Lai J H, Horvath G, Subleski J, Bruder J, Ghosh P, Tan T H
Department of Microbiology and Immunology, Baylor College of Medicine, Houston, Texas 77030, USA.
Mol Cell Biol. 1995 Aug;15(8):4260-71. doi: 10.1128/MCB.15.8.4260.
T-cell activation requires two different signals. The T-cell receptor's recognition of a specific antigen on antigen-presenting cells provides one, and the second signal comes from costimulatory molecules such as CD28. In contrast, T cells that are stimulated with antigen in the absence of the CD28 costimulatory signal can become anergic (nonresponsive). The CD28 response element (CD28RE) has been identified as the DNA element mediating interleukin 2 (IL-2) gene activation by CD28 costimulation. Our previous work demonstrates that the Rel/NF-kappa B family proteins c-Rel, RelA (p65), and NFKB1 (p50) are involved in the complex that binds to the CD28RE. We also showed that c-Rel, but not NFKB1 (p50), can bind to the CD28RE and activate CD28RE-driven transcription in cotransfection assays. However, the role of RelA (p65) in CD28 signaling has not yet been addressed. We provide evidence that RelA (p65) itself bound directly to the CD28RE of the IL-2 promoter and other lymphokine promoters. In addition, RelA (p65) was a potent transcriptional activator of the CD28RE in vivo. We show that a RelA (p65)-c-Rel heterodimer bound to the CD28RE and synergistically activated the CD28RE enhancer activity. We also demonstrate that activated Raf-1 kinase synergized with RelA (p65) in activating the CD28RE enhancer activity. Interestingly, a soluble anti-CD28 monoclonal antibody alone, in the absence of other stimuli, also synergized with RelA (p65) in activating the CD28RE. Furthermore, we show that RelA (p65) activated expression of the wild-type IL-2 promoter but not the CD28RE-mutated IL-2 promoter. A combination of RelA (p65) and NFKB1 (p50) also activated the IL-2 promoter through the CD28RE site. These results demonstrate the functional regulation of the CD28RE, within the IL-2 promoter, by Rel/NF-kappa B transcription factors.
T细胞活化需要两种不同的信号。T细胞受体识别抗原呈递细胞上的特定抗原提供一种信号，第二种信号来自共刺激分子，如CD28。相比之下，在没有CD28共刺激信号的情况下用抗原刺激的T细胞会变得无反应性（无应答）。CD28反应元件（CD28RE）已被确定为介导CD28共刺激激活白细胞介素2（IL-2）基因的DNA元件。我们之前的工作表明，Rel/NF-κB家族蛋白c-Rel、RelA（p65）和NFKB1（p50）参与了与CD28RE结合的复合物。我们还表明，在共转染实验中，c-Rel而非NFKB1（p50）能够结合CD28RE并激活由CD28RE驱动的转录。然而，RelA（p65）在CD28信号传导中的作用尚未得到研究。我们提供的证据表明，RelA（p65）自身直接结合到IL-2启动子和其他淋巴因子启动子的CD28RE上。此外，RelA（p65）在体内是CD28RE的有效转录激活因子。我们发现RelA（p65）-c-Rel异二聚体结合到CD28RE上，并协同激活CD28RE增强子活性。我们还证明活化的Raf-1激酶在激活CD28RE增强子活性方面与RelA（p65）协同作用。有趣的是，单独的可溶性抗CD28单克隆抗体在没有其他刺激的情况下，在激活CD28RE方面也与RelA（p65）协同作用。此外，我们表明RelA（p65）激活野生型IL-2启动子的表达，但不激活CD28RE突变的IL-2启动子的表达。RelA（p65）和NFKB1（p50）的组合也通过CD28RE位点激活IL-2启动子。这些结果证明了Rel/NF-κB转录因子对IL-2启动子内CD28RE的功能调节。
