Transforming growth factor β regulates clusterin gene expression via modulation of transcription factor c-Fos
Transforming growth factor β regulates clusterin gene expression via modulation of transcription factor c-Fos
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DOI：
10.1046/j.1432-1327.1999.00533.x
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发表时间：
1999-07-01
期刊：
影响因子：
--
通讯作者：
Howe, PH
中科院分区：
文献类型：
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作者：
Jin, G;Howe, PH
转化生长因子-β(TGF-β)通过一个共同的AP-1结合位点在多种细胞中诱导糖蛋白Clusterin的基因表达。在这里，我们通过超位移分析证明，JunB、Jund、Fra1、Fra2和c-Fos与AP-1结合，但转化生长因子β预先处理细胞后，c-Fos结合显著减少，表明c-Fos可能在Clusterin基因表达中发挥负调控作用。将人c-Fos表达载体与Clusterin启动子/报告载体或人工构建的转化生长因子β诱导的报告载体3TPLux联合应用于水貂肺上皮细胞的瞬时共转染实验表明，c-Fos确实对转化生长因子β诱导的启动子反式激活有抑制作用。进一步，我们证明在稳定的c-Fos过表达细胞系中，转化生长因子β对内源性Clusterin mRNA的诱导以及Clusterin启动子的反式激活被阻断。与c-Fos缺失构建体共转染发现，抑制Clusterin和3TPLux反式激活所需的C末端区域，包括同源框2基序和极端C末端丝氨酸磷酸化位点(Ser362和Ser374)。经转化生长因子β处理的CCL64细胞可诱导c-Fos基因的表达，但对总c-Fos蛋白水平无明显影响。结果表明，c-Fos抑制了Clusterin基因的表达，在没有转化生长因子β的情况下维持了较低的基础水平，转化生长因子β可能通过影响c-Fos蛋白的合成和/或稳定性，取消了对c-Fos的抑制，从而导致基因表达。
Transforming growth factor-beta (TGF beta) induces gene expression of the glycoprotein clusterin in a variety of cell types via a consensus AP-1 binding site. Here, we demonstrate, by supershift analysis, that JunB, JunD, Fra1, Fra2, and c-Fos bound to AP-1 but that prior treatment of the cells with TGF beta reduced dramatically c-Fos binding, suggesting that c-Fos might be playing a negative regulatory role in clusterin gene expression. Transient cotransfection assays in mink lung epithelial (CCL64) cells, using a human c-Fos expressing plasmid together with a clusterin promoter/reporter construct or the artificial TGF beta-inducible reporter construct 3TPLux, revealed that c-Fos was indeed repressive for TGF beta-induced promoter transactivation. Further, we demonstrate that in stable c-Fos-overexpressing cell lines, TGF beta induction of-endogenous clusterin mRNA, as well as clusterin promoter transactivation are blocked. Co-transfection with c-Fos deletion constructs revealed that the C-terminal region, including the homologue box 2 motif and the extreme C-terminal serine phosphorylation sites (Ser362 and Ser374) are required for repression of clusterin and 3TPLux transactivation. TGF beta treatment of CCL64 cells resulted in the induction of c-Fos mRNA but caused no alternation in total c-Fos protein levels. The results suggest that the c-Fos represses clusterin gene expression,, maintaining a low basal level in the absence of TGF beta, and that TGF beta, presumably through its effects on c-Fos protein synthesis and/or stability, abrogates the repression of c-Fos, thereby resulting in gene expression.