Defective domain-domain interactions within the ryanodine receptor as a critical cause of diastolic Ca2+ leak in failing hearts
Defective domain-domain interactions within the ryanodine receptor as a critical cause of diastolic Ca2+ leak in failing hearts
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DOI：
10.1093/cvr/cvn303
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发表时间：
2009-02-15
影响因子：
10.8
通讯作者：
Matsuzaki, Masunori
中科院分区：
文献类型：
--
作者：
Tateishi, Hiroki;Yano, Masafumi;Matsuzaki, Masunori
已知与心脏2型兰尼碱受体（RyR 2）的Gly（2460）-Pro（2495）区域匹配的结构域肽（DP）DPc 10模拟与儿茶酚胺能多形室性心动过速（CPVT）相关的通道功能障碍，这是由于其干扰N-末端（1-600）和中央（2000-2500）结构域的正常相互作用（即结构域解压缩）。本研究利用DPc 10和另外两个突变位点不同的DPs，从犬左心室肌分离肌浆网（SR）囊泡和心肌细胞，进行Ca ~（2+）泄漏和火花分析，以探讨衰竭心脏中Ca ~（2+）循环异常的潜在机制。使用对应于RyR 2的163-195和4090 - 4123区域的DP（分别为DP 163 - 195和DP 4090 - 4123）作为定点载体，用甲基香豆素乙酸酯（MCA）荧光标记SR的RyR 2部分。两种DP介导RyR 2的特异性MCA荧光标记。添加任何DP的MCA标记的SR诱导域解压缩，证明了一个大尺寸的荧光猝灭剂的结合MCA的可及性增加。衰竭心肌细胞SR Ca ~（2+）漏和Ca ~（2+）火花频率（SpF）均显著增加。在将DP 163 -195或DP 4090 -4123引入正常SR或心肌细胞后，Ca 2+渗漏和SpF均增加至与衰竭心肌细胞相当的水平。K201（JTV 519）抑制DP 163 -195诱导的所有效应（结构域解链和增加Ca 2+渗漏和SpF）或衰竭心肌细胞中的效应，但不抑制DP 4090 - 4123诱导的效应。N端和中央结构域之间的缺陷性结构域相互作用诱导舒张期Ca 2+渗漏，导致心力衰竭和致死性心律失常。在CPVT中观察到的C-末端区域的突变似乎不与上述N-末端和中央结构域间相互作用通信，尽管自发的Ca 2+泄漏被类似地诱导。
A domain peptide (DP) matching the Gly(2460)-Pro(2495) region of the cardiac type-2 ryanodine receptor (RyR2), DPc10, is known to mimic channel dysfunction associated with catecholaminergic polymorphic ventricular tachycardia (CPVT), owing to its interference in a normal interaction of the N-terminal (1-600) and central (2000-2500) domains (viz. domain unzipping). Using DPc10 and two other DPs harboring different mutation sites, we investigated the underlying mechanism of abnormal Ca2+ cycling in failing hearts.Sarcoplasmic reticulum (SR) vesicles and cardiomyocytes were isolated from dog left ventricular muscles for Ca2+ leak and spark assays. The RyR2 moiety of the SR was fluorescently labelled with methylcoumarin acetate (MCA) using DPs corresponding to the 163-195 and 4090-4123 regions of RyR2 (DP163-195 and DP4090-4123, respectively) as site-directed carriers. Both DPs mediated a specific MCA fluorescence labelling of RyR2. Addition of either DP to the MCA-labelled SR induced domain unzipping, as evidenced by an increased accessibility of the bound MCA to a large-size fluorescence quencher. Both SR Ca2+ leak and Ca2+ spark frequency (SpF) were markedly increased in failing cardiomyocytes. Upon introduction of DP163-195 or DP4090-4123 into normal SR or cardiomyocytes, both Ca2+ leak and SpF increased to the levels comparable with those of failing myocytes. K201 (JTV519) suppressed all of the effects induced by DP163-195 (domain unzipping and increased Ca2+ leak and SpF) or those in failing cardiomyocytes, but did not suppress the effects induced by DP4090-4123.Defective inter-domain interaction between N-terminal and central domains induces diastolic Ca2+ leak, leading to heart failure and lethal arrhythmia. Mutation at the C-terminal region seen in CPVT does not seem to communicate with the aforementioned N-terminal and central inter-domain interaction, although spontaneous Ca2+ leak is similarly induced.