A proteomic analysis of ataxia telangiectasia-mutated (ATM)/ATM-Rad3-related (ATR) substrates identifies the ubiquitin-proteasome system as a regulator for DNA damage checkpoints
A proteomic analysis of ataxia telangiectasia-mutated (ATM)/ATM-Rad3-related (ATR) substrates identifies the ubiquitin-proteasome system as a regulator for DNA damage checkpoints
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DOI：
10.1074/jbc.c700079200
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发表时间：
2007-06-15
影响因子：
4.8
通讯作者：
Qin, Jun
中科院分区：
文献类型：
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作者：
Mu, Jung-Jung;Wang, Yi;Qin, Jun
ATM（共济失调毛细血管扩张突变）和ATR（ATM-Rad 3相关）是调节DNA损伤反应（DDR）的近端检查点激酶。ATM/ ATR基质的识别和表征是理解DDR的关键。很少有技术可用于鉴定蛋白激酶底物。在这里，我们使用针对已知ATM/ ATR底物的磷酸特异性抗体筛选潜在的ATM/ ATR底物。我们确定了蛋白质交叉反应磷酸特异性抗体在DNA损伤的质谱。我们验证了一个子集的候选底物磷酸化的ATM/ATR依赖的方式在体内。结合功能性检查点筛选，我们鉴定了哺乳动物DNA损伤检查点控制，特别是G（1）细胞周期检查点所需的属于泛素-蛋白酶体系统（UPS）的蛋白质，从而揭示了蛋白质泛素化作为ATM/ ATR激活检查点控制下游的重要调节机制。
ATM (ataxia telangiectasia-mutated) and ATR (ATM-Rad3-related) are proximal checkpoint kinases that regulate DNA damage response (DDR). Identification and characterization of ATM/ ATR substrates hold the keys for the understanding of DDR. Few techniques are available to identify protein kinase substrates. Here, we screened for potential ATM/ ATR substrates using phospho-specific antibodies against known ATM/ ATR substrates. We identified proteins cross-reacting to phospho-specific antibodies in response to DNA damage by mass spectrometry. We validated a subset of the candidate substrates to be phosphorylated in an ATM/ ATR-dependent manner in vivo. Combining with a functional checkpoint screen, we identified proteins that belong to the ubiquitin-proteasome system (UPS) to be required in mammalian DNA damage checkpoint control, particularly the G(1) cell cycle checkpoint, thus revealing protein ubiquitylation as an important regulatory mechanism downstream of ATM/ ATR activation for checkpoint control.