Abstract：
NIK (NF-κB inducing kinase) belongs to the mitogen-activated protein kinase family, which activates NF-κB and plays a vital role in immunology, inflammation, apoptosis, and a series of pathological responses. In NF-κB noncanonical pathway, NIK and IKKα have been often studied in mammals and zebrafish. However, few have explored the relationship between NIK and other subunits of the IKK complex. As a classic kinase in the NF-κB canonical pathway, IKKβ has never been researched with NIK in fish. In this paper, the full-length cDNA sequence of grass carp (Ctenopharyngodon idella) NIK (CiNIK) was first cloned and identified. The expression level of CiNIK in grass carp cells was increased under GCRV stimuli. Under the stimulation of GCRV, poly (I:C), and LPS, the expression of NIK in various tissues of grass carp was also increased. This suggests that CiNIK responds to viral stimuli. To study the relationship between CiNIK and CiIKKβ, we co-transfected CiNIK-FLAG and CiIKKB-GFP into grass carp cells in coimmunoprecipitation and immunofluorescence experiments. The results revealed that CiNIK interacts with CiIKKβ. Besides, the degree of autophosphorylation of CiNIK was enhanced under poly (I:C) stimulation. CiIKKβ was phosphorylated by CiNIK and then activated the activity of p65. The activity change of p65 indicates that NF-κB downstream inflammatory genes will be functioning. CiNIK or CiIKKβ up-regulated the expression of IL-8. It got higher when CiNIK and CiIKKβ coexisted. This paper revealed that NF-κB canonical pathway and noncanonical pathway are not completely separated in generating benefits.
摘要：
NIK(NF-κB诱导激酶)属于丝裂原活化蛋白激酶家族,激活NF-κB并在免疫学中起着至关重要的作用，炎症,凋亡，和一系列的病理反应。在NF-κB非经典通路中，NIK和IKKα经常在哺乳动物和斑马鱼中进行研究。然而,很少有人探讨NIK与IKK复合体其他亚基之间的关系。作为NF-κB经典途径中的经典激酶，IKKβ从未在鱼类中与NIK一起研究过。在本文中,首次克隆并鉴定了草鱼（Ctenpharyngodonidella）NIK（CiNIK）的全长cDNA序列。在GCRV刺激下，草鱼细胞中CiNIK的表达水平升高。在GCRV的刺激下,聚（I：C），和LPS，NIK在草鱼各组织中的表达也增加。这表明CiNIK对病毒刺激有反应。为了研究CiNIK与CiIKKβ的关系，在共免疫沉淀和免疫荧光实验中，我们将CiNIK-FLAG和CiIKKB-GFP共转染到草鱼细胞中。结果显示CiNIK与CiIKKβ相互作用。此外,在poly（I：C）刺激下，CiNIK的自磷酸化程度增强。CiIKKβ被CiNIK磷酸化，然后激活p65的活性。p65的活性变化表明NF-κB下游炎症基因将起作用。CiNIK或CiIKKβ上调IL-8的表达。当CiNIK和CiIKKβ共存时，它变得更高。本文揭示了NF-κB经典途径和非经典途径在产生益处方面并没有完全分开。
