Evidence for mitochondrial localization of divalent metal transporter 1 (DMT1)

Evidence for mitochondrial localization of divalent metal transporter 1 (DMT1)
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二价金属转运蛋白 1 (DMT1) 线粒体定位的证据

DOI:
10.1096/fj.13-240564
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发表时间:
2014
期刊:
The FASEB Journal
影响因子:
--
通讯作者:
Thévenod
Thévenod
中科院分区:
--
文献类型:
--
作者:
Garrick;Garrick;Fenton;Thévenod

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在哺乳动物细胞中,线粒体接收最多的铁进入,但尚未确定铁进入线粒体膜外膜(OMM)的途径。我们的结果表明,OMM中存在二价金属转运蛋白1(DMT1)。免疫印迹检测到他们的OMM中的肺细胞模型的线粒体中有DMT1。利用裂解泛素酵母双杂交系统,我们发现细胞色素氧化酶亚基II(COXII)和OMM 6-kDa亚基转位酶(Tom6)同源物与DMT1相互作用。COXII与DMT1共沉淀。在N端和C端有4种不同的DMT1亚型。利用HEK293细胞可诱导表达DMT1的全部4个末端,免疫印迹法检测到DMT1的4个末端均在OMM中表达,免疫荧光法检测到DMT1的4个末端均存在于分离的线粒体中。免疫印迹分析从大鼠肾皮质纯化的细胞部分证实了这些结果,并将其扩展到表达高水平DMT1的肾脏。免疫金标记法检测到DMT1与OMM中表达的电压依赖性阴离子选择通道蛋白-1在线粒体中共存。-Wolff,N.A.,Ghio,A.J.,Garrick,L.M.,Garrick,M.D.,赵,L.,Fenton,R.A.,ThéVenod,F.二价金属转运蛋白1(DMT1)的线粒体定位证据.FASE B J.28,2134-2145(2014).Www.fasebj.org
In mammalian cells, mitochondria receive most incoming iron, yet no entry pathway for iron at the outer mitochondrial membrane (OMM) has been characterized. Our results show that the divalent metal transporter 1 (DMT1) occurs in the OMM. Immunoblots detected DMT1 in mitochondria from a pneumocyte cell model in their OMM. Using the split‐ubiquitin yeast 2‐hybrid system, we found that cytochromecoxidase subunit II (COXII) and the translocase of OMM 6‐kDa subunit (Tom6) homologue interact with DMT1. COXII coimmunoprecipitates with DMT1. There are 4 DMT1 isoforms that differ at the N and C termini. Using HEK293 cells that inducibly express all of the 4 ends of DMT1, we found all of them in the OMM, as detected by immunoblots after cell fractionation, and in isolated mitochondria, as detected by immunofluorescence. Immunoblot analysis of purified cell fractions from rat renal cortex confirmed and extended these results to the kidney, which expressed high levels of DMT1. Immunogold labeling detected DMT1 colocalization in mitochondria with the voltage‐dependent anion‐selective channel protein‐1, which is expressed in the OMM. We suggest that DMT1 not only exports iron from endosomes, but also serves to import the metal into the mitochondria.—Wolff, N. A., Ghio, A. J., Garrick, L. M., Garrick, M. D., Zhao, L., Fenton, R. A., Thévenod, F. Evidence for mitochondrial localization of divalent metal transporter 1 (DMT1).FASEB J. 28, 2134–2145 (2014). www.fasebj.org
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