Allosteric Modulation of HCN Channels

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PROJECT SUMMARY Hyperpolarization-activated, cyclic nucleotide-sensitive (HCN) channels are critical determinants of membrane potential and excitability in many types of cells throughout the body, including cardiac pacemaker cells, central and peripheral neurons, many types of sensory cells, and interstitial cells of Cajal in the colon and bladder. Consistent with this widespread distribution, HCN channels have been identified as potential drug targets for treatment of a long list of conditions including angina, heart failure, epilepsy, neuropathic pain, depression, gastrointestinal dysmotility, and neurogenic bladder. However, the single FDA-approved HCN channel drug (ivabradine) is limited, owing in part to its non-selective block of all four mammalian HCN channels isoforms and its off-target block of Kv11.1 (hERG), Nav1.5, and Cav1.2 channels. The need for new, isoform-specific HCN channel activators and inhibitors has been widely recognized but the lack of information about allosteric and isoform-specific regulation of HCN channels is a roadblock to the development of novel therapeutics. The long-term goals of this project are to identify naturally-occurring, allosteric regulators of HCN channels and to understand their mechanisms of action. Achieving these goals will advance understanding of the physiological and molecular functions of HCN channels and aid in the development of new HCN channel drugs. The current proposal focuses on our exciting discovery of LRMP and IRAG as two novel, isoform- specific protein interaction partners of HCN4 channels. LRMP and IRAG are homologous ER transmembrane proteins that have large cytoplasmic domains. Importantly, LRMP and IRAG only modulate the HCN4 isoform. Moreover, the two proteins have opposing effects on HCN4: LRMP causes a loss-of-function (LOF) by decreasing the canonical depolarizing shift in voltage-dependence induced by cAMP while IRAG causes gain- of-function (GOF) by shifting the basal voltage dependence of HCN4 to more positive potentials. Preliminary data establish that IRAG is co-expressed with HCN4 in cardiac pacemaker cells. Proposed aims will identify interaction sites on the three proteins, determine the molecular mechanisms for the distinct and isoform- specific effects, and evaluate their role in pacemaker cells.