pH-dependent pyridoxine transport by SLC19A2 and SLC19A3: Implications for absorption in acidic microclimates
pH-dependent pyridoxine transport by SLC19A2 and SLC19A3: Implications for absorption in acidic microclimates
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DOI：
10.1074/jbc.ra120.013610
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发表时间：
2020-12-11
影响因子：
4.8
通讯作者：
Yuasa, Hiroaki
中科院分区：
文献类型：
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作者：
Yamashiro, Takahiro;Yasujima, Tomoya;Yuasa, Hiroaki
SLC19A2和SLC19A3分别也称为硫胺转运体(THTR)1和2，它们将带正电荷的硫胺(维生素B1)转运到细胞内，使其能够有效利用。SLC19A2和SLC19A3也已知可以运输结构无关的阳离子药物，如二甲双胍，但这种电荷选择性是否延伸到其他分子，如吡哆醇(维生素B6)，尚不清楚。我们使用Madin-Darby犬肾II(MDCKII)细胞和人胚胎肾脏293(HEK293)细胞进行了转基因实验，并以Caco-2细胞作为人肠上皮细胞模型细胞。在MDCKII细胞中稳定表达SLC19A2和SLC19A3以及在HEK293细胞中瞬时表达SLC19A2和SLC19A3导致在pH 5.5时比对照细胞显著增加吡哆醇的摄取。诱导摄取是pH依赖的，有利于酸性条件，而不是中性到碱性条件，并且对原基敏感。SLC19A2和SLC19A3的表观K-m分别为37.8和18.5微米，并被吡哆醇类似物吡哆醛、吡哆胺和硫胺素抑制。我们还发现，用基因特异性siRNAs沉默Caco-2细胞的内源性SLC19A3，而不是SLC19A2，会导致载体介导的吡哆醇摄取显著减少。这些结果表明，SLC19A2和SLC19A3能够识别/转运吡哆醇，有利于操作的酸性条件，并提示这些转运蛋白可能主要在具有酸性环境的组织中发挥作用，如具有酸性表面小气候的小肠。
SLC19A2 and SLC19A3, also known as thiamine transporters (THTR) 1 and 2, respectively, transport the positively charged thiamine (vitamin B1) into cells to enable its efficient utilization. SLC19A2 and SLC19A3 are also known to transport structurally unrelated cationic drugs, such as metformin, but whether this charge selectivity extends to other molecules, such as pyridoxine (vitamin B6), is unknown. We tested this possibility using Madin-Darby canine kidney II (MDCKII) cells and human embryonic kidney 293 (HEK293) cells for transfection experiments, and also using Caco-2 cells as human intestinal epithelial model cells. The stable expression of SLC19A2 and SLC19A3 in MDCKII cells (as well as their transient expression in HEK293 cells) led to a significant induction in pyridoxine uptake at pH 5.5 compared with control cells. The induced uptake was pH-dependent, favoring acidic conditions over neutral to basic conditions, and protonophore-sensitive. It was saturable as a function of pyridoxine concentration, with an apparent K-m of 37.8 and 18.5 mu m, for SLC19A2 and SLC19A3, respectively, and inhibited by the pyridoxine analogs pyridoxal and pyridoxamine as well as thiamine. We also found that silencing the endogenous SLC19A3, but not SLC19A2, of Caco-2 cells with gene-specific siRNAs lead to a significant reduction in carrier-mediated pyridoxine uptake. These results show that SLC19A2 and SLC19A3 are capable of recognizing/transporting pyridoxine, favoring acidic conditions for operation, and suggest a possible role for these transporters in pyridoxine transport mainly in tissues with an acidic environment like the small intestine, which has an acidic surface microclimate.