Repression of endogenous Smad7 by Ski
Repression of endogenous Smad7 by Ski
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DOI：
10.1074/jbc.m404961200
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发表时间：
2004-07-02
影响因子：
4.8
通讯作者：
Liu, F
中科院分区：
文献类型：
--
作者：
Denissova, NG;Liu, F
Ski蛋白被认为是Smad 4的辅阻遏物，使转化生长因子-β（TGF-β）应答启动子维持在被阻遏的基础水平。然而，迄今为止还没有报道它确实作用于天然启动子。我们以前克隆了人Smad 7启动子，并表明它含有TGF-β诱导所必需的8个碱基对回文Smad结合元件（SBE）。在这份报告中，我们的特点是负调控Smad 7启动子基础活性滑雪。我们发现，滑雪抑制Smad 7启动子的基础活性在SBE依赖的方式。SBE的突变消除了Ski对Smad 7启动子的抑制作用。此外，SBE的突变增加了Smad 7启动子的基础活性。使用染色质免疫沉淀试验，我们进一步表明，滑雪连同Smad 4结合到内源性Smad 7启动子。最后，我们表明，RNAi敲除Ski增加了Smad 7报告基因的活性，在瞬时转染试验以及提高内源性水平的Smad 7 mRNA。综上所述，我们的研究结果提供了第一个证据，Ski确实是Smad 4的辅阻遏物，Smad 4可以抑制基础状态下的天然TGF-β反应基因。
The Ski protein has been proposed to serve as a corepressor for Smad4 to maintain a transforming growth factor-beta (TGF-beta)-responsive promoter at a repressed, basal level. However, there have been no reports so far that it indeed acts on a natural promoter. We have previously cloned the human Smad7 promoter and shown that it contains the 8-base pair palindromic Smad-binding element (SBE) necessary for TGF-beta induction. In this report, we have characterized the negative regulation of Smad7 promoter basal activity by Ski. We show that Ski inhibits the Smad7 promoter basal activity in a SBE-dependent manner. Mutation of the SBE abrogates the inhibitory effect of Ski on the Smad7 promoter. Moreover, mutation of the SBE increases the Smad7 promoter basal activity. Using the chromatin immunoprecipitation assay, we further show that Ski together with Smad4 binds to the endogenous Smad7 promoter. Finally, we show that RNAi knockdown of Ski increases Smad7 reporter gene activity in transient transfection assays as well as elevating the endogenous level of Smad7 mRNA. Taken together, our results provide the first evidence that Ski is indeed a corepressor for Smad4, which can inhibit a natural TGF-beta responsive gene at the basal state.