Kwon Woo Jong, Kim Sun Hee, Park Yeo Ok, Cho Mong, Kang Chi Dug, Lee Gwang, An Woo Gun, Joo Woo Hong, Kim Dong Wan
Department of Microbiology, College of Natural Sciences, Changwon National University, Changwon 641-773, Korea.
Mol Cells. 2004 Oct 31;18(2):200-6.
IKKgamma is a component of the IKK complex, which regulates NF-kappaB activity. To investigate the role of IKKgamma, we expressed wild type IKKgamma containing 412 amino acids, and deletion mutants containing residues 1-312 and 101-412, using murine IKKgamma cDNA. In a co-transfection assay with a CAT reporter plasmid, NIK activated NF-kappaB-dependent gene expression approximately two fold and this expression was inhibited by co-transfection of a wild type IKKgamma expression plasmid. In binding assays IKKgamma inhibited the association of IkappaBalpha with IKKbeta and the subsequent phosphorylation of IkappaBalpha that is activated by NIK. Inhibition by IKKgamma also occurred in an assay with a dominant negative mutant of NIK but not with a C-terminal deletion mutant of IKKgamma, indicating that the C-terminal 100 amino acids of IKKgamma are important for negative regulation of NF-kappaB activation. In addition, the interaction of IKKbeta with IKKgamma was inhibited by co-transfection with a NIK expression plasmid. Our results suggest that overexpression of IKKgamma inhibits activation of NF-kappaB by NIK by competing with NIK for interaction with IKKbeta.
IKKγ是IKK复合物的一个组成部分，该复合物调节核因子κB（NF-κB）的活性。为了研究IKKγ的作用，我们使用小鼠IKKγ cDNA表达了含412个氨基酸的野生型IKKγ以及含1 - 312位和101 - 412位残基的缺失突变体。在与CAT报告质粒的共转染实验中，NIK使NF-κB依赖的基因表达增加约两倍，而野生型IKKγ表达质粒的共转染抑制了这种表达。在结合实验中，IKKγ抑制了IkappaBα与IKKβ的结合以及随后由NIK激活的IkappaBα的磷酸化。IKKγ的抑制作用在使用NIK的显性负性突变体的实验中也出现，但在IKKγ的C末端缺失突变体的实验中未出现，这表明IKKγ的C末端100个氨基酸对于NF-κB激活的负调控很重要。此外，IKKβ与IKKγ的相互作用被NIK表达质粒的共转染所抑制。我们的结果表明，IKKγ的过表达通过与NIK竞争与IKKβ的相互作用来抑制NIK对NF-κB的激活。
