Nikolaienko Roman, Bovo Elisa, Zima Aleksey V
Department of Cell and Molecular Physiology, Strich School of Medicine, Loyola University Chicago, Maywood, Illinois.
Department of Cell and Molecular Physiology, Strich School of Medicine, Loyola University Chicago, Maywood, Illinois.
Biophys Rep (N Y). 2024 Dec 11;4(4):100183. doi: 10.1016/j.bpr.2024.100183. Epub 2024 Sep 27.
The type 2 ryanodine receptor (RyR2) is the major Ca release channel required for Ca-induced Ca release (CICR) and cardiac excitation-contraction coupling. The cluster organization of RyR2 at the dyad is critical for efficient CICR. Despite its central role in cardiac Ca signaling, the mechanisms that control CICR are not fully understood. As a single RyR2 Ca flux dictates local CICR that underlies Ca sparks, RyR2 density in a cluster, and therefore the distance between RyR2s, should have a profound impact on local CICR. Here, we studied the effect of the RyR2 expression level ([RyR2]) on CICR activation, termination, and amplitude. The endoplasmic reticulum (ER)-targeted Ca sensor RCEPIA-1er was used to directly measure the ER [Ca] (Ca]) in the T-Rex-293 the sarco/endoplasmic reticulum Ca-ATPase (SERCA2a) stable cell line expressing human RyR2. Cells coexpressing RyR2 and SERCA2a produced periodic [Ca] depletions in the form of spontaneous Ca waves due to propagating CICR. For each studied cell, the [Ca] at which Ca waves are activated and terminated was analyzed as a function of [RyR2]. CICR parameters, such as [Ca] activation, termination, and amplitude, were inversely proportional to [RyR2] at low-intermediate levels. Increasing the sensitivity of RyR2 to cytosolic Ca lowered the [Ca] at which CICR is activated and terminated. Decreasing the sensitivity of RyR2 to cytosolic Ca had the opposite effect on CICR. These results suggest that RyR2 density in the release cluster should have a significant impact on local CICR activation and termination. Since SR Ca load is evenly distributed throughout the SR network, clusters with higher RyR2 density would have a higher probability of initiating spontaneous CICR.
2型兰尼碱受体（RyR2）是钙诱导钙释放（CICR）和心脏兴奋-收缩偶联所需的主要钙释放通道。RyR2在二联体处的簇状组织对于有效的CICR至关重要。尽管其在心脏钙信号传导中起核心作用，但控制CICR的机制尚未完全了解。由于单个RyR2钙通量决定了作为钙火花基础的局部CICR，因此簇中RyR2的密度以及因此RyR2之间的距离应该对局部CICR产生深远影响。在这里，我们研究了RyR2表达水平（[RyR2]）对CICR激活、终止和幅度的影响。内质网（ER）靶向钙传感器RCEPIA-1er用于直接测量表达人RyR2的T-Rex-293肌浆网/内质网钙ATP酶（SERCA2a）稳定细胞系中的内质网[Ca]（Ca]）。共表达RyR2和SERCA2a细胞由于传播的CICR而以自发钙波的形式产生周期性[Ca]消耗。对于每个研究的细胞，分析作为[RyR2]函数的钙波激活和终止时的[Ca]。在低-中等水平时，CICR参数，如[Ca]激活、终止和幅度，与[RyR2]成反比。提高RyR2对胞质钙的敏感性会降低CICR激活和终止时的[Ca]。降低RyR2对胞质钙的敏感性对CICR有相反的影响。这些结果表明，释放簇中RyR2的密度应该对局部CICR激活和终止有显著影响。由于肌浆网钙负荷均匀分布在整个肌浆网网络中，具有较高RyR2密度的簇将有更高的概率引发自发CICR。
