A promoter element of the CD-RAP gene is required for repression of gene expression in non-cartilage tissues in vitro and in vivo
A promoter element of the CD-RAP gene is required for repression of gene expression in non-cartilage tissues in vitro and in vivo
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DOI：
10.1002/jcb.20648
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发表时间：
2006-03-01
影响因子：
4
通讯作者：
Sandell, LJ
中科院分区：
文献类型：
--
作者：
Okazaki, K;Yu, H;Sandell, LJ
软骨源性视黄酸敏感蛋白（CD-RAP）基因主要在软骨中表达。先前对转基因小鼠的研究表明，CD-RAP基因-2,251 bp至-2,068 bp的DNA启动子片段含有基因表达的关键元件。随后的研究揭示了这个183bp元件的正调控基序和负调控基序。在这里，我们表明该元件在体外和体内根据细胞类型和转录因子的含量表现出基因表达的激活或抑制。Sox（阳性）和C/EBP（阴性）转录因子在细胞系和小鼠组织中的分布与它们的阳性和阴性作用一致。在转基因小鼠中，当从3345 bp的软骨特异性CD-RAP启动子中去除183 bp的元件时，报告基因的表达变得广泛，除了软骨外，还在肌肉、骨骼、肺和肝脏中观察到。在体外，C/EBP位点的突变激活了肌母细胞中失活的3345 bp的CD-RAP基因启动子，表明该位点负责（- 2079 bp）抑制。这些结果表明，183-bp元件在软骨特异性基因表达中发挥重要作用，作为软骨细胞调节模块，抑制非软骨细胞的转录并促进软骨细胞的激活。这是第一次报道在体内非软骨组织中抑制所必需的功能性DNA元件。
The cartilage-derived retinoic acid-sensitive protein (CD-RAP) gene is expressed predominately in cartilage. Previous studies in transgenic mice have shown that the DNA promoter segment from -2,251 bp to -2,068 bp of the CD-RAP gene contains elements critical for gene expression. Subsequent studies revealed both positive and negative regulatory motifs in this 183 bp element. Here we show that this element demonstrates activation or repression of gene expression in vitro and in vivo based on cell type and content of transcription factors. The distribution of Sox (positive) and C/EBP (negative) transcription factors in cell lines and in mouse tissues is consistent with their positive and negative roles. in transgenic mice, when the 183-bp element was removed from a 3,345-bp cartilage-specific CD-RAP promoter, expression of the reporter gene became widespread, being observed in muscle, bone, lung, and liver in addition to cartilage. In vitro, mutation of the C/EBP site activated the inactive 3,345-bp CD-RAP gene promoter in myoblastic cells, suggesting that this site is responsible for (-2,079 bp) repression. These results indicate that the 183-bp element plays an important role in cartilage-specific gene expression by acting as a chondrocyte-regulatory module repressing transcription in non-chondrocytes and contributing to activation in chondrocytes. This is the first report of a functional DNA element necessary for repression in non-cartilage tissues in vivo.