A central core disease mutation in the Ca2+-binding site of skeletal muscle ryanodine receptor impairs single-channel regulation
A central core disease mutation in the Ca2+-binding site of skeletal muscle ryanodine receptor impairs single-channel regulation
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DOI：
10.1152/ajpcell.00052.2019
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发表时间：
2019-08-01
影响因子：
5.5
通讯作者：
Yamaguchi, Naohiro
中科院分区：
文献类型：
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作者：
Chirasani, Venkat R.;Xu, Le;Yamaguchi, Naohiro
冷冻电子显微镜和突变分析表明，1型Ryanodine受体(RyR1)氨基酸残基RyR1-E3893、-E3967和-T5001在钙离子介导的骨骼肌钙释放通道激活中起关键作用。报道了1例2岁男童的钙离子二级结合球区新生错义突变RyR1-Q3970K与中枢性核心病的相关性。在这里，我们通过细胞钙释放测量、单通道记录和计算方法来表征重组RyR1-Q3970K突变体。咖啡因诱导的钙释放研究表明，RyR1-Q3970K在HEK293细胞中形成了咖啡因敏感的钙离子传导通道。然而，在单声道录制中。RyR1-Q3970K的钙依赖通道活性较低，咖啡因或三磷酸腺苷的激活作用明显减弱。RyR1-Q3970E突变对应于错义突变RyR2-Q3925E，与心肌致心律失常综合征相关。与RyR1-Q3970K不同，RyR1-Q3970E在咖啡因诱导的HEK293细胞钙释放中也表现出较低的活性，但与RyR1-Q3970K不同的是，在单通道记录中，RyR1-Q3970K被ATP和咖啡因激活。计算分析表明，两个突变体的Ca~(2+)二级结合球结构重排明显，而Ca~(2+)与RyR1氨基酸残基Glu(3893)、Glu(3967)和Thr(5)(00)(1)的相互作用受影响很小。我们的结论是，RyR1-Q3970在钙依赖的激活RyR1中起着关键作用，RyR1-Q3970K错义突变可能导致骨骼肌病变。
Cryoelectron microscopy and mutational analyses have shown that type 1 ryanodine receptor (RyR1) amino acid residues RyR1-E3893, -E3967, and -T5001 are critical for Ca2+-mediated activation of skeletal muscle Ca' release channel. De novo missense mutation RyR1-Q3970K in the secondary binding sphere of Ca2+ was reported in association with central core disease (CCD) in a 2-yr-old boy. Here, we characterized recombinant RyR1-Q3970K mutant by cellular Ca2+ release measurements, single-channel recordings, and computational methods. Caffeine-induced Ca2+ release studies indicated that RyR1-Q3970K formed caffeine-sensitive, Ca2+-conducting channel in HEK293 cells. However, in single-channel recordings. RyR1-Q3970K displayed low Ca2+-dependent channel activity and greatly reduced activation by caffeine or ATP. A RyR1-Q3970E mutant corresponds to missense mutation RyR2-Q3925E associated with arrhythmogenic syndrome in cardiac muscle. RyR1-Q3970E also formed caffeine-induced Ca2+ release in HEK293 cells and exhibited low activity in the presence of the activating ligand Ca2+ but, in contrast to RyR1-Q3970K, was activated by ATP and caffeine in single-channel recordings. Computational analyses suggested distinct structural rearrangements in the secondary binding sphere of Ca2+ of the two mutants, whereas the interaction of Ca2+ with directly interacting RyR1 amino acid residues Glu(3893), Glu(3967), and Thr(5)(00)(1) was only minimally affected. We conclude that RyR1-Q3970 has a critical role in Ca2+-dependent activation of RyR1 and that a missense RyR1-Q3970K mutant may give rise to myopathy in skeletal muscle.