Characterization of SIS3, a novel specific inhibitor of Smad3, and its effect on transforming growth factor-β1-induced extracellular matrix expression
Characterization of SIS3, a novel specific inhibitor of Smad3, and its effect on transforming growth factor-β1-induced extracellular matrix expression
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DOI：
10.1124/mol.105.017483
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发表时间：
2006-02-01
影响因子：
3.6
通讯作者：
Tamaki, K
中科院分区：
文献类型：
--
作者：
Jinnin, M;Ihn, H;Tamaki, K
这是首次报道Smad3特异性抑制剂（SIS3）作为Smad3功能的有效和选择性抑制剂。在报告实验中，通过ALK-5组成活性形式的过表达而增加的p3TP-lux荧光素酶活性被SIS3以剂量依赖的方式消除。免疫沉淀显示，SIS3减弱了转化生长因子(TGF)- β 1诱导的Smad3磷酸化和Smad3与Smad4的相互作用。另一方面，该试剂不影响Smad2的磷酸化。随后，我们评估了SIS3抑制tgf - β 1诱导的人真皮成纤维细胞I型前胶原上调的能力。我们发现，添加SIS3通过降低转录活性来减弱tgf - β 1的作用。SIS3还能通过tgf - β 1抑制成纤维细胞向肌成纤维细胞的分化。此外，我们证明了SIS3完全降低了Smad3的组成磷酸化以及硬皮病成纤维细胞中I型胶原蛋白的上调表达。总之，我们的研究表明，SIS3是通过选择性抑制Smad3来评估tgf - β调节的细胞机制的有用工具。
This is the first report that characterizes specific inhibitor of Smad3 ( SIS3) as a potent and selective inhibitor of Smad3 function. In the reporter assay, the increased luciferase activity of p3TP-lux by the overexpression of constitutively active form of ALK-5 was abrogated by the treatment with SIS3 in a dose-dependent manner. Immunoprecipitation revealed that SIS3 attenuated the transforming growth factor ( TGF)-beta 1-induced phosphorylation of Smad3 and interaction of Smad3 with Smad4. On the other hand, this reagent did not affect the phosphorylation of Smad2. Thereafter, we evaluated the ability of SIS3 in the suppression of the TGF-beta 1-induced type I procollagen up-regulation in human dermal fibroblasts. We found that the addition of SIS3 attenuated the effects of TGF-beta 1 by reducing the transcriptional activity. SIS3 also inhibited the myofibroblast differentiation of fibroblasts by TGF-beta 1. Moreover, we demonstrated that SIS3 completely diminished the constitutive phosphorylation of Smad3 as well as the up-regulated type I collagen expression in scleroderma fibroblasts. Together, our study suggested that SIS3 is a useful tool to evaluate the TGF-beta-regulated cellular mechanisms via selective inhibition of Smad3.