Identification and characterization of human PCDH10 gene promoter
Identification and characterization of human PCDH10 gene promoter
复制标题
人PCDH10基因启动子的鉴定和表征
DOI：
10.1016/j.gene.2011.01.001
复制
发表时间：
2011-04-01
期刊：
影响因子：
3.5
通讯作者：
Cai, Zhiming
中科院分区：
文献类型：
--
作者：
Li, Zesong;Xie, Jun;Cai, Zhiming
最近的研究表明，PCDH 10作为一种新的肿瘤抑制基因的作用。在我们之前的工作中，我们将PCDH 10的核心启动子定位在一个462 bp的5 '侧翼区，该区域具有高GC含量的特征。在此，我们进一步鉴定并表征了PCDH 10的启动子。用一系列缺失的启动子构建体瞬时转染PC 3和LNCaP细胞表明，最小启动子区域在核苷酸-144和-99之间。该片段含有CAAT盒、GT盒和AP-4的推定转录因子结合位点。突变分析表明，CAAT盒和GT盒是启动子活性所必需的。NF-Ys的异位表达增加了报告基因的活性，而显性阴性NF-YA的表达降低了报告基因的活性。共转染Sp1/Sp3表达质粒以剂量依赖的方式增强报告基因的活性。光神霉素A Sp-DNA相互作用的抑制剂，降低PCDH 10启动子活性。电泳迁移率变动分析和染色质免疫沉淀证明了转录因子Sp1/Sp3在体外和体内与启动子区域的结合。我们的数据表明，Sp1/Sp3和CBF/NF-Y转录因子在人PCDH 10基因的基础表达中起着至关重要的作用。(C)2011 Elsevier B. V.保留所有权利。
Recent studies have suggested roles for PCDH10 as a novel tumor suppressor gene. In our previous work, we located the core promoter of PCDH10 to a 462-bp segment of 5'-flanking region characterized by a high GC content. Here we further identified and characterized the promoter for PCDH10. Transient transfection of PC3 and LNCaP cells with a series of deleted promoter constructs indicated that the minimal promoter region was between nucleotides - 144 and -99. This segment contained a CAAT box, a GT box, and a putative transcription factor binding site for AP-4. Mutational analysis identified that the CAAT box and GT box are necessary for promoter activity. Ectopic expression of NF-Ys increased reporter gene activity, whereas expression of a dominant-negative NF-YA decreased reporter gene activity. Co-transfection of Sp1/Sp3 expression plasmids enhanced reporter gene activity in a dose-dependent manner. Mithramycin A. an inhibitor of Sp-DNA interaction, reduced PCDH10 promoter activity. Electrophoretic mobility shift assays and chromatin immunoprecipitation demonstrated binding of transcription factors Sp1/Sp3 to the promoter region in vitro and in vivo. Our data show that Spl/Sp3 and CBF/NF-Y transcription factors play a crucial role in the basal expression of the human PCDH10 gene. (C) 2011 Elsevier B.V. All rights reserved.