A silencer element in the cartilage oligomeric matrix protein gene regulates chondrocyte-specific expression
A silencer element in the cartilage oligomeric matrix protein gene regulates chondrocyte-specific expression
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DOI：
10.1016/j.orthres.2003.12.006
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发表时间：
2004-07-01
影响因子：
2.8
通讯作者：
Di Cesare, PE
中科院分区：
文献类型：
--
作者：
Issack, PS;Liu, CJ;Di Cesare, PE
间充质细胞分化为软骨细胞的分子机制知之甚少。软骨寡聚基质蛋白基因（COMP）编码一种非胶原性细胞外基质蛋白，其表达模式与软骨细胞分化和关节炎相关。我们已经使用COMP启动子作为模型，以确定软骨细胞特异性表达所需的调控序列，并确定结合这些序列的细胞类型特异性蛋白质。我们以前克隆了鼠COMP基因5'侧翼启动子序列的1.9个酶，并通过缺失分析鉴定了两个空间上相距较远的软骨细胞特异性调节区。一个元件位于转录起始位点的近端（-125至-75），第二个区域位于转录起始位点的远端（-1925至-592）。在本研究中，我们对COMP启动子-1925到-592区域进行了更精细的缺失分析，并鉴定了位于-1775和-1725之间的沉默子区域。这种沉默剂结合序列特异性蛋白复合物;这些复合物的强度在两种不同的成纤维细胞百合（NIH 3 T3和10 T1/2）中比在软骨细胞RCS细胞中更大。竞争实验定位这些蛋白质复合物的结合位点从-1775到-1746;删除这个30 bp的位点导致COMP启动子活性在成纤维细胞中的选择性增加。这个30 bp的位点的四个串联重复序列足以在N1 H3 T3成纤维细胞中对异源启动子（SV 40）赋予负转录调控。这些结果表明，转录的负调控是软骨细胞特异性表达COMP基因的重要机制。(C)2003骨科研究学会。由爱思唯尔有限公司出版。保留所有权利。
The molecular mechanisms by which mesenchymal cells differentiate into chondrocytes are poorly understood. The cartilage oligomeric matrix protein gene (COMP) encodes a noncollagenous extracellular matrix protein whose expression pattern correlates with chondrocyte differentiation and arthritis. We have used the COMP promoter as a model to identify regulatory sequences necessary for chondrocyte-specific expression and to identify cell type-specific proteins that bind these sequences. We have previously cloned 1.9 kilobases of the 5' flanking promoter sequence of the murine COMP gene and by deletion analysis have identified two spatially distant chondrocyte-specific regulatory regions. One element is Situated proximally (-125 to -75), and a second region is located distally (-1925 to -592) relative to the transcription start site. In the present study, we performed a finer deletion analysis of the region of the COMP promoter from -1925 to -592 and identified a silencer region Situated between -1775 and -1725. This silencer binds sequence-specific protein complexes; the intensity of these complexes is greater in two different fibroblast cell lilies (NIH3T3 and 10T1/2) than in chondrocytic RCS cells. Competition experiments localized the binding site of these protein complexes from - 1775 to - 1746; deletion of this 30-bp site results in a selective increase in COMP promoter activity in fibroblasts. Four tandem repeats of this 30-bp site are sufficient to confer negative transcriptional regulation on a heterologous promoter (SV40) in N1H3T3 fibroblasts. These results Suggest that negative regulation of transcription is an important mechanism for chondrocyte-specific expression of the COMP gene. (C) 2003 Orthopaedic Research Society. Published by Elsevier Ltd. All rights reserved.