1)Sp1 upregulated the proximal promoter activity of the mouse collagen a1(XI) gene (Col11a1) in chondrocytes.
1)Sp1 upregulated the proximal promoter activity of the mouse collagen a1(XI) gene (Col11a1) in chondrocytes.
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1)Sp1上调软骨细胞中小鼠胶原蛋白a1(XI)基因(Col11a1)的近端启动子活性。
DOI：
10.1007/s11626-015-9959-y
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发表时间：
2016
期刊：
影响因子：
--
通讯作者：
Matsuo N.
中科院分区：
文献类型：
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作者：
Watanabe K;Hida M;Sasaki T;Yana H;Kawano K;Yoshioka H;Matsuo N.
XI型胶原是软骨特异性细胞外基质，并且对于胶原原纤维形成和骨骼形态发生是重要的。我们先前已经报道了NF-Y调节软骨细胞中小鼠胶原α1（XI）基因（Col 11 a1）的近端启动子活性（Hidaet.在VitroCell中。Dev. 2014）。然而，Col 11 a1基因在软骨细胞中的调控机制尚未完全阐明。在这项研究中，我们进一步的特点是近端启动子活动的mouseCol 11 a1基因在软骨细胞。用缺失和突变构建体进行的细胞转染实验表明，NF-γ结合位点的下游区域（-116至+1）对于调节小鼠Col 11 a1基因的近端启动子活性也是必需的。这个最小的启动子区域没有TATA盒和GC富集序列，因此我们研究了GC富集序列（-96至-67）是否是Col 11 a1基因转录调控所必需的。使用一系列突变构建体的荧光素酶测定表明，富含GC的序列是软骨细胞中Col 11 a1启动子活性的关键元件。此外，对该区域的计算机分析表明，最有效的候选者之一是转录因子Sp1。与预测一致，Sp1的过表达显著增加了启动子活性。此外，通过siRNA转染敲低Sp1表达抑制了小鼠Col 11 a1基因的近端启动子活性和内源性转录本的表达。综上所述，这些结果表明，转录因子Sp1上调近端启动子活性的小鼠Col 11 a1基因在软骨细胞中。
Type XI collagen is a cartilage-specific extracellular matrix, and is important for collagen fibril formation and skeletal morphogenesis. We have previously reported that NF-Y regulated the proximal promoter activity of the mouse collagen α1(XI) gene (Col11a1) in chondrocytes (Hidaet. al.In VitroCell. Dev. Biol. Anim.2014). However, the mechanism of theCol11a1gene regulation in chondrocytes has not been fully elucidated. In this study, we further characterized the proximal promoter activity of the mouseCol11a1gene in chondrocytes. Cell transfection experiments with deletion and mutation constructs indicated that the downstream region of the NF-Y binding site (-116 to +1) is also necessary to regulate the proximal promoter activity of the mouseCol11a1gene. This minimal promoter region has no TATA box and GC-rich sequence; we therefore examined whether the GC-rich sequence (-96 to -67) is necessary for the transcription regulation of theCol11a1gene. Luciferase assays using a series of mutation constructs exhibited that the GC-rich sequence is a critical element ofCol11a1promoter activity in chondrocytes. Moreover, in silico analysis of this region suggested that one of the most effective candidates was transcription factor Sp1. Consistent with the prediction, overexpression of Sp1 significantly increased the promoter activity. Furthermore, knockdown of Sp1 expression by siRNA transfection suppressed the proximal promoter activity and the expression of endogenous transcript of the mouseCol11a1gene. Taken together, these results indicate that the transcription factor Sp1 upregulates the proximal promoter activity of the mouseCol11a1gene in chondrocytes.