Vitamin D3 transactivates the zinc and manganese transporter SLC30A10 via the Vitamin D receptor
Vitamin D3 transactivates the zinc and manganese transporter SLC30A10 via the Vitamin D receptor
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DOI：
10.1016/j.jsbmb.2016.04.006
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发表时间：
2016-10-01
影响因子：
4.1
通讯作者：
Kullak-Ublick, Gerd A.
中科院分区：
文献类型：
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作者：
da Silva, Tatiana Claro;Hiller, Christian;Kullak-Ublick, Gerd A.
维生素D-3调节对人类健康至关重要的基因，其缺乏与骨质疏松症、癌症、糖尿病、多发性硬化症、高血压、炎症和免疫性疾病的风险增加有关。为了研究维生素D-3对营养物质和药物的运输和代谢相关基因的影响，我们采用了下一代测序（NGS），并分析了用500 nM维生素D-3处理的人源Caco-2细胞系的整体基因表达。参与神经肽信号传导、炎症、细胞粘附和形态发生的基因差异表达。值得注意的是，涉及锌，锰和铁稳态的基因在很大程度上增加了维生素D-3治疗。铜蓝蛋白增加10倍，结合珠蛋白基因表达增加4倍，表明维生素D和铁稳态之间可能存在关联。编码锌锰转运蛋白ZnT 10的基因SLC 30 A10是主要受影响的转运蛋白，其表达增加了约15倍。SLC 30 A10对锌和锰的体内平衡至关重要，该基因的突变导致ZnT 10功能或表达受损，导致锰中毒，伴有帕金森样症状。我们的NGS结果通过Caco-2细胞中的实时PCR以及从每天用0.5 μ g维生素D-3治疗10天的健康人受试者中采集的十二指肠活检进行了验证。除了增加SLC 30 A10和阳性对照TRPV 6的基因表达外，维生素D-3还增加ZnT 10蛋白的表达，如Western印迹和细胞荧光所示。在SLC 30 A10启动子的5 '侧翼区域中潜在的维生素D响应元件（VDRE）的计算机鉴定和双荧光素酶报告基因测定显示，在维生素D受体（VDR）和类维生素A X受体（RXR）构建体以及维生素D-3存在下，启动子活性增强，但当这些因子之一不存在时，启动子活性不增强。电泳迁移率变动分析（EMSA）和竞争性EMSA揭示了选择序列，即相对于转录起始位点的nt-16231 - 1588和nt-1758/-1723与含有VDR的核提取物的结合。总之，我们已经表明，维生素D-3反式激活SLC 30 A10基因的VDR依赖性的方式，导致增加ZnT 10蛋白的表达。由于SLC 30 A10在小肠中高度表达，因此锌和锰的全身水平的控制可能由肠道中的维生素D-3调节。锌、锰和维生素D对骨骼代谢和大脑健康很重要。未来对锌和锰的补充或螯合以及维生素D-3给药的可能作用的研究，将进一步了解其在治疗特定疾病（如骨质疏松症和帕金森病）方面的潜在益处。(C)2016作者（S）爱思唯尔有限公司出版
Vitamin D-3 regulates genes critical for human health and its deficiency is associated with an increased risk for osteoporosis, cancer, diabetes, multiple sclerosis, hypertension, inflammatory and immunological diseases. To study the impact of vitamin D-3 on genes relevant for the transport and metabolism of nutrients and drugs, we employed next-generation sequencing (NGS) and analyzed global gene expression of the human-derived Caco-2 cell line treated with 500 nM vitamin D-3. Genes involved in neuropeptide signaling, inflammation, cell adhesion and morphogenesis were differentially expressed. Notably, genes implicated in zinc, manganese and iron homeostasis were largely increased by vitamin D-3 treatment. An similar to 10-fold increase in ceruloplasmin and similar to 4-fold increase in haptoglobin gene expression suggested a possible association between vitamin D and iron homeostasis. SLC30A10, the gene encoding the zinc and manganese transporter ZnT10, was the chiefly affected transporter, with similar to 15-fold increase in expression. SLC30A10 is critical for zinc and manganese homeostasis and mutations in this gene, resulting in impaired ZnT10 function or expression, cause manganese intoxication, with Parkinson-like symptoms. Our NGS results were validated by real-time PCR in Caco-2 cells, as well as in duodenal biopsies taken from healthy human subjects treated with 0.5 mu g vitamin D-3 daily for 10 days. In addition to increasing gene expression of SLC30A10 and the positive control TRPV6, vitamin D-3 also increased ZnT10 protein expression, as indicated by Western blot and cytofluorescence. In silico identification of potential vitamin D responsive elements (VDREs) in the 5'-flanking region of the SLC30A10 promoter and dual-luciferase reporter assay showed enhanced promoter activity in the presence of vitamin D receptor (VDR) and retinoid X receptor (RXR) constructs, as well as vitamin D-3, but not when one of these factors was absent. Electrophoretic mobility shift assay (EMSA) and competition EMSA revealed binding of select sequences, namely, nt -16231-1588 and nt -1758/-1723 relative to the transcription start site, to VDR-containing nuclear extracts. In conclusion, we have shown that vitamin D-3 transactivates the SLC30A10 gene in a VDR-dependent manner, resulting in increased ZnT10 protein expression. Because SLC30A10 is highly expressed in the small intestine, it is possible that the control of zinc and manganese systemic levels is regulated by vitamin D-3 in the intestine. Zinc, manganese and vitamin D are important for bone metabolism and brain health. Future examination of a possible role for supplementation or chelation of zinc and manganese, alongside vitamin D-3 administration, will further our understanding of its potential benefit in the treatment of specific illnesses, such as osteoporosis and Parkinson's disease. (C) 2016 The Author(s). Published by Elsevier Ltd.