Endocytosis of hERG is clathrin-independent and involves Arf6

Karnik, R. , Ludlow, M. J., Abuarab, N., Smith, A. J., Hardy, M. E.L., Elliott, D. J.S. and Sivaprasadarao, A. (2013) Endocytosis of hERG is clathrin-independent and involves Arf6. PLoS ONE, 8(12), e85630. (doi:10.1371/journal.pone.0085630) (PMID:24392021) (PMCID:PMC3877390)

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Abstract

The hERG potassium channel is critical for repolarisation of the cardiac action potential. Reduced expression of hERG at the plasma membrane, whether caused by hereditary mutations or drugs, results in long QT syndrome and increases the risk of ventricular arrhythmias. Thus, it is of fundamental importance to understand how the density of this channel at the plasma membrane is regulated. We used antibodies to an extracellular native or engineered epitope, in conjunction with immunofluorescence and ELISA, to investigate the mechanism of hERG endocytosis in recombinant cells and validated the findings in rat neonatal cardiac myocytes. The data reveal that this channel undergoes rapid internalisation, which is inhibited by neither dynasore, an inhibitor of dynamin, nor a dominant negative construct of Rab5a, into endosomes that are largely devoid of the transferrin receptor. These results support a clathrin-independent mechanism of endocytosis and exclude involvement of dynamin-dependent caveolin and RhoA mechanisms. In agreement, internalised hERG displayed marked overlap with glycosylphosphatidylinositol-anchored GFP, a clathrin-independent cargo. Endocytosis was significantly affected by cholesterol extraction with methyl-β-cyclodextrin and inhibition of Arf6 function with dominant negative Arf6-T27N-eGFP. Taken together, we conclude that hERG undergoes clathrin-independent endocytosis via a mechanism involving Arf6.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Karnik, Dr Rucha
Authors: Karnik, R., Ludlow, M. J., Abuarab, N., Smith, A. J., Hardy, M. E.L., Elliott, D. J.S., and Sivaprasadarao, A.
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:PLoS ONE
ISSN:1932-6203
ISSN (Online):1932-6203
Copyright Holders:Copyright © Karnik et al.
First Published:First published in PLoS ONE 8(12):e85630
Publisher Policy:Reproduced under a Creative Commons License

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