Abstract：
BACKGROUND: Pathogenic variants in SCN5A, the gene encoding the cardiac Na+ channel α-subunit Nav1.5, result in life-threatening arrhythmias, e.g., Brugada syndrome, cardiac conduction defects and long QT syndrome. This variety of phenotypes is underlied by the fact that each Nav1.5 mutation has unique consequences on the channel trafficking and gating capabilities. Recently, we established that sodium channel α-subunits Nav1.5, Nav1.1 and Nav1.2 could dimerize, thus, explaining the potency of some Nav1.5 pathogenic variants to exert dominant-negative effect on WT channels, either by trafficking deficiency or coupled gating.
OBJECTIVE: The present study sought to examine whether Nav1.5 channels can cooperate, or transcomplement each other, to rescue the Na+ current (INa). Such a mechanism could contribute to explain the genotype-phenotype discordance often observed in family members carrying Na+-channel pathogenic variants.
METHODS: Patch-clamp and immunocytochemistry analysis were used to investigate biophysical properties and cellular localization in HEK293 cells and rat neonatal cardiomyocytes transfected respectively with WT and 3 mutant channels chosen for their particular trafficking and/or gating properties.
RESULTS: As previously reported, the mutant channels G1743R and R878C expressed alone in HEK293 cells both abolished INa, G1743R through a trafficking deficiency and R878C through a gating deficiency. Here, we showed that coexpression of both G1743R and R878C nonfunctioning channels resulted in a partial rescue of INa, demonstrating a cooperative trafficking of Nav1.5 α-subunits. Surprisingly, we also showed a cooperation mechanism whereby the R878C gating-deficient channel was able to rescue the slowed inactivation kinetics of the C-terminal truncated R1860X (ΔCter) variant, suggesting coupled gating.
CONCLUSIONS: Altogether, our results add to the evidence that Nav channels are able to interact and regulate each other\'s trafficking and gating, a feature that likely contributes to explain the genotype-phenotype discordance often observed between members of a kindred carrying a Na+-channel pathogenic variant.
OBJECTIVE: The present study sought to examine whether Nav1.5 channels can cooperate, or transcomplement each other, to rescue the Na+ current (INa). Such a mechanism could contribute to explain the genotype-phenotype discordance often observed in family members carrying Na+-channel pathogenic variants.
METHODS: Patch-clamp and immunocytochemistry analysis were used to investigate biophysical properties and cellular localization in HEK293 cells and rat neonatal cardiomyocytes transfected respectively with WT and 3 mutant channels chosen for their particular trafficking and/or gating properties.
RESULTS: As previously reported, the mutant channels G1743R and R878C expressed alone in HEK293 cells both abolished INa, G1743R through a trafficking deficiency and R878C through a gating deficiency. Here, we showed that coexpression of both G1743R and R878C nonfunctioning channels resulted in a partial rescue of INa, demonstrating a cooperative trafficking of Nav1.5 α-subunits. Surprisingly, we also showed a cooperation mechanism whereby the R878C gating-deficient channel was able to rescue the slowed inactivation kinetics of the C-terminal truncated R1860X (ΔCter) variant, suggesting coupled gating.
CONCLUSIONS: Altogether, our results add to the evidence that Nav channels are able to interact and regulate each other\'s trafficking and gating, a feature that likely contributes to explain the genotype-phenotype discordance often observed between members of a kindred carrying a Na+-channel pathogenic variant.
摘要：
背景：SCN5A的致病变异，编码心脏Na+通道α亚基的基因Nav1.5，导致危及生命的心律失常，例如，Brugada综合征,心脏传导缺陷和长QT综合征。每个Nav1.5突变对通道运输和门控能力具有独特后果的事实为这种表型的多样性奠定了基础。最近,我们确定钠通道α亚基Nav1.5，Nav1.1和Nav1.2可以二聚化，因此，解释了一些Nav1.5致病变体对WT通道产生显性负效应的效力，要么是贩运不足，要么是配对门控。
目的：本研究试图检查Nav1.5通道是否可以合作，或彼此互补，拯救Na+电流(INa)。这种机制可能有助于解释在携带Na通道致病变体的家族成员中经常观察到的基因型-表型不一致。
方法:使用膜片钳和免疫细胞化学分析来研究分别用WT和3种突变通道转染的HEK293细胞和大鼠新生心肌细胞中的生物物理特性和细胞定位,所述突变通道根据其特定运输和/或门控特性选择。
结果：如先前报道，在HEK293细胞中单独表达的突变通道G1743R和R878C均消除了INa，G1743R通过贩运缺陷和R878C通过门控缺陷。这里,我们显示G1743R和R878C无功能通道的共表达导致INa的部分挽救，展示了Nav1.5α亚基的合作贩运。令人惊讶的是,我们还显示了一种合作机制，即R878C门控缺陷通道能够挽救C末端截短的R1860X（ΔCter）变体的缓慢失活动力学，建议耦合门控。
结论：总而言之，我们的结果增加了Nav渠道能够相互作用和调节彼此的贩运和门控的证据，该特征可能有助于解释经常在携带Na通道致病变体的亲属成员之间观察到的基因型-表型不一致。
目的：本研究试图检查Nav1.5通道是否可以合作，或彼此互补，拯救Na+电流(INa)。这种机制可能有助于解释在携带Na通道致病变体的家族成员中经常观察到的基因型-表型不一致。
方法:使用膜片钳和免疫细胞化学分析来研究分别用WT和3种突变通道转染的HEK293细胞和大鼠新生心肌细胞中的生物物理特性和细胞定位,所述突变通道根据其特定运输和/或门控特性选择。
结果：如先前报道，在HEK293细胞中单独表达的突变通道G1743R和R878C均消除了INa，G1743R通过贩运缺陷和R878C通过门控缺陷。这里,我们显示G1743R和R878C无功能通道的共表达导致INa的部分挽救，展示了Nav1.5α亚基的合作贩运。令人惊讶的是,我们还显示了一种合作机制，即R878C门控缺陷通道能够挽救C末端截短的R1860X（ΔCter）变体的缓慢失活动力学，建议耦合门控。
结论：总而言之，我们的结果增加了Nav渠道能够相互作用和调节彼此的贩运和门控的证据，该特征可能有助于解释经常在携带Na通道致病变体的亲属成员之间观察到的基因型-表型不一致。
