Chen Danying, Xu Liang-Guo, Chen Lei, Li Lixia, Zhai Zhonghe, Shu Hong-Bing
Department of Cell Biology and Genetics, College of Life Sciences, Peking University, Beijing 100871, China.
Oncogene. 2003 Jul 10;22(28):4348-55. doi: 10.1038/sj.onc.1206532.
Nuclear factor kappaB-inducing kinase (NIK) is a member of the MAP kinase kinase kinase family that was first identified as a component of the TNF-R1-induced NF-kappaB activation pathway (TNF, tumor necrosis factor; nuclear factor kappaB, NF-kappaB). Gene knockout study, however, suggests that NIK is dispensable for TNF-R1- but required for lymphotoxin-beta receptor-induced NF-kappaB activation. A NIK kinase inactive mutant is a potent inhibitor of NF-kappaB activation triggered by various stimuli, suggesting that NIK is involved in a broad range of NF-kappaB activation pathways. To unambiguously identify signaling pathways that NIK participates in, we screened antibody arrays for proteins that are associated with NIK. This effort identified ErbB4, one of the EGF/heregulin receptors, and Grb7, an adapter protein associated with ErbB4 (ErbB, epidermal growth factor receptor family protein; EGF, epidermal growth factor; Grb, growth factor receptor bound). Coimmunoprecipitation experiments demonstrated that NIK interacted with Grb7, as well as Grb10 and Grb14, but not Grb2. Domain mapping experiments indicated that the central GM domain of Grb7 was sufficient for its interaction with NIK. Coimmunoprecipitation experiments also indicated that Grb7 and NIK could be simultaneously recruited into signaling complexes of all known EGF/heregulin receptors, including EGFR, ErbB2, ErbB3, and ErbB4. In reporter gene assays, NIK could potentiate Grb7, ErbB2/ErbB4, and EGF-induced NF-kappaB activation. A NIK kinase inactive mutant could block ErbB2/ErbB4 and EGF-induced NF-kappaB activation. Moreover, EGF/heregulin receptors activated NF-kappaB in wild-type, but not NIK-/- embryonic fibroblasts. Our findings suggest that NIK is a component of the EGF/heregulin receptor signaling complexes and involved in NF-kappaB activation triggered by these receptors.
核因子κB诱导激酶（NIK）是丝裂原活化蛋白激酶激酶激酶家族的成员，最初被鉴定为TNF-R1诱导的NF-κB激活途径（TNF，肿瘤坏死因子；核因子κB，NF-κB）的一个组成部分。然而，基因敲除研究表明，NIK对于TNF-R1诱导的NF-κB激活不是必需的，但对于淋巴毒素β受体诱导的NF-κB激活是必需的。一种NIK激酶失活突变体是由各种刺激引发的NF-κB激活的有效抑制剂，这表明NIK参与了广泛的NF-κB激活途径。为了明确鉴定NIK参与的信号通路，我们筛选了与NIK相关的蛋白质的抗体阵列。这项工作鉴定出了ErbB4，它是EGF/heregulin受体之一，以及Grb7，一种与ErbB4相关的衔接蛋白（ErbB，表皮生长因子受体家族蛋白；EGF，表皮生长因子；Grb，生长因子受体结合蛋白）。免疫共沉淀实验表明，NIK与Grb7以及Grb10和Grb14相互作用，但不与Grb2相互作用。结构域定位实验表明，Grb7的中央GM结构域足以使其与NIK相互作用。免疫共沉淀实验还表明，Grb7和NIK可以同时被招募到所有已知的EGF/heregulin受体的信号复合物中，包括EGFR、ErbB2、ErbB3和ErbB4。在报告基因测定中，NIK可以增强Grb7、ErbB2/ErbB4和EGF诱导的NF-κB激活。一种NIK激酶失活突变体可以阻断ErbB2/ErbB4和EGF诱导的NF-κB激活。此外，EGF/heregulin受体在野生型而非NIK基因敲除的胚胎成纤维细胞中激活NF-κB。我们的研究结果表明，NIK是EGF/heregulin受体信号复合物的一个组成部分，并参与由这些受体触发的NF-κB激活。
