Deubiquitylating enzyme USP2 counteracts Nedd4-2-mediated downregulation of KCNQ1 potassium channels.
BACKGROUND
KCNQ1 (Kv7.1), together with its KCNE β subunits, plays a pivotal role both in the repolarization of cardiac tissue and in water and salt transport across epithelial membranes. Nedd4/Nedd4-like (neuronal precursor cell-expressed developmentally downregulated 4) ubiquitin-protein ligases interact with the KCNQ1 potassium channel through a PY motif located in the C terminus of KCNQ1. This interaction induces ubiquitylation of KCNQ1, resulting in a reduced surface density of the channel. It was reported recently that the epithelial sodium channel is regulated by the reverse process-deubiquitylation-mediated by USP2 (ubiquitin-specific protease 2).


OBJECTIVE
In this article, we investigated whether deubiquitylation may regulate KCNQ1 channel complexes.


METHODS
In this study, we used electrophysiology, biochemistry, and confocal microscopy.


RESULTS
Electrophysiological investigations of KCNQ1/KCNE1 proteins coexpressed with USP2-45 or USP2-69 isoforms and Nedd4-2 in Xenopus laevis oocytes and mammalian cells revealed that both USP2 isoforms counter the Nedd4-2-specific downregulation of I(Ks). Biochemical studies showed that the total and surface-expressed KCNQ1 protein was more abundant when coexpressed with USP2 and Nedd4-2 as compared with Nedd4-2 alone. Western blotting revealed partial protection against covalent attachment of ubiquitin moieties on KCNQ1 when USP2 was coexpressed with Nedd4-2. Coimmunoprecipitation assays suggested that USP2 can bind to KCNQ1 independently of the PY motif. Immunocytochemistry confirmed that USP2 restores the membrane localization of KCNQ1.


CONCLUSIONS
These results demonstrate that USP2 can be a potent regulator of KCNQ1 surface density. USP2, which is well expressed in many tissues, may therefore be important in controlling the KCNQ1 channel dynamics in vivo.
Danish National Research Foundation Centre for Cardiac Arrhythmia, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark.
Katarzyna
K
Krzystanek
Hanne Borger
HB
Rasmussen
Morten
M
Grunnet
Olivier
O
Staub
Søren-Peter
SP
Olesen
Hugues
H
Abriel
Thomas
T
Jespersen
D000200:Action Potentials / Q000502:physiology; D000818:Animals; D002462:Cell Membrane / Q000502:physiology; D015536:Down-Regulation; D010450:Endopeptidases / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D015640:Ion Channel Gating / Q000502:physiology*; D051657:KCNQ1 Potassium Channel / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism; D000075702:Nedd4 Ubiquitin Protein Ligases; D021381:Protein Transport / Q000502:physiology; D025801:Ubiquitin / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D064570:Ubiquitin-Specific Proteases; D054875:Ubiquitination / Q000502:physiology*; D029867:Xenopus Proteins; D014982:Xenopus laevis
D056827:Endosomal Sorting Complexes Required for Transport; D051657:KCNQ1 Potassium Channel; D025801:Ubiquitin; D029867:Xenopus Proteins; D000075702:Nedd4 Ubiquitin Protein Ligases; C000618482:Nedd4 protein, Xenopus; C000618460:nedd4l protein, Xenopus; D044767:Ubiquitin-Protein Ligases; D010450:Endopeptidases; D064570:Ubiquitin-Specific Proteases