Grigat Silke, Harlfinger Stephanie, Pal Sonia, Striebinger Ralph, Golz Stefan, Geerts Andreas, Lazar Andreas, Schömig Edgar, Gründemann Dirk
Department of Pharmacology, University of Cologne, Gleueler Strasse 24, 50931 Cologne, Germany.
Biochem Pharmacol. 2007 Jul 15;74(2):309-16. doi: 10.1016/j.bcp.2007.04.015. Epub 2007 Apr 22.
Recently, we have identified the ergothioneine (ET) transporter ETT (gene symbol SLC22A4). Much interest in human ETT has been generated by case-control studies that suggest an association of polymorphisms in the SLC22A4 gene with susceptibility to chronic inflammatory diseases. ETT was originally designated a multispecific novel organic cation transporter (OCTN1). Here we reinvestigated, based on stably transfected 293 cells and with ET as reference substrate, uptake of quinidine, verapamil, and pyrilamine. ETT from human robustly catalyzed transport of ET (68micfrol/(minmgprotein)), but no transport of organic cations was discernible. With ET as substrate, ETT was relatively resistant to inhibition by selected drugs; the most potent inhibitor was verapamil (K(i)=11micromol/l). The natural compound hercynine and antithyroid drug methimazole are related in structure to ET. However, efficiency of ETT-mediated transport of methimazole (K(i)=7.5mmol/l) was 130-fold lower, and transport of hercynine (K(i)=1.4mmol/l) was 25-fold lower than transport of ET. ETT from mouse, upon expression in 293 cells, catalyzed high affinity, sodium-driven uptake of ET very similar to ETT from human. Additional real-time PCR experiments based on 16 human tissues revealed ETT mRNA levels considerably lower than in bone marrow. Our experiments establish that ETT is highly specific for its physiological substrate ergothioneine. ETT is not a cationic drug transporter, and it does not have high affinity for organic cation inhibitors. Detection of ETT mRNA or protein can therefore be utilized as a specific molecular marker of intracellular ET activity.
最近，我们鉴定出了麦角硫因（ET）转运体ETT（基因符号SLC22A4）。病例对照研究表明SLC22A4基因多态性与慢性炎症性疾病易感性相关，这引发了人们对人类ETT的极大兴趣。ETT最初被指定为一种多特异性新型有机阳离子转运体（OCTN1）。在此，我们以稳定转染的293细胞为基础，以ET作为参考底物，重新研究了奎尼丁、维拉帕米和吡拉明的摄取情况。来自人类的ETT能高效催化ET的转运（68微升/（分钟·毫克蛋白）），但未发现其对有机阳离子的转运。以ET为底物时，ETT对所选药物的抑制相对具有抗性；最有效的抑制剂是维拉帕米（抑制常数K(i)=11微摩尔/升）。天然化合物组氨酸和抗甲状腺药物甲巯咪唑在结构上与ET相关。然而，ETT介导的甲巯咪唑转运效率（抑制常数K(i)=7.5毫摩尔/升）比ET低130倍，组氨酸的转运效率（抑制常数K(i)=1.4毫摩尔/升）比ET低25倍。在293细胞中表达的小鼠ETT催化ET的高亲和力、钠依赖性摄取，这与人类ETT非常相似。基于16种人体组织的额外实时PCR实验显示，ETT mRNA水平远低于骨髓中的水平。我们的实验证实，ETT对其生理底物麦角硫因具有高度特异性。ETT不是阳离子药物转运体，对有机阳离子抑制剂也没有高亲和力。因此，检测ETT mRNA或蛋白可作为细胞内ET活性的特异性分子标记。
