Stoichiometry of KCNQ1-KCNE1 Ion Channel Complex is Flexible and Density-Dependent

Abstract

Many membrane proteins including ion channels form multi-molecular complexes. Because the composition of a molecular complex may define its functional properties, it is important to know its stoichiometry. KCNQ1 encodes a voltage-gated potassium channel alpha subunit, and four KCNQ1 subunits form one ion channel. KCNQ1 channel forms a molecular complex with auxiliary subunit KCNE proteins. In the heart the KCNQ1-KCNE1 complex underlies slowly-activating IKs current, which plays a significant role in regulation of the cardiac action potential. Assuming a fixed KCNQ1-KCNE1 stoichiometry macroscopic current measurements led earlier investigators to the conclusion that each 4-subunit channel is associated with two KCNE1 subunits (4:2 subunit stoichiometry). We asked whether the KCNQ1-KCNE1 stoichiometry is indeed fixed by counting subunits in many individual complexes using TIRF microscopy (Ulbrich and Isacoff, 2007, 2008). We expressed GFP-tagged KCNQ1 or KCNE1 in Xenopus oocytes at low density and counted bleaching steps in many fluorescent spots corresponding to single channel complexes. First, we confirmed that KCNQ1 forms a tetramer. Next we counted GFP-tagged KCNE1 subunits co-expressed with mCherry-tagged KCNQ1. We observed up to four bleaching steps from GFP-KCNE1 co-localized with mCherry, indicating that up to four KCNE1 subunits can bind to one KCNQ1 tetrameric channel. We find that the number of KCNE1 subunits per complex increases as the expression of KCNE1 is raised relative to that of KCNQ1. Our results suggest that modulation of KCNQ channels may be regulated by the level of expression of KCNE subunits.