Roles of macromolecular complexes in calcium-sensitivity of the cardiac I_Ks channel
Cardiac repolarization process is regulated by the slow component of the delayed rectifier potassium (I_Ks) channel composed of Ca²⁺-sensitive KCNQ1 and KCNE1. Although calmodulin and AKAP9 regulate I_Ks channel for instance, the entire macromolecular system has not been elucidated. Thus, to seek macromolecular complexes comprehensively, we performed proteomics analysis using a transgenic mouse overexpressing KCNQ1-KCNE1 fusion protein specifically in the heart. We identified 163 proteins as potential I_Ks binding partners by immunoprecipitation using a goat anti-KCNQ1 antibody. Pathway analysis of these proteins revealed that the most significant biofunction is the Calcium Signaling including 16 molecules. To understand how the protein-protein interactions affect Ca²⁺-dependent I_Ks regulation, we picked up the cardiac Na⁺-Ca²⁺ exchange transporter (NCX1). Pull-down assays in dog ventricles with purified GST-fusion proteins of full-length or parts of KCNQ1 identified that the N- and a proximal C-terminus of KCNQ1 contribute the interaction. These results imply that the Ca²⁺-sensitivity of I_Ks channel is regulated by the interaction between the I_Ks channel and NCX1. Pharmacological experiments will be performed to pursue its mechanistic insights.