Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan.
Departments of Medicine and Physiology/Biophysics, University of California, Irvine, California, USA; Department of Veterans Affairs Medical Center, Long Beach, California, USA.
J Biol Chem. 2020 Dec 11;295(50):16998-17008. doi: 10.1074/jbc.RA120.013610. Epub 2020 Oct 2.
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 of 37.8 and 18.5 μm, 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.
SLC19A2 和 SLC19A3，也分别被称为硫胺素转运体 (THTR)1 和 2，将带正电荷的硫胺素（维生素 B1）转运到细胞内以使其得到有效利用。SLC19A2 和 SLC19A3 也被发现可以转运结构上不相关的阳离子药物，如二甲双胍，但这种电荷选择性是否扩展到其他分子，如吡哆醇（维生素 B6），尚不清楚。我们使用 Madin-Darby 犬肾 II（MDCKII）细胞和人胚肾 293（HEK293）细胞进行转染实验，以及使用 Caco-2 细胞作为人肠道上皮模型细胞来测试这种可能性。在 MDCKII 细胞中稳定表达 SLC19A2 和 SLC19A3（以及在 HEK293 细胞中瞬时表达）导致与对照细胞相比，在 pH 5.5 时吡哆醇摄取显著增加。这种诱导的摄取依赖于 pH 值，在酸性条件下比中性至碱性条件下更有利，并且对质子载体敏感。它作为吡哆醇浓度的函数是饱和的，对于 SLC19A2 和 SLC19A3，分别有 37.8 和 18.5 μm 的表观，并且被吡哆醇类似物吡哆醛和吡哆胺以及硫胺素抑制。我们还发现，用基因特异性 siRNA 沉默 Caco-2 细胞中的内源性 SLC19A3，但不是 SLC19A2，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，有利于酸性条件下的操作，并提示这些转运体在吡哆醇转运中可能发挥作用，主要在像小肠这样具有酸性环境的组织中，其具有酸性表面微环境。
