Temporal modulation of the NF-?B RelA network in response to different types of DNA damage
Temporal modulation of the NF-?B RelA network in response to different types of DNA damage
复制标题
NF-κB RelA 网络响应不同类型 DNA 损伤的时间调节
DOI：
10.1101/2020.08.11.246504
复制
发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Campbell A
中科院分区：
文献类型：
--
作者：
Campbell A
不同类型的DNA损伤可以启动磷酸化介导的信号级联反应，导致刺激特异性促凋亡或抗凋亡细胞反应。在其众多作用中，NF-κB转录因子RelA是这些DNA损伤反应途径的中心。然而，我们仍然缺乏对协调信号传导机制的理解，这些机制允许不同的DNA损伤剂通过RelA诱导不同的细胞结果。在这里，我们使用无标记的定量磷酸化蛋白质组学检查暴露的U2 OS细胞的时间效应，无论是依托泊苷（ETO）或羟基脲（HU）通过监测RelA及其蛋白结合伙伴的磷酸化状态。虽然很少的刺激特异性差异被确定在磷酸化RelA相互作用组的成分暴露后，这些DNA损伤剂，我们观察到微妙的，但显着的，在他们的磷酸化状态的变化，作为一个功能的类型和治疗的持续时间。DNA双链断裂（DSB）诱导的ETO比HU引起更快速，持续的反应，主要涉及转录，细胞分裂和典型的DSB修复的调控目标。ETO调节磷酸化位点的激酶底物预测表明，除了已知的ATM/ATR诱导作用外，CDK和ERK 1信号传导也被消除。相比之下，HU诱导的复制应激介导时间动态调节，磷酸化RelA结合配偶体在rRNA/mRNA加工和翻译起始中发挥作用，其中许多含有14-3-3ε结合基序，并且是双特异性激酶的推定底物。因此，我们的数据指出，差异调节的关键细胞过程和参与不同的信号转导通路在调节DNA损伤的RelA特异性功能。
Different types of DNA damage can initiate phosphorylation-mediated signalling cascades that result in stimulus specific pro- or anti-apoptotic cellular responses. Amongst its many roles, the NF-κB transcription factor RelA is central to these DNA damage response pathways. However, we still lack understanding of the co-ordinated signalling mechanisms that permit different DNA damaging agents to induce distinct cellular outcomes through RelA. Here, we use label-free quantitative phosphoproteomics to examine the temporal effects of exposure of U2OS cells to either etoposide (ETO) or hydroxyurea (HU) by monitoring the phosphorylation status of RelA and its protein binding partners. Although few stimulus-specific differences were identified in the constituents of phosphorylated RelA interactome after exposure to these DNA damaging agents, we observed subtle, but significant, changes in their phosphorylation states, as a function of both type and duration of treatment. The DNA double strand break (DSB)-inducing ETO invoked more rapid, sustained responses than HU, with regulated targets primarily involved in transcription, cell division and canonical DSB repair. Kinase substrate prediction of ETO-regulated phosphosites suggest abrogation of CDK and ERK1 signalling, in addition to the known induction of ATM/ATR. In contrast, HU-induced replicative stress mediated temporally dynamic regulation, with phosphorylated RelA binding partners having roles in rRNA/mRNA processing and translational initiation, many of which contained a 14-3-3ε binding motif, and were putative substrates of the dual specificity kinase CLK1. Our data thus point to differential regulation of key cellular processes and the involvement of distinct signalling pathways in modulating DNA damage-specific functions of RelA.