Functional impact of the interaction between S1 segment and KCNE subunit in the modulation of KCNQ1 channels

KCNQ1 channel is a voltage-gated potassium channel broadly expressed in the human body, and KCNE proteins are modulatory subunits for the KCNQ1 channel. Among five members of the KCNE subfamily, at least KCNE1 and KCNE3 are considered to modulate the KCNQ1 channel through the voltage sensor domain (VSD), probably by stabilizing its intermediate state. However, the mechanisms of how the KCNE subunits interact and affect the VSD movements are still not fully understood. The recent cryo-EM structure of the KCNQ1-KCNE3 complex shows close interaction between the S1 segment and KCNE3 throughout the transmembrane region. Therefore, we hypothesized that the KCNE subunit controls the VSD equilibrium by interacting with the S1 segment. To test that, we mutated seven amino acid residues on the S1 (F123, F127, F130, L134, I138, L142, and I145) facing the KCNE3 subunit to four different hydrophobic amino acid residues with various sizes for each (“volume scanning”). The KCNE3 modulation depended on the size of introduced amino acid residues at most sites: The more different the size of amino acid residue from the wild-type, the more constitutive openness was weakened. A similar tendency was also observed in KCNE3 mutants: we introduced five or six different amino acid residues to each of the KCNE3 amino acid residues facing the S1 (S57, I61, M65, A69, G73, and I76), and the KCNE3 modulation depended on the size of introduced amino acid residues. Finally, we found two volume swap pairs (KCNQ1 F127A-KCNE3 G73L; KCNQ1 I145F-KCNE3 S57A), which restored the normal function (stabilizing the VSD at the intermediate state) of the KCNQ1-KCNE3 channels. These results indicate that the interaction between the S1 and KCNE3 is tightly optimized to regulate the VSD equilibrium.