Mahboubi Keyvan, Pober Jordan S
Interdepartmental Program in Vascular Biology and Transplantation, Boyer Center for Molecular Medicine, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
J Biol Chem. 2002 Mar 8;277(10):8012-21. doi: 10.1074/jbc.M107542200. Epub 2002 Jan 3.
We compared human endothelial cell (EC) responses to interferon-gamma (IFN gamma) and oncostatin M (OnM), cytokines that utilize Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling. Both cytokines cause phosphorylation of Tyr residue 701 and Ser residue 727 of STAT1, as shown by immunoblotting. Both activate DNA binding of STAT1 homodimers, shown by electrophoretic mobility shift assay. However, only IFN gamma increases expression of three STAT1-dependent gene products examined, namely transporter associated with antigen processing-1 (TAP1), interferon regulatory factor-1 (IRF1), and class I major histocompatibility complex (MHC) protein, as demonstrated by immunoblotting. Only IFN gamma increases TAP1 transcription assessed by reporter gene assay. OnM pretreatment or co-treatment does not inhibit IFN gamma responses. Interestingly, IFN gamma activation of STAT1 is considerably more long-lived than that produced by OnM. To determine whether duration is functionally significant, we transduced EC with a chimeric receptor containing extracellular domains of platelet-derived growth factor receptor beta and intracellular regions of gp130, the signaling subunit of the OnM receptor, mutated to prevent binding of the tyrosine phosphatase SHP-2. Addition of platelet-derived growth factor to such transduced cells produces STAT1 activation that is comparable in magnitude and duration to that caused by IFN gamma, but still fails to induce TAP1, IRF1, or class I MHC molecules. OnM also activates STAT1 but not transcription of STAT1-dependent genes in HepG2 cells. Transient transfection of HepG2 cells with a STAT-defective mouse IFN gamma receptor failed to complement the OnM STAT signal. We conclude that STAT1 activation is necessary but not sufficient for induction of transcription of IFN gamma-responsive genes. However, signals provided by IFN gamma other than STAT1 activation cannot be provided in trans to complement the response to OnM.
我们比较了人内皮细胞（EC）对干扰素-γ（IFNγ）和制瘤素M（OnM）的反应，这两种细胞因子都利用Janus激酶/信号转导子和转录激活子（JAK/STAT）信号通路。免疫印迹显示，这两种细胞因子都会导致STAT1的酪氨酸残基701和丝氨酸残基727磷酸化。电泳迁移率变动分析表明，二者均可激活STAT1同源二聚体的DNA结合。然而，免疫印迹显示，只有IFNγ能增加所检测的三种STAT1依赖性基因产物的表达，即抗原加工相关转运体-1（TAP1）、干扰素调节因子-1（IRF1）和I类主要组织相容性复合体（MHC）蛋白。报告基因检测表明，只有IFNγ能增加TAP1转录。OnM预处理或共处理不会抑制IFNγ反应。有趣的是，IFNγ对STAT1的激活作用比OnM产生的激活作用持续时间长得多。为了确定持续时间是否具有功能意义，我们用一种嵌合受体转导EC，该嵌合受体包含血小板衍生生长因子受体β的细胞外结构域和gp130的细胞内区域，gp130是OnM受体的信号亚基，经过突变以防止酪氨酸磷酸酶SHP-2结合。向这种转导细胞中添加血小板衍生生长因子会产生与IFNγ引起的STAT1激活在幅度和持续时间上相当的激活，但仍无法诱导TAP1、IRF1或I类MHC分子。OnM也能激活HepG2细胞中的STAT1，但不能激活STAT1依赖性基因的转录。用STAT缺陷型小鼠IFNγ受体瞬时转染HepG2细胞无法补充OnM的STAT信号。我们得出结论，STAT1激活对于诱导IFNγ反应性基因的转录是必要的，但不是充分的。然而，除STAT1激活外，IFNγ提供的信号不能通过转导来补充对OnM的反应。
