Zhang Nianxiang, Kaur Ramandeep, Akhter Shamima, Legerski Randy J
Department of Genetics, The University of Texas MD Anderson Cancer Center, University of Texas, 1515 Holcombe Boulevard, Houston, Texas 77030, USA.
EMBO Rep. 2009 Sep;10(9):1029-35. doi: 10.1038/embor.2009.122. Epub 2009 Jul 24.
Cell division cycle 5-like protein (Cdc5L) is a core component of the putative E3 ubiquitin ligase complex containing Prp19/Pso4, Plrg1 and Spf27. This complex has been shown to have a role in pre-messenger RNA splicing from yeast to humans; however, more recent studies have described a function for this complex in the cellular response to DNA damage. Here, we show that Cdc5L interacts physically with the cell-cycle checkpoint kinase ataxia-telangiectasia and Rad3-related (ATR). Depletion of Cdc5L by RNA-mediated interference methods results in a defective S-phase cell-cycle checkpoint and cellular sensitivity in response to replication-fork blocking agents. Furthermore, we show that Cdc5L is required for the activation of downstream effectors or mediators of ATR checkpoint function such as checkpoint kinase 1 (Chk1), cell cycle checkpoint protein Rad 17 (Rad17) and Fanconi anaemia complementation group D2 protein (FancD2). In addition, we have mapped the ATR-binding region in Cdc5L and show that a deletion mutant that is unable to interact with ATR is defective in the rescue of the checkpoint deficiency in Cdc5L-depleted cells. These findings show a new function for Cdc5L in the regulation of the ATR-mediated cell-cycle checkpoint in response to genotoxic agents.
细胞分裂周期5样蛋白（Cdc5L）是包含Prp19/Pso4、Plrg1和Spf27的假定E3泛素连接酶复合物的核心组分。已证明该复合物在从酵母到人类的信使核糖核酸前体剪接中起作用；然而，最近的研究描述了该复合物在细胞对DNA损伤的反应中的功能。在此，我们表明Cdc5L与细胞周期检查点激酶共济失调毛细血管扩张症和Rad3相关蛋白（ATR）发生物理相互作用。通过RNA介导的干扰方法消耗Cdc5L会导致S期细胞周期检查点缺陷以及细胞对复制叉阻断剂的敏感性。此外，我们表明Cdc5L是激活ATR检查点功能的下游效应器或介质（如检查点激酶1（Chk1）、细胞周期检查点蛋白Rad 17（Rad17）和范可尼贫血互补组D2蛋白（FancD2））所必需的。另外，我们已绘制出Cdc5L中的ATR结合区域，并表明无法与ATR相互作用的缺失突变体在挽救Cdc5L缺失细胞中的检查点缺陷方面存在缺陷。这些发现表明Cdc5L在响应遗传毒性剂时对ATR介导的细胞周期检查点的调节中具有新功能。
