期刊
CHANNELS
卷 14, 期 1, 页码 268-286出版社
TAYLOR & FRANCIS INC
DOI: 10.1080/19336950.2020.1805999
关键词
Calmodulin; sodium channels; SCN5A; Na(v)1; 5 N-terminal domain; Brugada syndrome; dominant-negative effect
资金
- Schweizerischer Nationalfonds zur Forderung der Wissenschaftlichen Forschung [310030_165741]
The cardiac voltage-gated sodium channel Na(v)1.5 conducts the rapid inward sodium current crucial for cardiomyocyte excitability. Loss-of-function mutations in its geneSCN5Aare linked to cardiac arrhythmias such as Brugada Syndrome (BrS). Several BrS-associated mutations in the Na(v)1.5 N-terminal domain (NTD) exert a dominant-negative effect (DNE) on wild-type channel function, for which mechanisms remain poorly understood. We aim to contribute to the understanding of BrS pathophysiology by characterizing three mutations in the Na(v)1.5 NTD: Y87C-here newly identified-, R104W, and R121W. In addition, we hypothesize that the calcium sensor protein calmodulin is a new NTD binding partner. Recordings of whole-cell sodium currents in TsA-201 cells expressing WT and variant Na(v)1.5 showed that Y87C and R104W but not R121W exert a DNE on WT channels. Biotinylation assays revealed reduction in fully glycosylated Na(v)1.5 at the cell surface and in whole-cell lysates. Localization of Na(v)1.5 WT channel with the ER did not change in the presence of variants, as shown by transfected and stained rat neonatal cardiomyocytes. We demonstrated that calmodulin binds the Na(v)1.5 NTD usingin silicomodeling, SPOTS, pull-down, and proximity ligation assays. Calmodulin binding to the R121W variant and to a Na(v)1.5 construct missing residues 80-105, a predicted calmodulin-binding site, is impaired. In conclusion, we describe the new natural BrS Na(v)1.5 variant Y87C and present first evidence that calmodulin binds to the Na(v)1.5 NTD, which seems to be a determinant for the DNE.
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