Characterization of the human peptide transporter PEPT1 promoter: Sp1 functions as a basal transcriptional regulator of human PEPT1
Characterization of the human peptide transporter PEPT1 promoter: Sp1 functions as a basal transcriptional regulator of human PEPT1
复制标题
DOI：
10.1152/ajpgi.00025.2005
复制
发表时间：
2005-09-01
影响因子：
4.5
通讯作者：
Inui, KI
中科院分区：
文献类型：
--
作者：
Shimakura, J;Terada, T;Inui, KI
H+偶联肽转运蛋白1（PEPT 1，SLC 15 A1）定位于肠上皮细胞刷状缘膜，在小分子肽和多种拟肽药物的肠道吸收中起重要作用。PEPT 1受多种因素调节，包括激素、饮食条件、某些药物和昼夜节律。但是关于PEPT 1的转录调控的信息很少。因此，在本研究中，我们克隆了人（h）PEPT 1启动子区域，并使用人肠细胞系Caco-2检测其启动子活性。对hPEPT 1启动子的缺失分析表明，跨越-172至-35 bp的区域是基础转录活性所必需的。该区域缺乏TATA盒，但含有一些富含GC的位点，据推测这些位点与转录因子Sp1结合。突变分析表明，这些推定的Sp1位点的转录活性作出了贡献。EMSA分析表明Sp1与两个富含GC的位点结合。此外，通过光神霉素A处理抑制Sp1结合显著降低了转录活性。最后，Sp1的过表达以剂量依赖的方式增加转录活性。本研究报告的第一个特征的hPEPT 1启动子，并显示了显着的作用，Sp1的基础转录调控的hPEPT 1。
H+-coupled peptide transporter 1 (PEPT1, SLC15A1) localized at the brush-border membranes of intestinal epithelial cells plays an important role in the intestinal absorption of small peptides and a variety of peptidemimetic drugs. PEPT1 is regulated by various factors, including hormones, dietary conditions, some pharmaceutics, and diurnal rhythm. But there is little information about the transcriptional regulation of PEPT1. In the present study, therefore, we cloned the human (h) PEPT1 promoter region and examined its promoter activity using a human intestinal cell line, Caco-2. Deletion analysis of the hPEPT1 promoter suggested that the region spanning -172 to -35 bp was essential for basal transcriptional activity. This region lacked a TATA-box but contained some GC-rich sites that supposedly bind with the transcription factor Sp1. Mutational analysis revealed that three of these putative Sp1 sites contributed to the transcriptional activity. EMSA showed that Sp1 bound to two GC-rich sites. Furthermore, inhibition of Sp1 binding by mithramycin A treatment significantly reduced the transcriptional activity. Finally, overexpression of Sp1 increased the transcriptional activity in a dose-dependent manner. This study reports the first characterization of the hPEPT1 promoter and shows the significant role of Sp1 in the basal transcriptional regulation of hPEPT1.