A1152D mutation of the Na+ channel causes paramyotonia congenita and emphasizes the role of DIII/S4-S5 linker in fast inactivation
A1152D mutation of the Na+ channel causes paramyotonia congenita and emphasizes the role of DIII/S4-S5 linker in fast inactivation
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DOI：
10.1113/jphysiol.2004.081018
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发表时间：
2005-06-01
影响因子：
5.5
通讯作者：
Tabti, N
中科院分区：
文献类型：
--
作者：
Bouhours, M;Luce, S;Tabti, N
人骨骼肌Na+通道α亚基（hSkM 1）中的错义突变导致许多肌肉兴奋性疾病。其中，先天性副肌强直（PC）的特点是由寒冷引起的肌肉僵硬发作和运动加重。我们已经确定了一个新的PC相关的突变，它取代天冬氨酸的保守的丙氨酸在S4-S5连接域III（A1152 D）。这个残基是特别感兴趣的，因为它在大鼠脑II型Na+通道中的同系物已被认为是快速失活颗粒的重要受体位点。为了确定A1152 D突变引起的生物物理变化，我们在HEK 293（人胚肾）细胞中稳定表达hSkM 1突变体或野生型（WT）通道，并用膜片钳技术记录全细胞Na+电流。在21和IVC下进行实验以更好地理解副肌强直对冷的敏感性。A1152 D突变破坏通道快速失活。与WT相比，突变体通道失活的动力学较慢，并显示了5 mV的去极化位移的电压依赖性的稳态。A1152 D突变通道的另一个明显缺陷是从失活状态的失活速率加快。温度降低10 ℃放大了突变体和WT之间通道门控动力学的差异，并揭示了持续电流和通道失活与开放状态的差异。总的来说，冷加剧突变缺陷可能导致足够过量的Na+流入，产生重复放电和肌强直。根据以前的报告，我们的数据点的功能以及表型之间的差异的突变保守的S4-S5残基的结构域II和III的人骨骼肌Na+通道。
Missense mutations in the human skeletal muscle Na+ channel alpha subunit (hSkM1) are responsible for a number of muscle excitability disorders. Among them, paramyotonia congenita (PC) is characterized by episodes of muscle stiffness induced by cold and aggravated by exercise. We have identified a new PC-associated mutation, which substitutes aspartic acid for a conserved alanine in the S4-S5 linker of domain III (A1152D). This residue is of particular interest since its homologue in the rat brain type II Na+ channel has been suggested as an essential receptor site for the fast inactivation particle. To identify the biophysical changes induced by the A1152D mutation, we stably expressed hSkM1 mutant or wild-type (WT) channels in HEK293 (human embryonic kidney) cells, and recorded whole-cell Na+ currents with the patch-clamp technique. Experiments were performed both at 21 and I VC to better understand the sensitivity to cold of paramyotonia. The A1152D mutation disrupted channel fast inactivation. In comparison to the WT, mutant channels inactivated with slower kinetics and displayed a 5 mV depolarizing shift in the voltage dependence of the steady-state. The other noticeable defect of A1152D mutant channels was an accelerated rate of deactivation from the inactivated state. Decreasing temperature by 10 degrees C amplified the differences in channel gating kinetics between mutant and WT, and unveiled differences in both the sustained current and channel deactivation from the open state. Overall, cold-exacerbated mutant defects may result in a sufficient excess of Na+ influx to produce repetitive firing and myotonia. In the light of previous reports, our data point to functional as well as phenotypic differences between mutations of conserved S4-S5 residues in domains II and III of the human skeletal muscle Na+ channel.