Artikel
The Zinc importer ZIP9 as mediator of glioma invasion
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Veröffentlicht: | 8. Juni 2016 |
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Gliederung
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Objective: A hallmark of gliomas is their diffuse infiltration in surrounding CNS tissue. Zinc is an essential element for the observed glioma migration and invasion. Zinc utilisation is controlled by zinc transporters and through buffering by metallothioneins and glutathione. Thus, zinc could mediate invasiveness and metastatic potential of glioma cells. Since both zinc- treatment and zinc-chelation can have detrimental effects on tumor cells, we aimed to investigate the individual expression profiles of zinc importers (ZIPs) and transporters (ZnT) in 6 human glioma cell lines (U373, U251, G28, G112, A172, U87) and their vulnerability to TPEN (N,N,N′,N′-tetrakis 2-pyridylmethyl ethylenediamine) induced zinc chelation.
Method: Resilience of glioma cell lines to TPEN treatment was tested by MTT proliferation assays. Using sublethal doses of TPEN (≤ 1μM), we analysed expression levels of ZnTs and ZIPs by qRT-PCR in the six glioma cell lines. In addition, migration behaviour of glioma cell lines was assessed by performing scratch assays. Selected ZnTs and ZIPs that were induced by sublethal doses of TPEN were analysed further by overexpression and knockdown experiments and the effect on cell migration and on intracellular kinase activation was tested.
Results: Upon zinc depletion using 1μM TPEN, we demonstrated by qPCR and Western Blotting that ZIP9 is selectively upregulated in expression levels in G28 and A172 glioma cells. When ZIP9 was expressed in these cell lines, enhanced cell migration of glioma cells was observed (G28: 25±5 for ZIP9 vs. 10±3 for GFP control; p<0.001). Analysis of intracellular signalling using a kinase array with 43 different kinases revealed that ZIP9 overexpression causes enhanced phosphorylation of GSK-3β. Application of a GSK-3β inhibitor caused reduced glioma migration, suggesting that ZIP9 can directly activate the Wnt signalling pathway via phophorylation of GSK-3β.
Conclusions: We have demonstrated that in particular ZIP9 is regulated by zinc depletion and that ZIP9 causes enhanced cell migration in glioma cells. Furthermore, ZIP9 acts on intracellular signalling as it causes phosphorylation of GSK-3β, an important effector of cancer cell migration. Taken together, we defined particularly ZIP9 as an important mediator of cell migration in glioma cells.