The Rad9-Hus1-Rad1 checkpoint interaction of TopBP1 with ATR
The Rad9-Hus1-Rad1 checkpoint interaction of TopBP1 with ATR
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DOI：
10.1074/jbc.m704635200
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发表时间：
2007-09-21
影响因子：
4.8
通讯作者：
Dunphy, William G.
中科院分区：
文献类型：
--
作者：
Lee, Joon;Kumagai, Akiko;Dunphy, William G.
TopBP1作为ATR-ATRIP复合物的激活剂，响应不完全复制或受损DNA的存在。该过程涉及ATR与TopBP1的ATR激活域的结合，该域位于BRCT结构域VI和VII之间。在含有检查点诱导DNA模板的爪蟾卵提取物中，TopBP1与ATR-ATRIP的结合增加。我们发现，包含BRICT重复序列I-II的TopBP1的n端区域对于这种检查点刺激的TopBP1与ATR-ATRIP的结合是必不可少的。TopBP1的BRCT I-II区域也特异性结合爪蟾卵提取物中的Rad9-Hus1-Rad1（9-1-1）复合体。这种结合通过Rad9的c端结构域发生，依赖于其Ser-373残基的磷酸化。含有缺乏BRCT I-II重复的TopBP1突变体或含有Ser-373丙氨酸取代的Rad9突变体的鸡蛋提取物在检查点调节中存在缺陷。此外，从Rad9分离的c端片段是鸡蛋提取物中检查点信号传导的有效抑制剂。这些发现表明，9-1-1复合物与TopBP1的BRCT I-II区域的相互作用是ATR- atrip结合到TopBP1的ATR激活域并随后激活ATR的必要条件。
TopBP1 serves as an activator of the ATR-ATRIP complex in response to the presence of incompletely replicated or damaged DNA. This process involves binding of ATR to the ATR-activating domain of TopBP1, which is located between BRCT domains VI and VII. TopBP1 displays increased binding to ATR-ATRIP in Xenopus egg extracts containing checkpoint-inducing DNA templates. We show that an N-terminal region of TopBP1 containing BRICT repeats I-II is essential for this checkpoint-stimulated binding of TopBP1 to ATR-ATRIP. The BRCT I-II region of TopBP1 also binds specifically to the Rad9-Hus1-Rad1 (9-1-1) complex in Xenopus egg extracts. This binding occurs via the C-terminal domain of Rad9 and depends upon phosphorylation of its Ser-373 residue. Egg extracts containing either a mutant of TopBP1 lacking the BRCT I-II repeats or a mutant of Rad9 with an alanine substitution at Ser-373 are defective in checkpoint regulation. Furthermore, an isolated C-terminal fragment from Rad9 is an effective inhibitor of checkpoint signaling in egg extracts. These findings suggest that interaction of the 9-1-1 complex with the BRCT I-II region of TopBP1 is necessary for binding of ATR-ATRIP to the ATR-activating domain of TopBP1 and the ensuing activation of ATR.