Science Topics – 182
The hyperpolarization-activated cyclic nucleotide-gated (HCN) channel is a voltage-dependent ion channel abundantly expressed in cardiac pacemaker cells and neurons. Dysfunction of HCN channels can lead to cardiac arrhythmias such as bradycardia, as well as neurological disorders. HCN channels open upon hyperpolarization, during which the voltage sensor S4 moves downward.
Recent cryo-electron microscopy studies have revealed that the S4 segment of HCN4 extends toward the extracellular side. Although the contribution of this elongated extracellular S4 region to channel gating remains unclear, previous studies have shown that mutations in positively charged residues within this region (e.g., R375C and R378C) are associated with sick sinus syndrome and bradycardia.
In this study, we investigated the functional role of salt bridges formed between the extracellular portion of S4, the S5 segment, and the S1–S2 linker. Using charge-reversal and double-cysteine mutants, we demonstrated that positively charged residues in the extracellular S4 region interact stepwise with negatively charged residues in S5 and the S1–S2 linker to form a salt bridge network. Voltage-clamp fluorometry confirmed that this network influences S4 movement. Mutations disrupting the network impaired channel function and may contribute to disease pathogenesis.
Our findings reveal that the extracellular portion of the S4 segment plays a critical role in the voltage-dependent gating of HCN channels. Disruption of this salt bridge network may underlie cardiac and neurological disorders, suggesting a potential therapeutic target for future interventions.
Extracellular salt bridge networks around S4 implicated in HCN channel gating and heart disease. Kaei Ryu, Go Kasuya, Koichi Nakajo. Proceedings of the National Academy of Sciences of the United States of America 122(37): e2502136122, 2025
＜Figure Legends＞
(A) HCN4 Channel Structure and Salt Bridge Formation
Side and extracellular views of the cryo-EM structure of the HCN4 channel (PDB: 6GYO) are shown. In this structure, positively charged residues R378 and K381 on the extracellular side form salt bridges with negatively charged residues D444 (in the S5 segment) and E290 (in the S2 segment), respectively.
(B) Salt Bridge Transition During Hyperpolarization Gating
Upon membrane hyperpolarization, the S4 segment moves downward, leading to channel opening. Mutations at residues such as R375 or R378 disrupt the salt bridge network, impairing the smooth movement of the S4 segment. This disruption results in channel dysfunction and may contribute to disorders such as bradycardia or sick sinus syndrome.
Division of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Jichi Medical University, Japan