Mutation-linked defective interdomain interactions within ryanodine receptor cause aberrant Ca²⁺release leading to catecholaminergic polymorphic ventricular tachycardia.
Mutation-linked defective interdomain interactions within ryanodine receptor cause aberrant Ca²⁺release leading to catecholaminergic polymorphic ventricular tachycardia.
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DOI：
10.1161/circulationaha.111.023259
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发表时间：
2011-08-09
期刊：
影响因子：
37.8
通讯作者：
Matsuzaki M
中科院分区：
文献类型：
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作者：
Suetomi T;Yano M;Uchinoumi H;Fukuda M;Hino A;Ono M;Xu X;Tateishi H;Okuda S;Doi M;Kobayashi S;Ikeda Y;Yamamoto T;Ikemoto N;Matsuzaki M
心脏Ryanodine受体（RyR 2）内的单个氨基酸突变诱导的儿茶酚胺能多态性室性心动过速（CPVT）的分子机制仍然难以捉摸。在这里，我们研究了突变诱导的构象缺陷的RyR 2使用敲入（KI）小鼠模型表达的人CPVT相关的RyR 2突变体（S2246 L;丝氨酸亮氨酸突变的残基2246）。我们检查的所有KI小鼠在跑步机上运动后产生VT。cAMP依赖性增加的频率的Ca 2+火花更明显的皂苷渗透KI心肌细胞比WT心肌细胞。DP 2246（对应于2232-2266区域的肽：2246结构域）特异性结合的定点荧光标记和石英微量天平测定显示，DP 2246与RyR 2的K201结合序列（1741- 2270）结合。将S2246 L突变引入DP 2246中增加了肽结合的亲和力。RyR 2内结构域间相互作用的荧光猝灭测定显示，2246结构域/K201结合结构域的紧密相互作用以变构方式与N-末端（1-600）/中心（2000-2500）结构域对的结构域解压缩偶联。丹曲林纠正了突变引起的域开关的域解压缩，并停止了运动诱发的室性心动过速。RyR 2的CPVT连锁突变S2246 L在涉及突变位点的中心结构域内引起异常紧密的局部亚结构域/亚结构域相互作用，这诱导N-末端和中心结构域之间的缺陷性相互作用。这导致在舒张状态下Ca 2+通道的错误激活，反映在增加的Ca 2+火花频率上，然后导致致命的心律失常。
The molecular mechanism by which catecholaminergic polymorphic ventricular tachycardia (CPVT) is induced by single amino acid mutations within the cardiac ryanodine receptor (RyR2) remains elusive. Here, we investigated mutation-induced conformational defects of RyR2 using a knock-in (KI) mouse model expressing the human CPVT-associated RyR2 mutant (S2246L; Serine to Leucine mutation at the residue 2246). All KI mice we examined produced VT after exercise on a treadmill. cAMP-dependent increase in the frequency of Ca2+ sparks was more pronounced in saponin-permeabilized KI cardiomyocytes than in WT cardiomyocytes. Site-directed fluorescent labeling and quartz microbalance assays of the specific binding of DP2246 (a peptide corresponding to the 2232–2266 region: the 2246 domain) showed that DP2246 binds with the K201-binding sequence of RyR2 (1741– 2270). Introduction of S2246L mutation into the DP2246 increased the affinity of peptide binding. Fluorescence quench assays of inter-domain interactions within RyR2 showed that tight interaction of the 2246 domain/K201-binding domain is coupled with domain unzipping of the N-terminal (1-600)/central (2000–2500) domain pair in an allosteric manner. Dantrolene corrected the mutation-caused domain unzipping of the domain switch, and stopped the exercise-induced ventricular tachycardia. The CPVT-linked mutation of RyR2, S2246L, causes an abnormally tight local sub-domain/sub-domain interaction within the central domain involving the mutation site, which induces defective interaction between the N-terminal and central domains. This results in an erroneous activation of Ca2+ channel in a diastolic state reflecting on the increased Ca2+ spark frequency, which then leads to lethal arrhythmia.