Cussiol José R, Jablonowski Carolyn M, Yimit Askar, Brown Grant W, Smolka Marcus B
Department of Molecular Biology and Genetics, Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, NY, USA.
Donnelly Centre and Department of Biochemistry, University of Toronto, Toronto, ON, Canada.
EMBO J. 2015 Jun 12;34(12):1704-17. doi: 10.15252/embj.201490834. Epub 2015 Apr 20.
In response to DNA damage, checkpoint signalling protects genome integrity at the cost of repressing cell cycle progression and DNA replication. Mechanisms for checkpoint down-regulation are therefore necessary for proper cellular proliferation. We recently uncovered a phosphatase-independent mechanism for dampening checkpoint signalling, where the checkpoint adaptor Rad9 is counteracted by the repair scaffolds Slx4-Rtt107. Here, we establish the molecular requirements for this new mode of checkpoint regulation. We engineered a minimal multi-BRCT-domain (MBD) module that recapitulates the action of Slx4-Rtt107 in checkpoint down-regulation. MBD mimics the damage-induced Dpb11-Slx4-Rtt107 complex by synergistically interacting with lesion-specific phospho-sites in Ddc1 and H2A. We propose that efficient recruitment of Dpb11-Slx4-Rtt107 or MBD via a cooperative 'two-site-docking' mechanism displaces Rad9. MBD also interacts with the Mus81 nuclease following checkpoint dampening, suggesting a spatio-temporal coordination of checkpoint signalling and DNA repair via a combinatorial mode of BRCT-domains interactions.
作为对DNA损伤的反应，检查点信号传导以抑制细胞周期进程和DNA复制为代价来保护基因组完整性。因此，检查点下调机制对于细胞的正常增殖是必要的。我们最近发现了一种不依赖磷酸酶的减弱检查点信号传导的机制，其中检查点衔接蛋白Rad9被修复支架Slx4-Rtt107所拮抗。在此，我们确定了这种新的检查点调节模式的分子要求。我们设计了一个最小化的多BRCT结构域（MBD）模块，该模块概括了Slx4-Rtt107在检查点下调中的作用。MBD通过与Ddc1和H2A中的损伤特异性磷酸化位点协同相互作用，模拟了损伤诱导的Dpb11-Slx4-Rtt107复合物。我们提出，通过协同的“双位点对接”机制有效招募Dpb11-Slx4-Rtt107或MBD会取代Rad9。在检查点减弱后，MBD还与Mus81核酸酶相互作用，这表明通过BRCT结构域相互作用的组合模式实现了检查点信号传导和DNA修复的时空协调。
