Cardiovascular Research Advance Access [Accepted Manuscript] published online on February 7, 2009
Cardiovascular Research, doi:10.1093/cvr/cvp047
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MinK-dependent internalization of the IKs potassium channel
1 Greenberg Division of Cardiology, Dept. of Medicine
2 Dept. of Pharmacology, Weill Medical College of Cornell University, New York, NY
3 CV Ingenuity, San Francisco, CA
To whom correspondence should be addressed: Dr. Geoffrey W. Abbott, Starr 463, Greenberg Division of Cardiology, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY, 10065. Tel: 212 7466275; Fax: 212 7467984; email: gwa2001{at}med.cornell.edu
Aims: KCNQ1-MinK potassium channel complexes (4
:2β stoichiometry) generate IKs, the slowly-activating human cardiac ventricular repolarization current. The MinK ancillary subunit slows KCNQ1 activation, eliminates its inactivation and increases its unitary conductance. However, KCNQ1 transcripts outnumber MinK transcripts five-to-one in human ventricles, suggesting KCNQ1 also forms other heteromeric or even homomeric channels there. Mechanisms governing which channel types prevail have not previously been reported, despite their significance: normal cardiac rhythm requires tight control of IKs density and kinetics, and inherited mutations in KCNQ1 and MinK can cause ventricular fibrillation and sudden death. Here, we describe a novel mechanism for this control.
Methods: Whole-cell patch-clamping, confocal immunofluorescence microscopy, antibody feeding, biotin feeding, fluorescent transferrin feeding and protein biochemistry techniques were applied to COS-7 cells heterologously expressing KCNQ1 with wild-type or mutant MinK and dynamin 2, and to native IKs channels in guinea-pig myocytes.
Results: KCNQ1-MinK complexes, but not homomeric KCNQ1 channels, were found to undergo clathrin- and dynamin 2-dependent internalization (DDI). Three sites on the MinK intracellular C-terminus were, in concert, necessary and sufficient for DDI. Gating kinetics and sensitivity to XE991 indicated that DDI decreased cell-surface KCNQ1-MinK channels relative to homomeric KCNQ1, decreasing whole-cell current but increasing net activation rate; inhibiting DDI did the reverse.
Conclusions: The data redefine MinK as an endocytic chaperone for KCNQ1 and present a dynamic mechanism for controlling net surface Kv channel subunit composition - and thus current density and gating kinetics - that may also apply to other -β type Kv channel complexes.
Time for primary review: 18 Days
* these authors contributed equally to this study
¶ present address: Charite Campus Berlin-Buch, Experimental and Clinical Research Center, Franz Volhard Clinic and HELIOS Klinikum, Berlin-Buch, Berlin, Germany
Abbreviations: CME, clathrin-mediated endocytosis; DDI, dynamin-dependent internalization; EEA1, early endosomal antigen 1; GPCR, G-protein coupled receptor; HA, hemaglutinin; IP, immunoprecipitate; Kv channel, voltage-gated potassium channel; MiRP, MinK-related peptide; TCL, total cell lysate.
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Cardiovasc Res 2009 82: 390-391.
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