Riedersberger Maria, Woltereck Madalina, Wagner Paul J, Focke Kristin, Höller Sarina, Lampert Angelika, Alzheimer Christian, Düsterhöft Stefan, Huth Tobias
Institut für Physiologie und Pathophysiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
Institut für Neurophysiologie, Universitätsklinikum RWTH Aachen University, Aachen, Germany.
Biophys J. 2025 Jul 15;124(14):2263-2279. doi: 10.1016/j.bpj.2025.05.030. Epub 2025 Jun 2.
Resurgent sodium currents (INaRs) result from an unorthodox gating behavior of voltage-activated sodium channels (Na), allowing transient re-openings during repolarization from an apparently inactivated state. In both native cells not normally exhibiting INaR and in heterologous expression systems, intracellular delivery of small positively charged peptides through the recording pipette elicits robust INaRs, suggesting that INaRs arise from a peptide-mediated open-channel block that is relieved upon repolarization. Here we examined the hNa1.5 ΔKPQ mutant, which causes a long QT syndrome in the heart, to probe the open-channel block hypothesis of INaR in a channel with altered inactivation properties due to the deletion near the canonical inactivation gate. We investigated INaRs with peptides derived from the hNaβ4 subunit, FGF13-1a and FGF14-1a, using the HEK293T expression system. Surprisingly, the peptides not only gave rise to pronounced INaRs with unusually fast kinetics but also altered the late sodium current of the channel mutant. To elucidate the molecular basis of these effects, we employed AlphaFold modeling of hNa1.5, incorporating the ΔKPQ mutation and the β4 peptide. This model supports the open-channel block mechanism of INaR and its mutual exclusivity with fast inactivation. It also demonstrates a lack of interaction between the IFM linker and the C-terminal domain in hNa1.5 ΔKPQ, offering a plausible explanation for why the peptides are capable of affecting both INaR and persistent currents (INaPs). Finally, the peptides generated a considerable increase in repolarization-associated Na currents with the ΔKPQ mutant, highlighting the presumed impact of pathologically enhanced INaR on cardiac electrophysiology.
复苏钠电流（INaRs）源于电压激活钠通道（Na）的非传统门控行为，允许在从明显失活状态复极化期间短暂重新开放。在通常不表现出INaR的天然细胞和异源表达系统中，通过记录微电极向细胞内递送带正电的小肽可引发强大的INaRs，这表明INaRs源于肽介导的开放通道阻滞，而复极化时该阻滞得以解除。在这里，我们研究了hNa1.5 ΔKPQ突变体，该突变体在心脏中会导致长QT综合征，以探究在一个由于经典失活门附近缺失而具有改变的失活特性的通道中INaR的开放通道阻滞假说。我们使用HEK293T表达系统，研究了源自hNaβ4亚基、FGF13 - 1a和FGF14 - 1a的肽对INaRs的影响。令人惊讶的是，这些肽不仅产生了具有异常快速动力学的明显INaRs，还改变了通道突变体的晚期钠电流。为了阐明这些效应的分子基础，我们采用了hNa1.5的AlphaFold建模，纳入了ΔKPQ突变和β4肽。该模型支持INaR的开放通道阻滞机制及其与快速失活的相互排斥性。它还表明在hNa1.5 ΔKPQ中IFM连接子与C末端结构域之间缺乏相互作用，这为肽为何能够同时影响INaR和持续性电流（INaPs）提供了一个合理的解释。最后，这些肽使ΔKPQ突变体的复极化相关钠电流显著增加，突出了病理性增强的INaR对心脏电生理学的假定影响。
