The Basic Cleft of RPA70N Binds Multiple Checkpoint Proteins, Including RAD9, To Regulate ATR Signaling
The Basic Cleft of RPA70N Binds Multiple Checkpoint Proteins, Including RAD9, To Regulate ATR Signaling
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DOI：
10.1128/mcb.01079-08
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发表时间：
2008-12-15
影响因子：
5.3
通讯作者：
Cortez, David
中科院分区：
文献类型：
--
作者：
Xu, Xin;Vaithiyalingam, Sivaraja;Cortez, David
ATR激酶激活需要将ATR-ATRIP和RAD 9-HUS 1-RAD 1（9-1-1）检查点复合物募集到DNA损伤或复制应激的位点。与单链DNA结合的复制蛋白A（RPA）至少是招募这些检查点复合物的分子识别元件的一部分。我们已经发现RPA 70 N-末端寡核苷酸-寡糖折叠（OB-折叠）结构域的基本裂缝是检查点激活的关键决定因素。这种蛋白质-蛋白质相互作用表面能够结合几种检查点蛋白，包括ATRIP，RAD 9和MRE 11。RAD 9与RPA的结合由C末端RAD 9尾内的酸性肽介导，该酸性肽与ATRIP的检查点募集结构域（CRD）中的主要RPA结合表面具有序列相似性。RAD 9 CRD的突变削弱了其在DNA损伤或复制应激位点的定位，而不干扰其形成9-1-1复合物或结合ATR激活剂TopBP 1的能力。RAD 9-RPA相互作用的破坏也会损害到CHK 1的ATR信号传导，并导致对DNA损伤和复制应激的超敏反应。因此，RPA 70 N-末端OB-折叠结构域的碱性裂缝结合多种检查点蛋白，包括RAD 9，以促进ATR信号传导。
ATR kinase activation requires the recruitment of the ATR-ATRIP and RAD9-HUS1-RAD1 (9-1-1) checkpoint complexes to sites of DNA damage or replication stress. Replication protein A (RPA) bound to single-stranded DNA is at least part of the molecular recognition element that recruits these checkpoint complexes. We have found that the basic cleft of the RPA70 N-terminal oligonucleotide-oligosaccharide fold (OB-fold) domain is a key determinant of checkpoint activation. This protein-protein interaction surface is able to bind several checkpoint proteins, including ATRIP, RAD9, and MRE11. RAD9 binding to RPA is mediated by an acidic peptide within the C-terminal RAD9 tail that has sequence similarity to the primary RPA-binding surface in the checkpoint recruitment domain (CRD) of ATRIP. Mutation of the RAD9 CRD impairs its localization to sites of DNA damage or replication stress without perturbing its ability to form the 9-1-1 complex or bind the ATR activator TopBP1. Disruption of the RAD9-RPA interaction also impairs ATR signaling to CHK1 and causes hypersensitivity to both DNA damage and replication stress. Thus, the basic cleft of the RPA70 N-terminal OB-fold domain binds multiple checkpoint proteins, including RAD9, to promote ATR signaling.