SOX9 enhances aggrecan gene promoter/enhancer activity and is up-regulated by retinoic acid in a cartilage-derived cell line, TC6
SOX9 enhances aggrecan gene promoter/enhancer activity and is up-regulated by retinoic acid in a cartilage-derived cell line, TC6
复制标题
DOI：
10.1074/jbc.275.15.10738
复制
发表时间：
2000-04-14
影响因子：
4.8
通讯作者：
Noda, M
中科院分区：
文献类型：
--
作者：
Sekiya, I;Tsuji, K;Noda, M
SOX 9是一种在软骨形成中起关键作用的转录因子，聚集蛋白聚糖是软骨的主要结构成分之一;然而，聚集蛋白聚糖基因调控的分子机制尚未完全阐明，TC 6是来源于关节软骨的克隆软骨细胞系，本研究的目的是检测SOX 9是否调节聚集蛋白聚糖基因的表达，并进一步鉴定在TC6中调节SOX 9表达的分子。细胞SOX 9在TC6细胞中的过表达增强了AgCAT-8构建体的转录活性的3倍，该构建体含有聚集蛋白聚糖基因的S-丝氨酸蛋白酶（kb）启动子/第一外显子/第一内含子片段。当我们从对应于包括第一内含子的区域的8-kb片段的3 '-末端缺失4.5-kb片段时，S0X9对聚集蛋白聚糖启动子活性的增强丧失。在TC6细胞中，S0X9使含有Sry/Sox共有序列的报告构建体的转录活性增强> 10倍。在成骨细胞骨肉瘤细胞（ROS17/2.8）中没有观察到SOX 9增强聚集蛋白聚糖基因启动子活性和SOX 9通过Sry/Sox共有序列的反式激活，表明依赖于细胞背景。北方印迹分析表明，TC6细胞组成性表达相对较低水平的Sox 9 mRNA。为了研究促钙激素和细胞因子对Sox 9基因表达的调节作用，我们研究了促钙激素和细胞因子对Sox 9基因表达的影响。Sox9表达的基础水平和RA的增强在允许温度（33 ℃）和非允许温度（39 ℃）下观察到相似。此外，RA处理增强了TC6细胞中含有Sry/Sox共有序列的报告构建体的转录活性。此外，RA治疗还增强了另一个报告构建体的转录活性，该报告构建体含有TC6细胞中II型前胶原基因的增强子区域。这些观察结果表明，SOX 9增强聚集蛋白聚糖启动子活性，并且其表达在TC6细胞中被RA上调。
SOX9 is a transcription factor that plays a key role in chondrogenesis, Aggrecan is one of the major structural components in cartilage; however, the molecular mechanism of aggrecan gene regulation has not yet been fully elucidated, TC6 is a clonal chondrocytic cell line derived from articular cartilage, The purpose of this study was to examine whether SOX9 modulates aggrecan gene expression and to further identify molecules that regulate Sox9 expression in TC6 cells. SOX9 overexpression in TC6 cells enhanced by similar to 3-fold the transcriptional activity of the AgCAT-8 construct containing S-kilobase (kb) promoter/first exon/first intron fragments of the aggrecan gene. SOX9 enhancement of aggrecan promoter activity was lost when we deleted a 4.5-kb fragment from the 3'-end of the 8-kb fragment corresponding to the region including the first intron, In TC6 cells, SOX9 enhanced the transcriptional activity of a reporter construct containing the Sry/Sox consensus sequence >10-fold. SOX9 enhancement of aggrecan gene promoter activity and SOX9 transactivation through the Sry/Sox consensus sequence were not observed in osteoblastic osteosarcoma cells (ROS17/2.8), indicating the dependence on the cellular background. Northern blot analysis indicated that TC6 cells constitutively express Sox9 mRNA at relatively low levels. To examine regulation of Sox9 gene expression, we investigated the effects of calciotropic hormones and cytokines, Among these, retinoic acid (RA) specifically enhanced Sox9 mRNA expression in TC6 cells. The basal levels of Sox9 expression and its enhancement by RA were observed similarly at both permissive (33 degrees C) and nonpermissive (39 degrees C) temperatures. Furthermore, RA treatment enhanced the transcriptional activity of a reporter construct containing the Sry/Sox consensus sequence in TC6 cells. Moreover, RA treatment also enhanced the transcriptional activity of another reporter construct containing the enhancer region of the type II procollagen gene in TC6 cells. These observations indicate that SOX9 enhances aggrecan promoter activity and that its expression is up-regulated by RA in TC6 cells.