Newton T R, Patel N M, Bhat-Nakshatri P, Stauss C R, Goulet R J, Nakshatri H
Department of Surgery, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
J Biol Chem. 1999 Jun 25;274(26):18827-35. doi: 10.1074/jbc.274.26.18827.
The transcription factor NF-kappaB regulates the expression of genes involved in cancer cell invasion, metastasis, angiogenesis, and resistance to chemotherapy. In normal cells NF-kappaB is maintained in the cytoplasm by protein-protein interaction with inhibitor IkappaBs. In contrast, in cancer cells a substantial amount of NF-kappaB is in the nucleus and constitutively activates target genes. To understand the mechanisms of constitutive NF-kappaB activation, we have analyzed the function of IkappaBalpha and IkappaBbeta in breast cancer cells. In most cases, constitutive NF-kappaB DNA binding correlated with reduced levels of either IkappaBalpha or IkappaBbeta isoforms. Overexpression of IkappaBalpha but not IkappaBbeta1 resulted in reduced constitutive DNA binding of NF-kappaB in MDA-MB-231 cells. Unexpectedly, IkappaBbeta1 overexpression moderately increased 12-O-tetradecanoylphorbol-13-acetate- and interleukin-1-inducible NF-kappaB DNA binding. 12-O-Tetradecanoylphorbol-13-acetate- and interleukin-1-induced transactivation by NF-kappaB, however, was lower in IkappaBbeta1-overexpressing cells. Mutants of IkappaBbeta1 lacking the C-terminal casein kinase II phosphorylation sites, which form a stable complex with DNA bound NF-kappaB without inhibiting its transactivation in other cell types, repressed the transactivation by NF-kappaB in MDA-MB-231 cells. Consistent with the results of transient transfections, the expression of urokinase plasminogen activator, an NF-kappaB target gene, was reduced in IkappaBbeta1-overexpressing cells. These results suggest that depending on the cell type, IkappaBbeta1 represses the expression of NF-kappaB-regulated genes by inhibiting either DNA binding or transactivation function of NF-kappaB.
转录因子核因子-κB（NF-κB）调控参与癌细胞侵袭、转移、血管生成及化疗耐药相关基因的表达。在正常细胞中，NF-κB通过与抑制蛋白IκB发生蛋白质-蛋白质相互作用而维持在细胞质中。相反，在癌细胞中，大量的NF-κB存在于细胞核中并持续激活靶基因。为了解NF-κB持续性激活的机制，我们分析了IκBα和IκBβ在乳腺癌细胞中的功能。在大多数情况下，NF-κB的持续性DNA结合与IκBα或IκBβ亚型水平降低相关。IκBα的过表达而非IκBβ1的过表达导致MDA-MB-231细胞中NF-κB的持续性DNA结合减少。出乎意料的是，IκBβ1的过表达适度增加了12-O-十四酰佛波醇-13-乙酸酯（TPA）和白细胞介素-1诱导的NF-κB DNA结合。然而，在IκBβ1过表达的细胞中，TPA和白细胞介素-1诱导的NF-κB反式激活作用较低。缺乏C末端酪蛋白激酶II磷酸化位点的IκBβ1突变体，在其他细胞类型中可与结合DNA的NF-κB形成稳定复合物而不抑制其反式激活作用，但在MDA-MB-231细胞中却抑制了NF-κB的反式激活作用。与瞬时转染结果一致，NF-κB靶基因尿激酶型纤溶酶原激活剂在IκBβ1过表达细胞中的表达降低。这些结果表明，取决于细胞类型，IκBβ1通过抑制NF-κB的DNA结合或反式激活功能来抑制NF-κB调控基因 的表达。
