Human erythrocyte nucleoside transporter ENT1 functions at ice-cold temperatures.
Human erythrocyte nucleoside transporter ENT1 functions at ice-cold temperatures.
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DOI：
10.2133/dmpk.dmpk-09-rg-099
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发表时间：
2010
影响因子：
2.1
通讯作者：
M. Takano;E. Kimura;Satoshi Suzuki;J. Nagai;R. Yumoto
中科院分区：
文献类型：
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作者：
M. Takano;E. Kimura;Satoshi Suzuki;J. Nagai;R. Yumoto
在冰冷温度(ICT；测量温度，0.5-0.7摄氏度)下，使用直立式膜泡(ROV)检测了人红细胞核苷转运体ENT1的功能。ENT1底物尿苷的摄取呈饱和动力学，并在23℃和ICT条件下被特异性ENT1抑制剂S-(4-硝基苄基)-6-硫代肌苷抑制。用未标记的尿苷或利巴韦林(另一种ENT1底物)预加载ROV可显著反式刺激[~3H]尿苷摄取，并在ICT观察到超调现象。同样，在ICT中，未标记的利巴韦林或尿苷显著反式刺激[~3H]利巴韦林摄取。ENT1抑制剂/底物如NBMPR、双嘧达莫、腺苷和利巴韦林以浓度依赖的方式抑制ICT反式刺激的[~3H]尿苷摄取。在完整红细胞上，NBMPR和潘生丁对[~3H]尿苷摄取的抑制作用也被观察到。与尿苷摄取一样，由促进性葡萄糖转运体GLUT1介导的ROV摄取[~3H]D-葡萄糖在ICT时被未标记的D-葡萄糖反式刺激，并观察到超调现象。相反，通过多药耐药相关蛋白5转运5-(和-6)-羧基-2‘，7’-二氯荧光素的能力在ICT时消失。这些结果清楚地表明，人类红细胞转运体，如ENT1，即使在接近0摄氏度的非常低的温度下也能发挥作用。本文讨论了这些发现在转运体研究中的意义。
The functionality of human erythrocyte nucleoside transporter ENT1 was examined at ice-cold temperatures (ICT; measured temperature, 0.5-0.7 degrees C) using rightside-out membrane vesicles (ROVs). The uptake of uridine, an ENT1 substrate, showed saturation kinetics and was inhibited by S-(4-nitrobenzyl)-6-thioinosine (NBMPR), a specific ENT1 inhibitor, at both 23 degrees C and ICT. [3H]Uridine uptake was markedly trans-stimulated by preloading ROVs with unlabeled uridine or ribavirin, another ENT1 substrate, and the overshoot phenomenon was observed at ICT. Similarly, [3H]ribavirin uptake was markedly trans-stimulated by unlabeled ribavirin or uridine at ICT. The trans-stimulated uptake of [3H]uridine at ICT was inhibited by ENT1 inhibitors/substrates such as NBMPR, dipyridamole, adenosine, and ribavirin in a concentration-dependent manner. The inhibition of [3H]uridine uptake by NBMPR and dipyridamole at ICT was also observed in intact red blood cells. Like uridine uptake, [3H]D-glucose uptake by ROVs, which is mediated by facilitative glucose transporter GLUT1, was trans-stimulated by unlabeled D-glucose at ICT, and the overshoot phenomenon was observed. In contrast, the ability of ATP-dependent transport of 5-(and-6)-carboxy-2',7'-dichlorofluorescein via multidrug resistance-associated protein 5 in inside-out membrane vesicles disappeared at ICT. These results clearly indicate that human erythrocyte transporters such as ENT1 function even at very low temperatures near 0 degrees C. The significance of these findings in transporter research is discussed.