Direct phosphorylation of NF-κB1 p105 by the IκB kinase complex on serine 927 is essential for signal-induced p105 proteolysis
Direct phosphorylation of NF-κB1 p105 by the IκB kinase complex on serine 927 is essential for signal-induced p105 proteolysis
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DOI：
10.1074/jbc.m101754200
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发表时间：
2001-06-22
影响因子：
4.8
通讯作者：
Ley, SC
中科院分区：
文献类型：
--
作者：
Salmerón, A;Janzen, J;Ley, SC
NF-κ B1的p105前体蛋白作为NF-κ B抑制蛋白，在未刺激细胞的细胞质中保留相关的Rel亚基。肿瘤坏死因子α（TNF α）和白细胞介素-1 α（IL-1 α）刺激p105降解，释放相关的Rel亚单位易位到细胞核中。通过使用敲除胚胎成纤维细胞，首先确定I κ B激酶（IKK）复合物对于这些促炎细胞因子有效地触发p105降解是必需的。p105 PEST结构域包含一个基序（Asp-Ser（927）Gly-Val Glu-Thr），与I κ B α中的IKK靶序列相关，在人、小鼠、大鼠和鸡p105之间是保守的。分析一组人p105突变体，其中丝氨酸/苏氨酸残基内和邻近该基序分别改变为丙氨酸，建立了只有丝氨酸927是必不可少的p105蛋白水解引发IKK 2过表达。TNF α和IL-1 α刺激p105降解也需要该残基。通过使用特异性抗磷酸肽抗体，证实IKK 2过表达诱导共转染的p105的丝氨酸927磷酸化，并且在TNF α或IL-1 α刺激后内源性p105也在该残基上快速磷酸化。用纯化的蛋白质进行的体外激酶测定证明IKK 1和IKK 2都可以直接磷酸化丝氨酸927上的p105。总之，这些实验表明，IKK复合物调节信号诱导的NF-κ B1 p105蛋白水解的直接磷酸化丝氨酸927在其PEST结构域。
The p105 precursor protein of NF-kappa B1 acts as an NF-KB inhibitory protein, retaining associated Rel subunits in the cytoplasm of unstimulated cells. Tumor necrosis factor alpha (TNF alpha) and interleukin-1 alpha (IL-1 alpha) stimulate p105 degradation, releasing associated Rel subunits to translocate into the nucleus. By using knockout embryonic fibroblasts, it was first established that the I kappaB kinase (IKK) complex is essential for these pro-inflammatory cytokines to trigger efficiently p105 degradation. The p105 PEST domain contains a motif (Asp-Ser(927) Gly-Val Glu-Thr), related to the IKK target sequence in I kappaB alpha, which is conserved between human, mouse, rat, and chicken p105. Analysis of a panel of human p105 mutants in which serine/threonine residues within and adjacent to this motif were individually changed to alanine established that only serine 927 is essential for p105 proteolysis triggered by IKK2 overexpression. This residue is also required for TNF alpha and IL-1 alpha to stimulate p105 degradation. By using a specific anti-phosphopeptide antibody, it was confirmed that IKK2 overexpression induces serine 927 phosphorylation of co transfected p105 and that endogenous p105 is also rapidly phosphorylated on this residue after TNF alpha or IL-1 alpha stimulation. lie vitro kinase assays with purified proteins demonstrated that both IKK1 and IKK2 can directly phosphorylate p105 on serine 927. Together these experiments indicate that the IKK complex regulates the signal-induced proteolysis of NF-kappa B1 p105 by direct phosphorylation of serine 927 in its PEST domain.