Section of Digestive Diseases & Nutrition, Department of Medicine, University of Illinois at Chicago and Jesse Brown VA Medical Center, 60612, USA.
Am J Physiol Gastrointest Liver Physiol. 2011 Apr;300(4):G627-36. doi: 10.1152/ajpgi.00563.2010. Epub 2011 Jan 27.
Serotonin transporter (SERT) regulates extracellular availability of serotonin and is a potential pharmacological target for gastrointestinal disorders. A decrease in SERT has been implicated in intestinal inflammatory and diarrheal disorders. However, little is known regarding regulation of SERT in the intestine. Epidermal growth factor (EGF) is known to influence intestinal electrolyte and nutrient transport processes and has protective effects on intestinal mucosa. Whether EGF regulates SERT in the human intestine is not known. The present studies examined the regulation of SERT by EGF, utilizing Caco-2 cells grown on Transwell inserts as an in vitro model. Treatment with EGF from the basolateral side (10 ng/ml, 24 h) significantly stimulated SERT activity (∼2-fold, P < 0.01) and mRNA levels compared with control. EGF increased the activities of the two alternate promoter constructs for human SERT gene: SERT promoter 1 (hSERTp1, upstream of exon 1a) and SERT promoter 2 (hSERTp2, upstream of exon 2). Inhibition of EGF receptor (EGFR) tyrosine kinase activity by PD168393 (1 nM) blocked the stimulatory effects of EGF on SERT promoters. Progressive deletions of the SERT promoter indicated that the putative EGF-responsive elements are present in the -672/-472 region of the hSERTp1 and regions spanning -1195/-738 and -152/+123 of hSERTp2. EGF markedly increased the binding of Caco-2 nuclear proteins to the potential AP-1 cis-elements present in EGF-responsive regions of hSERTp1 and p2. Overexpression of c-jun but not c-fos specifically transactivated hSERTp2, with no effects on hSERTp1. Our findings define novel mechanisms of transcriptional regulation of SERT by EGF via EGFR at the promoter level that may contribute to the beneficial effects of EGF in gut disorders.
5-羟色胺转运体（SERT）调节 5-羟色胺的细胞外含量，是胃肠道疾病的潜在药物靶点。SERT 减少与肠道炎症性和腹泻性疾病有关。然而，关于 SERT 在肠道中的调节作用知之甚少。表皮生长因子（EGF）已知会影响肠道电解质和营养物质转运过程，并对肠道黏膜具有保护作用。EGF 是否调节人肠道中的 SERT 尚不清楚。本研究利用 Caco-2 细胞在 Transwell 插入物上生长的体外模型，研究了 EGF 对 SERT 的调节作用。与对照相比，从基底外侧侧（10ng/ml，24 小时）用 EGF 处理显著刺激 SERT 活性（约 2 倍，P<0.01）和 mRNA 水平。EGF 增加了人类 SERT 基因的两个替代启动子构建体的活性：SERT 启动子 1（hSERTp1，exon1a 上游）和 SERT 启动子 2（hSERTp2，exon2 上游）。PD168393（1nM）抑制 EGF 受体（EGFR）酪氨酸激酶活性，阻断 EGF 对 SERT 启动子的刺激作用。SERT 启动子的逐步缺失表明，假定的 EGF 反应元件存在于 hSERTp1 的-672/-472 区域和跨越 hSERTp2 的-1195/-738 和-152/+123 的区域。EGF 显著增加了 Caco-2 核蛋白与 hSERTp1 和 p2 的 EGF 反应区存在的潜在 AP-1 顺式元件的结合。c-jun 的过表达而非 c-fos 特异性地转激活 hSERTp2，而对 hSERTp1 没有影响。我们的研究结果定义了 EGF 通过 EGFR 在启动子水平上对 SERT 的转录调节的新机制，这可能有助于 EGF 在肠道疾病中的有益作用。
