Devajyothi C, Kalvakolanu I, Babcock G T, Vasavada H A, Howe P H, Ransohoff R M
Research Institute, Cleveland Clinic Foundation, Ohio 44195.
J Biol Chem. 1993 Sep 5;268(25):18794-800.
To address mechanisms by which cytokines inhibit interferon-gamma (IFN gamma)-induced gene expression in astrocytic cells, we have been studying effects of type beta 1 transforming growth factor (TGF beta 1) and interferon-beta (IFN beta) on IFN gamma-induced expression of the well-characterized human major histocompatibility complex (MHC) class II gene DRA. This investigation was motivated by the observations that IFN gamma-induced expression of MHC class II antigen expression on astrocytic cells can be blocked in a tissue-specific fashion by several cytokines and neurotransmitters in tissue culture and that astrocyte expression of MHC class II is severely restricted in vivo. We previously showed that IFN beta inhibited IFN gamma-induced DRA expression at the transcriptional level. This inhibition was not global, since IFN gamma-induction of intercellular adhesion molecule-1 was not affected. Here, TGF beta 1-mediated inhibition of DRA is shown to exhibit similar characteristics. To address the mechanism of this inhibition, sequence requirements for IFN beta and TGF beta 1 to suppress IFN gamma-induced transcription of DRA were determined. A 135-base pair DRA sequence element containing the IFN gamma-responsive region and transcriptional start site was sufficient to direct IFN beta- or TGF beta 1-mediated suppression of a reporter gene. These experiments suggested that either IFN beta or TGF beta 1 could repress IFN gamma-induced DRA transcription directly, without requiring a cis-element extrinsic to the IFN gamma-inducible DRA sequences. Consistently similar effects of IFN beta and TGF beta 1 observed in these experiments prompted comparison of other gene regulatory effects of the two cytokines. TGF beta 1, unlike IFN beta, induced gene expression directed by upstream elements of the gene encoding plasminogen activator inhibitor type-1. IFN beta, unlike TGF beta 1, enhanced levels of 2'-5'-oligoadenylate synthetase activity. Additionally, direct assay of TGF beta 1 and monoclonal anti-TGF beta antibody blocking experiments were used to determine that cells treated with IFN beta did not produce increased amounts of active or latent TGF beta. These data argued that IFN beta and TGF beta 1 utilized distinct pathways to mediate the inhibition of IFN gamma-induced DRA transcription.
为了探究细胞因子抑制星形胶质细胞中干扰素-γ（IFNγ）诱导的基因表达的机制，我们一直在研究β1型转化生长因子（TGFβ1）和干扰素-β（IFNβ）对IFNγ诱导的、已被充分表征的人类主要组织相容性复合体（MHC）II类基因DRA表达的影响。这项研究的动机源于以下观察结果：在组织培养中，几种细胞因子和神经递质可以以组织特异性方式阻断IFNγ诱导的星形胶质细胞上MHC II类抗原的表达，并且MHC II类在星形胶质细胞中的表达在体内受到严重限制。我们之前表明，IFNβ在转录水平上抑制IFNγ诱导的DRA表达。这种抑制并非全局性的，因为细胞间黏附分子-1的IFNγ诱导不受影响。在此，TGFβ1介导的DRA抑制表现出类似的特征。为了探究这种抑制的机制，确定了IFNβ和TGFβ1抑制IFNγ诱导的DRA转录的序列要求。一个包含IFNγ反应区域和转录起始位点的135个碱基对的DRA序列元件足以指导IFNβ或TGFβ1介导的报告基因抑制。这些实验表明，IFNβ或TGFβ1可以直接抑制IFNγ诱导的DRA转录，而无需IFNγ诱导的DRA序列之外的顺式元件。在这些实验中观察到的IFNβ和TGFβ1始终相似的作用促使我们比较这两种细胞因子的其他基因调控作用。与IFNβ不同，TGFβ1诱导由纤溶酶原激活物抑制剂1型编码基因的上游元件指导的基因表达。与TGFβ1不同，IFNβ提高了2'-5'-寡腺苷酸合成酶的活性水平。此外，通过对TGFβ1的直接检测和单克隆抗TGFβ抗体阻断实验来确定用IFNβ处理的细胞不会产生增加量的活性或潜伏性TGFβ。这些数据表明，IFNβ和TGFβ1利用不同的途径来介导对IFNγ诱导的DRA转录的抑制。
