Unnatural amino acid photo-crosslinking of the IKs channel complex demonstrates a KCNE1:KCNQ1 stoichiometry of up to4:4.

Christopher I Murray,Maartje Westhoff,Robert Emes,Emely Thompson,David Fedida,Jodene Eldstrom

doi:10.7554/elife.11815

Abstract

Cardiac repolarization is determined in part by the slow delayed rectifier current (IKs), through the tetrameric voltage-gated ion channel, KCNQ1, and its β-subunit, KCNE1. The stoichiometry between α and β-subunits has been controversial with studies reporting either a strict 2 KCNE1:4 KCNQ1 or a variable ratio up to 4:4. We used IKs fusion proteins linking KCNE1 to one (EQ), two (EQQ) or four (EQQQQ) KCNQ1 subunits, to reproduce compulsory 4:4, 2:4 or 1:4 stoichiometries. Whole cell and single-channel recordings showed EQQ and EQQQQ to have increasingly hyperpolarized activation, reduced conductance, and shorter first latency of opening compared to EQ - all abolished by the addition of KCNE1. As well, using a UV-crosslinking unnatural amino acid in KCNE1, we found EQQQQ and EQQ crosslinking rates to be progressively slowed compared to KCNQ1, which demonstrates that no intrinsic mechanism limits the association of up to four β-subunits within the IKs complex.

Highlights

Kv7.1 (KCNQ1) is a voltage gated potassium channel expressed throughout the body
In addition to their role in the heart, KCNQ1 and KCNE1 are involved in hearing, where mutations have been linked with Jervell and Lange-Nielsen syndrome, an inherited form of long QT (LQT) syndrome accompanied by deafness (Jervell and Lange-Nielsen, 1957; Neyroud et al, 1997; Schulze-Bahr et al, 1997)
The C-terminus of KCNE1 was connected with the N-terminus of KCNQ1 via a flexible linker (EQ), forcing a 4:4 stoichiometry of b and a subunits

Summary

Introduction

Kv7.1 (KCNQ1) is a voltage gated potassium channel expressed throughout the body. When paired with the accessory b-subunit, KCNE1, it has unique properties in several tissues, the myocardium and inner ear. KCNQ1 and KCNE1 together conduct the slow delayed rectifier current (IKs). Several inherited mutations in KCNQ1 or KCNE1 result in the life-threating arrhythmogenic syndromes long QT (LQT) types 1 and 5, short QT type 2 or familial atrial fibrillation (Splawski et al, 2000;Bellocq et al, 2004; Chen et al, 2003). In addition to their role in the heart, KCNQ1 and KCNE1 are involved in hearing, where mutations have been linked with Jervell and Lange-Nielsen syndrome, an inherited form of LQT syndrome accompanied by deafness (Jervell and Lange-Nielsen, 1957; Neyroud et al, 1997; Schulze-Bahr et al, 1997). KCNQ1 and KCNE1 complexes have been found in the proximal tubule of the kidney, where they participate in secretory transduction (Vallon et al, 2001)

Methods

Results

Discussion

Conclusion