Abstract
cAMP mediates autonomic regulation of heart rate by means of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which underlie the pacemaker current If. cAMP binding to the C-terminal cyclic nucleotide binding domain enhances HCN open probability through a conformational change that reaches the pore via the C-linker. Using structural and functional analysis, we identified a binding pocket in the C-linker of HCN4. Cyclic dinucleotides, an emerging class of second messengers in mammals, bind the C-linker pocket (CLP) and antagonize cAMP regulation of the channel. Accordingly, cyclic dinucleotides prevent cAMP regulation of If in sinoatrial node myocytes, reducing heart rate by 30%. Occupancy of the CLP hence constitutes an efficient mechanism to hinder β-adrenergic stimulation on If. Our results highlight the regulative role of the C-linker and identify a potential drug target in HCN4. Furthermore, these data extend the signaling scope of cyclic dinucleotides in mammals beyond their first reported role in innate immune system.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Binding Sites
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Blotting, Western
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Crystallography, X-Ray
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Cyclic AMP / metabolism*
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Cyclic GMP / analogs & derivatives*
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Cyclic GMP / chemistry
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Cyclic GMP / metabolism
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Dinucleoside Phosphates / chemistry
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Dinucleoside Phosphates / metabolism*
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HEK293 Cells
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High-Throughput Screening Assays
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Humans
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Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels / genetics
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Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels / metabolism*
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Ion Channel Gating / drug effects
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Ion Channel Gating / physiology*
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Ligands
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Mice
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Mice, Inbred C57BL
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Molecular Docking Simulation
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Molecular Structure
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Muscle Proteins / genetics
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Muscle Proteins / metabolism*
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Myocytes, Cardiac / drug effects
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Myocytes, Cardiac / metabolism
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Patch-Clamp Techniques
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Potassium Channels / genetics
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Potassium Channels / metabolism*
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Sinoatrial Node / cytology
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Sinoatrial Node / drug effects
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Sinoatrial Node / metabolism
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Small Molecule Libraries / chemistry
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Small Molecule Libraries / pharmacology
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Transfection
Substances
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Dinucleoside Phosphates
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HCN4 protein, human
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Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels
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Ligands
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Muscle Proteins
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Potassium Channels
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Small Molecule Libraries
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cyclic diadenosine phosphate
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bis(3',5')-cyclic diguanylic acid
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Cyclic AMP
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Cyclic GMP
Associated data
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PDB/4KL1
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PubChem-Substance/175441889
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PubChem-Substance/175441890
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PubChem-Substance/175441891
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PubChem-Substance/175441892
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PubChem-Substance/175441893
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PubChem-Substance/175441894
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PubChem-Substance/175441895
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PubChem-Substance/175441896
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PubChem-Substance/175441897
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PubChem-Substance/175441898
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PubChem-Substance/175441899