K+ channel complexes: assembly, trafficking and function

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DESCRIPTION (provided by applicant): The long-term goal of this research is to elucidate the molecular and cellular mechanisms that ensure potassium channels assemble with the appropriate membrane-embedded partnering proteins for proper physiological function. The KCNQ1 K+ channel is a key player in many areas of human physiology from repolarization of the cardiac action potential to salt and water transport in epithelia cells. In order to generate the diverse set of K+ currents needed for these varied physiological roles, KCNQ1 channels must form membrane-embedded complexes with the KCNE-family of transmembrane peptides. Genetic mutations in either the KCNQ1 channel or the KCNE peptide that perturb complex function or reduce its cell surface expression cause cardiac arrhythmias and deafness. This proposal is directed at determining the basic mechanisms of KCNQ1-KCNE assembly and function. There are two aims to this proposal: (1) we will identify the molecular determinants required for KCNQ1-KCNE assembly and function using mutagenic perturbation analysis and cysteine-specific chemistries in combination with electrophysiological measurements; (2) we will determine whether the cellular stability and trafficking rates of the KCNQ1-KCNE1 complex are the cellular mechanisms responsible for their preferential cell surface expression. For this aim, we will metabolically and chemically label the proteins and determine their cellular sites of assembly and compartmentalization using subcellular membrane fractionation. The results from these aims will provide a molecular and cellular framework to aid in the understanding of KCNE-related diseases as well as providing potential targets for new therapies. Furthermore, as the list of K+ channels that assemble with KCNE peptides grows, these results will be applicable to all electrically excitable cells from neurons to muscle.