Analysis of SOCS-3 promoter responses to interferon γ
Analysis of SOCS-3 promoter responses to interferon γ
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DOI：
10.1074/jbc.m308999200
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发表时间：
2004-04-02
影响因子：
4.8
通讯作者：
Bazzoni, F
中科院分区：
文献类型：
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作者：
Gatto, L;Berlato, C;Bazzoni, F
SOCS-3（细胞因子信号传导抑制因子3）是一种细胞内蛋白，其被适当的激动剂选择性地和快速地诱导，并且通过干扰Janus激酶/信号转导子和转录激活子（Jak/STAT）途径来调节免疫细胞对细胞因子的应答。在观察到干扰素γ（IFN γ）上调小鼠原代巨噬细胞、J774巨噬细胞系和胚胎成纤维细胞中SOCS-3基因和蛋白表达的基础上，我们研究了5'端SOCS-3基因的哪些序列对IFN γ应答。通过启动子缺失分析，我们确定了一个功能性IFN γ响应元件，位于转录起始上游的核苷酸-72/-64处，其存在和完整性是确保对IFN γ响应所必需的。该元件包含STAT共有结合序列（SOCS-3/STAT结合元件（SBE）），其特异性突变完全消除了对IFN γ的响应性。相反，SOCS-3启动子的其它5'区的离散缺失基本上不改变IFN γ的诱导。电迁移率变动分析表明，IFN γ促进特异性DNA结合活性的寡核苷酸探针含有SOCS-3/SBE序列。尽管IFN γ在巨噬细胞和J774细胞中触发了STAT 1和STAT 3的酪氨酸磷酸化，但只有STAT 1被适当激活，因此发现其特异性结合SOCS-3/SBE寡核苷酸探针。因此，IFN γ诱导的SOCS-3蛋白表达在STAT 3缺陷的胚胎成纤维细胞中没有受损。综上所述，这些结果表明IFN γ诱导SOCS-3依赖于SOCS-3启动子中STAT结合元件的存在，该元件被STAT 1特异性激活。
SOCS-3 ( suppressor of cytokine signaling 3) is an intracellular protein that is selectively and rapidly induced by appropriate agonists and that modulates responses of immune cells to cytokines by interfering with the Janus kinase/signal transducer and activator of transcription (Jak/STAT) pathway. On the basis of the observations that interferon gamma(IFNgamma) up-regulates SOCS-3 gene and protein expression in primary mouse macrophages, J774 macrophage cell line and embryonal fibroblasts, we investigated which sequences of the 5' SOCS-3 gene are responsive to IFNgamma. By promoter deletion analysis we identified a functional IFNgamma-responsive element, located at nucleotides - 72/ - 64 upstream from the transcription initiation, whose presence and integrity is necessary to ensure responsiveness to IFNgamma. This element contains a STAT consensus binding sequence (SOCS-3/STAT-binding element (SBE)) whose specific mutation totally abolished the responsiveness to IFNgamma. In contrast, discrete deletion of other 5' regions of the SOCS-3 promoter did not substantially modify the inducibility by IFNgamma. Electromobility shift assay analyses revealed that IFNgamma promotes specific DNA binding activities to an oligonucleotide probe containing the SOCS-3/SBE sequence. Even though IFNgamma triggered tyrosine phosphorylation of both STAT1 and STAT3 in macrophages and J774 cells, only STAT1 was appropriately activated and thus found to specifically bind to the SOCS-3/SBE oligonucleotide probe. Accordingly, IFNgamma-induced SOCS-3 protein expression was not impaired in STAT3-deficient embryonal fibroblasts. Taken together, these results demonstrate that the induction of SOCS-3 by IFNgamma depends upon the presence of a STAT-binding element in the SOCS-3 promoter that is specifically activated by STAT1.