点突变减少了Sil的有丝分裂磷酸化，Pin1结合和纺锤体检查点持续时间。当磷酸化位点突变体Sil稳定表达时，在紫杉醇或诺可唑刺激的细胞中，纺锤体检查点的持续时间缩短，细胞恢复到G(2)样状态。这一事件与Cdc2/细胞周期蛋白B1复合物的激酶活性下调和Cdc2亚基上苏氨酸161的去磷酸化有关。质粒介导的RNA干扰的Sil下调限制了细胞激活纺锤体检查点的能力，并与Cdc2/cyclin B1活性的降低和Cdc2亚基上T161的磷酸化相关。这些数据表明，在纺锤体检查点阻滞期间，需要Sil的一个关键区域来介导Cdc2活性的呈现。
SIL is an immediate-early gene that is essential for embryonic development and is implicated in T-cell leukemia-associated translocations. We now show that the Sil protein is hyperphosphorylated during mitosis or in cells blocked at prometaphase by microtubule inhibitors. Cell cycle-dependent phosphorylation of Sil is required for its interaction with Pin1, a regulator of mitosis. Point mutation of the seven (S/T)P sites between amino acids 567 and 760 reduces mitotic phosphorylation of Sil, Pin1 binding, and spindle checkpoint duration. When a phosphorylation site mutant Sil is stably expressed, the duration of the spindle checkpoint is shortened in cells challenged with taxol or nocodazole, and the cells revert to a G(2)-like state. This event is associated with the downregulation of the kinase activity of the Cdc2/cyclin B1 complex and the dephosphorylation of the threonine 161 on the Cdc2 subunit. Sil downregulation by plasmid-mediated RNA interference limited the ability of cells to activate the spindle checkpoint and correlated with a reduction of Cdc2/cyclin B1 activity and phosphorylation on T161 on the Cdc2 subunit. These data suggest that a critical region of Sil is required to mediate the presentation of Cdc2 activity during spindle checkpoint arrest.