Regulation of sinoatrial myocyte pacemaker function by a novel HCN4 interacting protein

University Of Colorado Denver, Aurora CO

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Abstract

? DESCRIPTION (provided by applicant): Control of heart rate, both up and down, is clinically important. Increasing heart rate via surgically-implanted electronic pacemakers is beneficial for many elderly patients, whereas decreasing heart rate via sinus node inhibitors is advantageous for patients suffering angina and heart failure. However these treatments have significant limitations. Heart rate is controlled by the pacemaker activity of specialized myocytes in the sinoatrial node. Sinoatrial myocytes (SAMs) generate spontaneous action potentials (APs) via spontaneous depolarization of the membrane during diastole. The funny current (If) is critical for pacemaker activity in SAMs and is produced by hyperpolarization-activated, cyclic nucleotide sensitive (HCN) channels, with HCN4 being the predominant isoform in the heart. If is a target for emerging treatments for controlling heart rate, however little is known about the molecular mechanisms by which HCN4 channels are modulated. We recently discovered lymphoid-restricted membrane protein (LRMP) as a novel HCN4-associated protein that can profoundly alter the cAMP sensitivity and voltage-dependence of If in heterologous expression systems. LRMP was previously identified in a genome wide association study as being in close proximity to a gene locus related to resting heart rate7, and knockout in mice of a longer homolog has been shown to reduce heart rate8. Furthermore, we recently demonstrated expression of LRMP in the sinoatrial node. These data support a possible role for LRMP in heart rate regulation via modulation of If in SAMs, and could suggest that it could be a novel therapeutic target for heart rate control. The central goals of this project are to define the functional interactions between LRMP and HCN4 channels, and to determine how LRMP modulates If and pacemaker activity in SAMs. In Specific Aim 1, I will test the hypothesis that there is a direct interaction between LRMP and HCN4 using protein affinity chromatography, site-directed mutagenesis and patch clamp electrophysiology. In Specific Aim 2, I will use siRNA to knockdown expression of LRMP in cultured SAMs, and will perform patch clamp electrophysiology to determine if LRMP modulates the cAMP sensitivity and voltage- dependence of If in SAMs. Successful completion of these Aims will yield novel information about modulation of HCN4 by LRMP, and could provide the framework for development of better therapeutic strategies to control heart rate.