Tenascin-C upregulation by transforming growth factor-β in human dermal broblasts involves Smad3, Sp1, and Ets1
Tenascin-C upregulation by transforming growth factor-β in human dermal broblasts involves Smad3, Sp1, and Ets1
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DOI：
10.1038/sj.onc.1207064
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发表时间：
2004-03-04
期刊：
影响因子：
8
通讯作者：
Tamaki, K
中科院分区：
文献类型：
--
作者：
Jinnin, M;Ihn, H;Tamaki, K
在培养的人真皮成纤维细胞中，转化生长因子（TGF）-β诱导腱生蛋白-C（TN-C）的mRNA表达。该过程的分子机制目前尚不清楚。在这项研究中，我们进行了连续50缺失和瞬时转染分析，以确定。在TN-C启动子中的一个区域，介导对TGF-β的诱导反应。该区域含有转录调节因子Smad家族的非典型核苷酸识别元件。DNA亲和沉淀分析表明，Smad 2/Smad 3结合到这个网站在一个短暂的和特定的方式。Smad 3或Smad 4的过表达激活了TN-C启动子的活性，并超诱导了TGF-β刺激的TN-C启动子活性。此外，同时共转染Smad 3和Smad 4以协同方式激活TN-C启动子活性。TN-C启动子中Smad结合位点、Ets结合位点或Sp1/3结合位点的突变废除了TGF-β/Smad诱导型启动子活性。免疫沉淀分析表明，Smad 3，Sp1和Ets 1形成一个转录活性复合物。此外，Smads和CBP/p300在TGF-β信号转导中的相互作用得到证实。这些研究结果表明，在TN-C启动子的近端区域存在一种新的功能性结合元件，介导对TGF-β的反应，涉及Smad 3/4，Sp1，Ets 1和CBP/p300。
In cultured human dermal fibroblasts, transforming growth factor ( TGF)-beta induced the mRNA expression of tenascin-C (TN-C). The molecular mechanism(s) underlying this process is not presently understood. In this study, we performed serial 50 deletion and a transient transfection analysis to de. ne a region in the TN-C promoter mediating the inducible responsiveness to TGF-beta. This region contains an atypical nucleotide recognition element for the Smad family of transcriptional regulators. A DNA affinity precipitation assay revealed that Smad2/ Smad3 bound to this site in a transient and specific manner. Overexpression of Smad3 or Smad4 activated the TN-C promoter activity and superinduced the TN-C promoter activity stimulated by TGF-beta. Moreover, simultaneous cotransfection of Smad3 and Smad4 activated the TN-C promoter activity in a synergistic manner. Mutation of the Smad-binding sites, the Ets-binding sites, or Sp1/3-binding sites in the TN-C promoter abrogated the TGF-beta/Smad-inducible promoter activity. Immunoprecipitation analysis revealed that Smad3, Sp1, and Ets1 form a transcriptionally active complex. Furthermore, the interaction between Smads and CBP/p300 in TGF-beta signaling was confirmed. These findings demonstrate the existence of a novel, functional binding element in the proximal region of the TN-C promoter mediating responsiveness to TGF-beta involving Smad3/4, Sp1, Ets1, and CBP/p300.