Abstract
Malignant melanoma, characterized by invasive local growth and early formation of metastases, is the most aggressive type of skin cancer. Melanoma inhibitory activity (MIA), secreted by malignant melanoma cells, interacts with the cell adhesion receptors, integrins α4β1 and α5β1, facilitating cell detachment and promoting formation of metastases. In the present study, we demonstrate that MIA secretion is confined to the rear end of migrating cells, while in non-migrating cells MIA accumulates in the actin cortex. MIA protein takes a conventional secretory pathway including coat protein complex I (COPI)- and coat protein complex II (COPII)-dependent protein transport to the cell periphery, where its final release depends on intracellular Ca2+ ions. Interestingly, the Ca2+-activated K+-channel, subfamily N, member 4 (KCa3.1), known to be active at the rear end of migrating cells, was found to support MIA secretion. Secretion was diminished by the specific KCa3.1 channel inhibitor TRAM-34 and by expression of dominant-negative mutants of the channel. In summary, we have elucidated the migration-associated transport of MIA protein to the cell rear and also disclosed a new mechanism by which KCa3.1 potassium channels promote cell migration.
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Abbreviations
- COPI:
-
(coat protein complex I)
- COPII:
-
(coat protein complex II)
- DMEM:
-
(Dulbecco's modified Eagle's medium)
- KCa3.1:
-
(Ca2+-activated K+-channel, subfamily N, member 4)
- MIA:
-
(melanoma inhibitory activity)
- MTOC:
-
(microtubule organizing center)
- SNARE:
-
(soluble N-ethylmaleimide-sensitive factor attachment receptor)
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Acknowledgements
We are indebted to Dr J Johnson (University of Munich, Germany) for providing the melanoma cell lines, Mel Im and Mel Ju, and to Dr B Distel (University of Amsterdam, Netherlands) for providing the Sar1p (H79G) vector. We also thank Dr A Schwab (University of Muenster, Germany) for the generous donation of the expression vector of the KCa3.1 channel and Dr C Voelker (University of Bonn, Germany) for the Golgi-GFP (endomannosidase) plasmid. This work was supported by grants from the DFG and the German Cancer Aid (Melanoma Research Network).
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(Supplementary information is linked to the online version of the paper on the Cell Research website.)
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Supplementary information, Figure S1
Effect of BFA treatment on the subcellular distribution of MMP14 and bFGF in the human melanoma cell line Mel Im. (PDF 110 kb)
Supplementary information, Figure S2
Effect of KCa3.1 activator 1-EBIO and inhibitor TRAM-34 on MIA protein secretion and migration in the presence of BAPTA AM in the human melanoma cell lines Mel Im and Mel Ju. (PDF 41 kb)
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Schmidt, J., Friebel, K., Schönherr, R. et al. Migration-associated secretion of melanoma inhibitory activity at the cell rear is supported by KCa3.1 potassium channels. Cell Res 20, 1224–1238 (2010). https://doi.org/10.1038/cr.2010.121
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DOI: https://doi.org/10.1038/cr.2010.121
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