Identification and characterization of a novel family of membrane magnesium transporters, MMgT1 and MMgT2
Identification and characterization of a novel family of membrane magnesium transporters, MMgT1 and MMgT2
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DOI：
10.1152/ajpcell.00238.2007
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发表时间：
2008-02-01
影响因子：
5.5
通讯作者：
Quamme, Gary A.
中科院分区：
文献类型：
--
作者：
Goytain, Angela;Quamme, Gary A.
镁是一种必需的金属，但很少有选择性转运蛋白已被确定在分子水平上。微阵列分析被用来确定两个类似的转录上调与低细胞外Mg 2+。相应的cDNA编码具有两个预测的跨膜结构域的131和123个氨基酸的蛋白质。这两个独立的基因产物组成了我们称之为“膜Mg 2+转运蛋白”（MMgTs）的家族，因为这些蛋白质存在于膜中并介导Mg 2+转运。当在非洲爪蟾卵母细胞中表达时，MMgT 1和MMgT 2介导Mg 2+转运，如用双电极电压钳分析和荧光测量所确定的。转运是可饱和的Mg 2+摄取，米氏常数分别为1.47 +/- 0.17和0.58 +/- 0.07 mM。实时荧光定量RT-PCR证实MMgT mRNA存在于多种细胞中。免疫组织化学亚细胞定位确定MMgT 1-血凝素（HA）和MMgT 2-V5融合蛋白存在于高尔基复合体和高尔基后囊泡中，包括转染相应标记构建体的COS-7细胞中的早期内体。有趣的是，MMgT 1-HA和MMgT 2-V5被发现在不同的群体后高尔基体囊泡。MMgT 1和MMgT 2 mRNA分别增加了约3倍，在肾上皮细胞培养在低镁培养基相对于正常的媒体和维持低镁饮食的小鼠的肾皮质相比，这些动物消耗正常饮食。随着转录本的增加，高尔基体和高尔基体后囊泡中的MMgT 1和MMgT 2蛋白明显增加。这些实验表明，MMgT蛋白可能提供调节途径的Mg 2+运输的高尔基体和后高尔基体细胞器的上皮细胞。
Magnesium is an essential metal, but few selective transporters have been identified at the molecular level. Microarray analysis was used to identify two similar transcripts that are upregulated with low extracellular Mg2+. The corresponding cDNAs encode proteins of 131 and 123 amino acids with two predicted transmembrane domains. The two separate gene products comprise the family that we have termed '' membrane Mg2+ transporters '' (MMgTs), because the proteins reside in the membrane and mediate Mg2+ transport. When expressed in Xenopus laevis oocytes, MMgT1 and MMgT2 mediate Mg2+ transport as determined with two-electrode voltage-clamp analysis and fluorescence measurements. Transport is saturable Mg2+ uptake with Michaelis constants of 1.47 +/- 0.17 and 0.58 +/- 0.07 mM, respectively. Real-time RT-PCR demonstrated that MMgT mRNAs are present in a wide variety of cells. Subcellular localization with immunohistochemistry determined that the MMgT1-hemagglutinin (HA) and MMgT2-V5 fusion proteins reside in the Golgi complex and post-Golgi vesicles, including the early endosomes in COS-7 cells transfected with the respective tagged constructs. Interestingly, MMgT1-HA and MMgT2-V5 were found in separate populations of post-Golgi vesicles. MMgT1 and MMgT2 mRNA increased by about threefold, respectively, in kidney epithelial cells cultured in low-magnesium media relative to normal media and in the kidney cortex of mice maintained on low-magnesium diets compared with those animals consuming normal diets. With the increase in transcripts, there was an apparent increase in MMgT1 and MMgT2 protein in the Golgi and post-Golgi vesicles. These experiments suggest that MMgT proteins may provide regulated pathways for Mg2+ transport in the Golgi and post-Golgi organelles of epithelium-derived cells.