SOX9 protein is stabilized by TGF-β and regulates PAPSS2 mRNA expression in chondrocytes.
SOX9 protein is stabilized by TGF-β and regulates PAPSS2 mRNA expression in chondrocytes.
复制标题
SOX9蛋白通过TGF-β稳定，并调节软骨细胞中的PAPSS2 mRNA表达。
DOI：
10.1016/j.joca.2016.10.007
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发表时间：
2017-02
影响因子：
7
通讯作者：
Serra R
中科院分区：
文献类型：
--
作者：
Chavez RD;Coricor G;Perez J;Seo HS;Serra R
我们先前发现PAPSS2是转化生长因子-β在软骨细胞中的转录靶点。PAPSS2是软骨中蛋白多糖正确硫酸盐化所必需的。基质中有缺陷的硫酸盐会导致软骨的机械性能发生变化，这可能会导致软骨退化。这项研究的目的是确定调节PAPSS2表达的因素，并与已知的转化生长因子-β反应基因PrG4进行比较。本研究对转化生长因子β介导的SOX9的调控进行了研究，并探讨了SOX9对PAPSS2mRNA的调控作用。以微团培养的原代牛关节软骨细胞和ATDC5细胞为模型系统。用腺病毒表达SOX9和Smad3。用siRNA敲除Sox9和Smad3。用Western印迹和实时定量RT-PCR检测治疗后蛋白质和mRNA水平的变化。SOX9的过度表达足以上调PAPSS2基因的表达。转化生长因子-β作用于表达SOX9的细胞后，PAPSS2mRNA表达上调，提示SOX9与转化生长因子-β信号通路具有协同作用。此外，SOX9对于转化生长因子-β介导的PAPSS2的完全诱导也是必需的。相比之下，PRG4受SMAD3调控，而不受SOX9调控。经转化生长因子-β处理后，SOX9蛋白水平升高，而SOX9基因表达水平无明显变化。经转化生长因子-β处理后，SOX9蛋白在翻译后稳定。转化生长因子β稳定SOX9蛋白，而SOX9是转化生长因子β介导的PAPSS2mRNA调控的充分必要条件，为转化生长因子β介导的软骨细胞基因调控提供了一种新的机制。
We previously identified PAPSS2 as a transcriptional target of TGF-β in chondrocytes. PAPSS2 is required for proper sulfation of proteoglycans in cartilage. Defective sulfation in the matrix results in alterations in mechanical properties of the cartilage that would be expected to result in degeneration. The objective of this study was to identify factors that regulate PAPSS2 expression and compare to a known TGF-β responsive gene, PRG4. In this study, TGF-β-mediated regulation of SOX9 was characterized, and the involvement of SOX9 in regulation of PAPSS2 mRNA was investigated. Primary bovine articular chondrocytes grown in micromass culture and ATDC5 cells were used as the model system. Adenoviruses were used to express SOX9 and SMAD3. siRNA was used to knock-down Sox9 and Smad3. Western blot and real-time quantitative RT-PCR were used to measure changes in protein and mRNA levels in response to treatment. Over-expression of SOX9 was sufficient to up-regulate PAPSS2 mRNA. TGF-β treatment of SOX9-expressing cells resulted in enhanced up-regulation of PAPSS2 mRNA, suggesting that SOX9 cooperates with TGF-β signaling. Furthermore, Sox9 was required for full TGF-β-mediated induction of Papss2. In contrast, PRG4 was regulated by SMAD3 but not SOX9. SOX9 protein levels were increased after treatment with TGF-β although SOX9 mRNA was not. SOX9 protein was post-translationally stabilized after treatment with TGF-β. TGF-β stabilizes SOX9 protein, and SOX9 is sufficient and necessary for TGF-β-mediated regulation of PAPSS2 mRNA, providing a novel mechanism for TGF-β-mediated gene regulation in chondrocytes.