Timeless functions independently of the Tim-Tipin complex to promote sister chromatid cohesion in normal human fibroblasts
Timeless functions independently of the Tim-Tipin complex to promote sister chromatid cohesion in normal human fibroblasts
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DOI：
10.4161/cc.10.10.15613
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发表时间：
2011-05-15
期刊：
影响因子：
4.3
通讯作者：
Kaufmann, William K.
中科院分区：
文献类型：
--
作者：
Smith-Roe, Stephanie L.;Patel, Shivani S.;Kaufmann, William K.
Timeless-Tipin 复合物和 Claspin 是 ATR 依赖性 Chk1 激活的介导物，在 S 内检查点对停滞的 DNA 复制叉的反应中。 Tim-Tipin 和 Claspin 还可能通过复制耦合过程对各种生物体中的姐妹染色单体凝聚力 (SCC) 做出贡献。 SCC 建立的一些模型认为，粘连蛋白环和复制体之间的相互作用可能导致需要叉稳定的生理复制应激。在遗传稳定的人二倍体成纤维细胞系中研究了 Timeless、Tipin、Claspin、Chk1 和 ATR 对 SCC 的贡献。虽然 Timeless、Tipin 和 Claspin 对 UVC 诱导的 Chk1 激活表现出类似的贡献，但 siRNA 介导的 Timeless 敲除诱导姐妹染色单体分裂增加 100 倍，而敲除 Tipin、Claspin 和 ATR 的诱导效果是 4-20 倍。 Chk1 的敲低并没有显着影响 SCC。在两个独立衍生的人二倍体成纤维细胞系中获得了一致的结果，并支持这样的结论：人类细胞中的 SCC 强烈依赖于 Timeless 但独立于 Chk1。此外，消耗 Timeless 与 Tipin 时观察到的脱粘差异有 10 倍，这表明 Timeless 在 SCC 中具有独立于 Tim-Tipin 复合物的功能，尽管当 Tipin 被靶向消耗时 Timeless 的丰度会降低。更好地了解 Timeless、Tipin 和 Claspin 如何促进 SCC，将阐明这些蛋白质在 DNA 复制叉上的非检查点功能，并为 SCC 的建立模型提供信息。
The Timeless-Tipin complex and Claspin are mediators of the ATR-dependent activation of Chk1 in the intra-S checkpoint response to stalled DNA replication forks. Tim-Tipin and Claspin also contribute to sister chromatid cohesion (SCC) in various organisms, likely through a replication-coupled process. Some models of the establishment of SCC posit that interactions between cohesin rings and replisomes could result in physiological replication stress requiring fork stabilization. The contributions of Timeless, Tipin, Claspin, Chk1 and ATR to SCC were investigated in genetically stable, human diploid fibroblast cell lines. Whereas Timeless, Tipin and Claspin showed similar contributions to UVC-induced activation of Chk1, siRNA-mediated knockdown of Timeless induced a 100-fold increase in sister chromatid discohesion, whereas the inductive effects of knocking down Tipin, Claspin and ATR were 4-20-fold. Knockdown of Chk1 did not significantly affect SCC. Consistent findings were obtained in two independently derived human diploid fibroblast lines and support a conclusion that SCC in human cells is strongly dependent on Timeless but independent of Chk1. Furthermore, the 10-fold difference in discohesion observed when depleting Timeless versus Tipin indicates that Timeless has a function in SCC that is independent of the Tim-Tipin complex even though the abundance of Timeless is reduced when Tipin is targeted for depletion. A better understanding of how Timeless, Tipin and Claspin promote SCC will elucidate non-checkpoint functions of these proteins at DNA replication forks and inform models of the establishment of SCC.