Rad4（TopBP1）通过磷酸化依赖性组装和协调 DNA 损伤检查点装置。
Phosphorylation-dependent assembly and coordination of the DNA damage checkpoint apparatus by Rad4(TopBP1).
机构信息
National Institute of Biological Sciences, 7 Science Park Road, ZGC Life Science Park, Beijing 102206, China.
Cancer Research UK DNA Repair Enzymes Group, Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Falmer, BN1 9RQ, UK.
出版信息
Mol Cell. 2013 Sep 26;51(6):723-736. doi: 10.1016/j.molcel.2013.08.030.
The BRCT-domain protein Rad4(TopBP1) facilitates activation of the DNA damage checkpoint in Schizosaccharomyces pombe by physically coupling the Rad9-Rad1-Hus1 clamp, the Rad3(ATR) -Rad26(ATRIP) kinase complex, and the Crb2(53BP1) mediator. We have now determined crystal structures of the BRCT repeats of Rad4(TopBP1), revealing a distinctive domain architecture, and characterized their phosphorylation-dependent interactions with Rad9 and Crb2(53BP1). We identify a cluster of phosphorylation sites in the N-terminal region of Crb2(53BP1) that mediate interaction with Rad4(TopBP1) and reveal a hierarchical phosphorylation mechanism in which phosphorylation of Crb2(53BP1) residues Thr215 and Thr235 promotes phosphorylation of the noncanonical Thr187 site by scaffolding cyclin-dependent kinase (CDK) recruitment. Finally, we show that the simultaneous interaction of a single Rad4(TopBP1) molecule with both Thr187 phosphorylation sites in a Crb2(53BP1) dimer is essential for establishing the DNA damage checkpoint.
BRCT 结构域蛋白 Rad4（TopBP1）通过物理连接 Rad9-Rad1-Hus1 夹、Rad3（ATR）-Rad26（ATRIP）激酶复合物和 Crb2（53BP1）介体，促进裂殖酵母有丝分裂检查点的激活。我们现在已经确定了 Rad4（TopBP1）BRCT 重复序列的晶体结构，揭示了其独特的结构域架构，并对其与 Rad9 和 Crb2（53BP1）的磷酸化依赖性相互作用进行了表征。我们鉴定了 Crb2（53BP1）N 端区域的一组磷酸化位点，这些位点介导与 Rad4（TopBP1）的相互作用，并揭示了一个分层的磷酸化机制，其中 Crb2（53BP1）残基 Thr215 和 Thr235 的磷酸化促进了非典型 Thr187 位点的磷酸化，通过支架环化酶依赖性激酶（CDK）募集。最后，我们表明，单个 Rad4（TopBP1）分子与 Crb2（53BP1）二聚体中两个 Thr187 磷酸化位点的同时相互作用对于建立 DNA 损伤检查点至关重要。
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