Mechanism of calsequestrin regulation of single cardiac ryanodine receptor in normal and pathological conditions
Mechanism of calsequestrin regulation of single cardiac ryanodine receptor in normal and pathological conditions
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DOI：
10.1085/jgp.201311022
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发表时间：
2013-08-01
影响因子：
3.8
通讯作者：
Volpe, Pompeo
中科院分区：
文献类型：
--
作者：
Chen, Haiyan;Valle, Giorgia;Volpe, Pompeo
肌浆网（SR）释放的Ca 2+驱动心肌细胞的收缩功能。腔Ca 2+调节SR Ca 2+释放不仅在生理学上而且在生理病理学上是基本的，因为已知异常的腔Ca 2+调节导致心律失常、儿茶酚胺能性多形性室性心动过速（CPVT）和/或心脏骤停，如从动物模型研究推断的。腔Ca 2+通过位于SR内部的机制调节ryanodine受体（RyR）2介导的SR Ca 2+释放;其中之一涉及腔Ca 2+与钙螯合蛋白（CASQ）、三聚体和/或连接蛋白相互作用以调节RyR 2功能。在单个RyR 2通道水平上检查CASQ 2-RyR 2调节。通过融合从野生型（WT）、CASQ 2敲除（KO）或R33 Q-CASQ 2敲入（KI）小鼠分离的天然SR囊泡，将单个RyR 2掺入平面脂质双层中。KO和KI小鼠具有CPVT样表型。我们表明，CASQ 2（WT）对RyR 2功能的作用（激活或抑制）受到胞浆MgATP存在的强烈影响。重构的CASQ 2（WT）-RyR 2复合物的功能不受管腔游离[Ca 2 +]（从0.1至1 mM）变化的影响。CASQ 2（WT）与RyR 2结合产生的抑制作用决定了细胞溶质Ca 2+激活敏感性的降低。来自KO小鼠的RyR 2对胞质Ca 2+活化显著更敏感，并且具有比来自WT小鼠的RyR 2显著更长的平均开放时间。来自KI小鼠的RyR 2s的敏感性介于来自KO和WT小鼠的RyR 2通道之间。增强的胞质RyR 2 Ca 2+敏感性和较长的RyR 2开放时间可能解释了KO和KI小鼠的CPVT样表型。
Release of Ca2+ from the sarcoplasmic reticulum (SR) drives contractile function of cardiac myocytes. Luminal Ca2+ regulation of SR Ca2+ release is fundamental not only in physiology but also in physiopathology because abnormal luminal Ca2+ regulation is known to lead to arrhythmias, catecholaminergic polymorphic ventricular tachycardia (CPVT), and/or sudden cardiac arrest, as inferred from animal model studies. Luminal Ca2+ regulates ryanodine receptor (RyR) 2-mediated SR Ca2+ release through mechanisms localized inside the SR; one of these involves luminal Ca2+ interacting with calsequestrin (CASQ), triadin, and/or junctin to regulate RyR2 function. CASQ2-RyR2 regulation was examined at the single RyR2 channel level. Single RyR2s were incorporated into planar lipid bilayers by the fusion of native SR vesicles isolated from either wild-type (WT), CASQ2 knockout (KO), or R33Q-CASQ2 knock-in (KI) mice. KO and KI mice have CPVT-like phenotypes. We show that CASQ2(WT) action on RyR2 function (either activation or inhibition) was strongly influenced by the presence of cytosolic MgATP. Function of the reconstituted CASQ2(WT)-RyR2 complex was unaffected by changes in luminal free [Ca2+] (from 0.1 to 1 mM). The inhibition exerted by CASQ2(WT) association with the RyR2 determined a reduction in cytosolic Ca2+ activation sensitivity. RyR2s from KO mice were significantly more sensitive to cytosolic Ca2+ activation and had significantly longer mean open times than RyR2s from WT mice. Sensitivity of RyR2s from KI mice was in between that of RyR2 channels from KO and WT mice. Enhanced cytosolic RyR2 Ca2+ sensitivity and longer RyR2 open times likely explain the CPVT-like phenotype of both KO and KI mice.