A central role for CK1 in catalyzing phosphorylation of the p53 transactivation domain at serine 20 after HHV-6B viral infection
A central role for CK1 in catalyzing phosphorylation of the p53 transactivation domain at serine 20 after HHV-6B viral infection
复制标题
DOI：
10.1074/jbc.m804433200
复制
发表时间：
2008-10-17
影响因子：
4.8
通讯作者：
Hupp, Ted R.
中科院分区：
文献类型：
--
作者：
MacLaine, Nicola J.;Oster, Bodil;Hupp, Ted R.
肿瘤抑制蛋白p53可被不同的细胞应激激活，包括辐射、缺氧、I型干扰素和DNA/RNA病毒感染。p53的反激活结构域在Ser（20）上包含一个磷酸化位点，其修饰稳定了转录共激活子p300的结合，并且在小鼠转基因中其突变可诱导b细胞淋巴瘤。虽然检查点激酶CHK2与辐照后促进Ser（20）位点磷酸化有关，但在DNA病毒感染后触发这种磷酸化的酶尚不清楚。使用人类疱疹病毒6B （HHV-6B）作为一种诱导t细胞中p53丝氨酸（20）位点磷酸化的病毒，我们试图确定负责这种病毒诱导的p53修饰的激酶。使用阳离子、阴离子和染料-配体交换层析对p53 Ser（20）激酶进行分离和纯化。质谱鉴定出酪蛋白激酶1 （CK1）和牛痘相关激酶1 （VRK1）是与病毒诱导的丝氨酸（20）位点激酶活性结合的酶。免疫缺失CK1而非VRK1将激酶活性从峰值部分移除，细菌表达的CK1表现出与病毒诱导的天然CK1相当的Ser（20）位点激酶活性。CK1以对接依赖的方式修饰p53，这与其他已知的Ser（20）位点p53激酶类似。低水平的CK1抑制剂D4476选择性地抑制hhv - 6b诱导的p53的Ser（20）位点磷酸化。然而，x射线诱导的p53的Ser（20）位点磷酸化并未被D4476阻断。这些数据强调了CK1在DNA病毒感染细胞中作为p53的丝氨酸（20）位点激酶的核心作用，但也表明不同的应激可能选择性地触发不同的蛋白激酶来修饰p53的丝氨酸（20）的反活化结构域。
The tumor suppressor protein p53 is activated by distinct cellular stresses including radiation, hypoxia, type I interferon, and DNA/RNA virus infection. The transactivation domain of p53 contains a phosphorylation site at Ser(20) whose modification stabilizes the binding of the transcriptional co-activator p300 and whose mutation in murine transgenics induces B-cell lymphoma. Although the checkpoint kinase CHK2 is implicated in promoting Ser(20) site phosphorylation after irradiation, the enzyme that triggers this phosphorylation after DNA viral infection is undefined. Using human herpesvirus 6B (HHV-6B) as a virus that induces Ser(20) site phosphorylation of p53 in T-cells, we sought to identify the kinase responsible for this virus-induced p53 modification. The p53 Ser(20) kinase was fractionated and purified using cation, anion, and dye-ligand exchange chromatography. Mass spectrometry identified casein kinase 1 (CK1) and vaccinia-related kinase 1 (VRK1) as enzymes that coeluted with virus-induced Ser(20) site kinase activity. Immunodepletion of CK1 but not VRK1 removed the kinase activity from the peak fraction, and bacterially expressed CK1 exhibited Ser(20) site kinase activity equivalent to that of the virus-induced native CK1. CK1 modified p53 in a docking-dependent manner, which is similar to other known Ser(20) site p53 kinases. Low levels of the CK1 inhibitor D4476 selectively inhibited HHV-6B-induced Ser(20) site phosphorylation of p53. However, x-ray-induced Ser(20) site phosphorylation of p53 was not blocked by D4476. These data highlight a central role for CK1 as the Ser(20) site kinase for p53 in DNA virus-infected cells but also suggest that distinct stresses may selectively trigger different protein kinases to modify the transactivation domain of p53 at Ser(20).