The role of calsequestrin, triadin, and junctin in conferring cardiac ryanodine receptor responsiveness to luminal calcium
The role of calsequestrin, triadin, and junctin in conferring cardiac ryanodine receptor responsiveness to luminal calcium
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DOI：
10.1016/s0006-3495(04)74271-x
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发表时间：
2004-04-01
影响因子：
3.4
通讯作者：
Györke, S
中科院分区：
文献类型：
--
作者：
Györke, I;Hester, N;Györke, S
肌浆网（SR）内的Ca水平是心肌中Ca释放通道/ryanodine受体（RyR）功能活性的重要决定因素。然而，RyR调节管腔钙的分子基础仍然在很大程度上未知。在本研究中，我们研究了心脏SIR管腔辅助蛋白calsequestrin（CSQ），triadin 1和junctin在形成心脏RyR的管腔钙传感器中的潜在作用。记录的单一RyR通道纳入脂质双层，从SR囊泡或纯化的RyR制剂，在MgATP的存在下进行使用Cs+作为电荷载体。将管腔[Ca]从20 μ M提高到5 mM增加了SR囊泡中天然RyR的开放通道概率（P-O），但不增加纯化RyR的开放通道概率。将CSQ添加到纯化通道的管腔侧不会产生P-o的显著变化，也不会恢复RyR响应管腔Ca的能力。当将三联蛋白1和连接蛋白加入到纯化通道的管腔侧时，RyR P-0显著增加;然而，通道仍然对管腔[Ca]的变化无反应。在与三醇溶蛋白1和连接蛋白重新关联的RyR中，添加鲁米那CSQ产生活性的显著降低。在与所有三种蛋白质重新结合后，RyRs通过增加它们的P-o来响应管腔[Ca]的升高。这些结果表明，CSQ，三联蛋白1和连接蛋白的复合物赋予RyR管腔Ca敏感性。CSQ显然充当在低管腔[Ca]下抑制通道的管腔Ca传感器，而可能需要三聚体蛋白1和/或连接蛋白来介导CSQ与RyR的相互作用。
The level of Ca inside the sarcoplasmic reticulum (SR) is an important determinant of functional activity of the Ca release channel/ryanodine receptor (RyR) in cardiac muscle. However, the molecular basis of RyR regulation by luminal Ca remains largely unknown. In the present study, we investigated the potential role of the cardiac SIR luminal auxiliary proteins calsequestrin (CSQ), triadin 1, and junctin in forming the luminal calcium sensor for the cardiac RyR. Recordings of single RyR channels incorporated into lipid bilayers, from either SR vesicle or purified RyR preparations, were performed in the presence of MgATP using Cs+ as the charge carrier. Raising luminal [Ca] from 20 muM to 5 mM increased the open channel probability (P-o) of native RyRs in SR vesicles, but not of purified RyRs. Adding CSQ to the luminal side of the purified channels produced no significant changes in P-o, nor did it restore the ability of RyRs to respond to luminal Ca. When triadin 1 and junctin were added to the luminal side of purified channels, RyR P-o increased significantly; however, the channels still remained unresponsive to changes in luminal [Ca]. In RyRs reassociated with triadin 1 and junctin, adding luminal CSQ produced a significant decrease in activity. After reassociation with all three proteins, RyRs responded to rises of luminal [Ca] by increasing their P-o. These results suggest that a complex of CSQ, triadin 1, and junctin confer RyR luminal Ca sensitivity. CSQ apparently serves as a luminal Ca sensor that inhibits the channel at low luminal [Ca], whereas triadin 1 and/or junctin may be required to mediate interactions of CSQ with RyR.