Alport syndrome

ABSTRACT The function of the KCNE5 (KCNE1-like) protein has not previously been described. Here we show that KCNE5 induces both a time- and voltage-dependent modulation of the KCNQ1 current. Interaction of the KCNQ1 channel with KCNE5 shifted the voltage activation curve of KCNQ1 by more than 140 mV in the positive direction. The activation threshold of the KCNQ1 +KCNE5 complex was +40 mV and the midpoint of activation was +116 mV. The KCNQ1+KCNE5 current activated slowly and deactivated rapidly as compared to the KCNQ1+KCNE1 at 22 degC; however, at physiological temperature, the activation time constant of the KCNQ1 +KCNE5 current decreased fivefold, thus exceeding the activation rate of the KCNQ1+KCNE1 current. The KCNE5 subunit is specific for the KCNQ1 channel, as none of other members of the KCNQ-family or the human ether a-go-go related channel (hERG1) was affected by KCNE5. Four residues in the transmembrane domain of the KCNE5 protein were found to be important for the control of the voltage-dependent activation of the KCNQ1 current. We speculate that since KCNE5 is expressed in cardiac tissue it may here along with the KCNE1 beta-subunit regulate KCNQ1 channels. It is possible that KCNE5 shapes the /^sub Ks^ current in certain parts of the mammalian heart.

INTRODUCTION

It has become clear that channels are composed of numerous units, which engage into larger complexes. Often the pore-forming alpha-subunits interact with other membrane proteins and/or couple to cytosolic components that contribute to the features of the channel. Some accessory proteins change the sensitivity of the alpha-subunit to e.g., Ca^sup 2+^, pH, temperature, cell volume changes, or second messengers. They may direct the membrane localization of channels, couple channels to the cytoskeleton, or regulate the expression level of channels. Basic channel properties such as selectivity, conductance, voltage-dependency and pharmacological profile can be changed by these subunit-interactions to an extent where the biophysical characteristics of the alpha-subunit become difficult to recognize. Several important native currents have been proven to be composed of such subunits interacting in a non-trivial fashion (Seino, 1999; Peleg et al., 2002).

The KCNE-family is a group of small proteins, which interact with voltage-gated potassium channels. The length of the KCNE proteins is between 130 and 167 amino acids and they contain one transmembrane domain flanked by an extracellular N-terminal and a cytosolic C-terminal. The first member of the family, KCNE1 (minK), was cloned in 1988 (Takumi et al., 1988) and in 1999, Abbott et al. isolated KCNE2, 3 and 4 (MiRP1-3) (Abbott et al., 1999). The functional effect of KCNE on the KCNQ1 channel is that the current activates and deactivates slower than when the KCNQI alpha-subunit is alone. The voltage sensitivity of the KCNQ1+KCNE1 current is shifted toward positive potentials and the whole-cell current density is increased. Furthermore, assembly of KCNQI with the KCNE1 subunit increases the unitary conductance by fourfold (Sesti and Goldstein, 1998). The current formed by the KCNQ1 +KCNE1 complex corresponds to the slow delayed rectifier current, I^sub Ks^, in cardiac myocytes (Sanguinetti et al., 1996; Barhanin et al., 1996). An interaction between KCNE3 and KCNQI gives rise to a constitutively open potassium channel that plays a role in cyclic AMP-stimulated C1^sup -^ secretion (Schroeder et al., 2000; Grahammer et al., 2001). Expression of KCNQ1 together with KCNE2 in COS cells results in an effect similar to that of the KCNE3; the KCNQ1 channel transforms into a voltage-- independent channel (Tinel et al., 2000a).

In 1999 Piccini et al. isolated a gene, which they refer to as the KCNE1-like gene. The KCNE1-like gene is one of the four genes that are deleted in the AMME contiguous gene syndrome (Jonsson et al., 1998). The human KCNE1-like protein shows 56% homology with KCNE1. It is composed of 142 amino acids and like the other members of the KCNE-family it has a single putative transmembrane domain. The KCNE1-like gene has lately been suggested to be the fifth member of the KCNE family and is now referred to as KCNE5 (Abbott et al., 2001). To date the function of the KCNE5 protein has not been established. Here we demonstrate that the KCNQI current is markedly influenced by the presence of KCNE5. We show that the KCNE5 beta-subunit affects the activation kinetics of the KCNQ1 current in the same direction as observed for the KCNE1+KCNQI complex; however, to a much greater extend. The voltage-activation curve of the KCNQ I current is shifted in the positive direction toward an activation threshold of +40 mV. Thus, the KCNQ1 +KCNE5 complex only conducts current upon strong and continued depolarization. The effect of KCNE5 was specific for KCNQ1 channels; the other KCNQ channels or the human ether a-go-go related channel (hERG1) was not affected by coexpressed KCNE5. Mutagenesis experiments showed that four specific amino acids in the transmembrane domain of KCNE5 are responsible for the regulatory effect.

We thank Inge Kjeldsen and Pia Hageman for technical assistance. This study was supported by the Danish Heart Association.

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Kamilla Angelo, Thomas Jespersen, Morten Grunnet, Morten Schak Nielsen, Dan A. Klaerke, and Soren-Peter Olesen

Department of Medical Physiology, University of Copenhagen, The Panum Institute, Copenhagen, Denmark

Submitted March 12, 2002, and accepted for publication May 30, 2002.

Address reprint requests to Kamilla Angelo, Dept. of Medical Physiology, University of Copenhagen, The Panum Inst, Blegdamsvej 3, Copenhagen 2200 N, Denmark. Tel.: 45 35327445; Fax: 45 35327555; E-mail: angelo@mfi.ku.dk.

Copyright Biophysical Society Oct 2002
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