Ero1α 依赖性 ERp44 从 RyR2 上解离导致心律失常。
Ero1α-Dependent ERp44 Dissociation From RyR2 Contributes to Cardiac Arrhythmia.
机构信息
Department of Physiology and Cell Biology (S.H., R.T., V.B., F.P., J.Y., X.A., A.E.B., J.P.D., S.G., D.T.), The Ohio State University.
Dorothy M. Davis Heart and Lung Research Institute, College of Medicine, The Ohio State University Wexner Medical Center, Columbus (S.H., R.T., V.B., F.P., J.Y., X.A., C.A.C., A.E.B., J.P.D., S.G., D.T.).
出版信息
Circ Res. 2022 Mar 4;130(5):711-724. doi: 10.1161/CIRCRESAHA.121.320531. Epub 2022 Jan 28.
BACKGROUND
Oxidative stress in cardiac disease promotes proarrhythmic disturbances in Ca homeostasis, impairing luminal Ca regulation of the sarcoplasmic reticulum (SR) Ca release channel, the RyR2 (ryanodine receptor), and increasing channel activity. However, exact mechanisms underlying redox-mediated increase of RyR2 function in cardiac disease remain elusive. We tested whether the oxidoreductase family of proteins that dynamically regulate the oxidative environment within the SR are involved in this process.
METHODS
A rat model of hypertrophy induced by thoracic aortic banding (TAB) was used for ex vivo whole heart optical mapping and for Ca and reactive oxygen species imaging in isolated ventricular myocytes (VMs).
RESULTS
The SR-targeted reactive oxygen species biosensor ERroGFP showed increased intra-SR oxidation in TAB VMs that was associated with increased expression of Ero1α (endoplasmic reticulum oxidoreductase 1 alpha). Pharmacological (EN460) or genetic Ero1α inhibition normalized SR redox state, increased Ca transient amplitude and SR Ca content, and reduced proarrhythmic spontaneous Ca waves in TAB VMs under β-adrenergic stimulation (isoproterenol). Ero1α overexpression in Sham VMs had opposite effects. Ero1α inhibition attenuated Ca-dependent ventricular tachyarrhythmias in TAB hearts challenged with isoproterenol. Experiments in TAB VMs and human embryonic kidney 293 cells expressing human RyR2 revealed that an Ero1α-mediated increase in SR Ca-channel activity involves dissociation of intraluminal protein ERp44 (endoplasmic reticulum protein 44) from the RyR2 complex. Site-directed mutagenesis and molecular dynamics simulations demonstrated a novel redox-sensitive association of ERp44 with RyR2 mediated by intraluminal cysteine 4806. ERp44-RyR2 association in TAB VMs was restored by Ero1α inhibition, but not by reducing agent dithiothreitol, as hypo-oxidation precludes formation of covalent bond between RyR2 and ERp44.
CONCLUSIONS
A novel axis of intraluminal interaction between RyR2, ERp44, and Ero1α has been identified. Ero1α inhibition exhibits promising therapeutic potential by stabilizing RyR2-ERp44 complex, thereby reducing spontaneous Ca release and Ca-dependent tachyarrhythmias in hypertrophic hearts, without causing hypo-oxidative stress in the SR.
背景
心脏疾病中的氧化应激会导致钙稳态的致心律失常紊乱，损害肌浆网（SR）钙释放通道 RyR2（ryanodine 受体）的管腔钙调节，并增加通道活性。然而，氧化还原介导的心脏疾病中 RyR2 功能增加的确切机制仍不清楚。我们测试了是否氧化还原酶家族蛋白，这些蛋白动态调节 SR 内的氧化环境，是否参与了这一过程。
方法
使用胸主动脉缩窄（TAB）诱导的大鼠肥厚模型进行离体心脏光学映射和分离心室肌细胞（VMs）中的 Ca 和活性氧成像。
结果
SR 靶向活性氧生物传感器 ERroGFP 显示 TAB VMs 中 SR 内氧化增加，与 Ero1α（内质网氧化还原酶 1α）表达增加相关。药理学（EN460）或遗传 Ero1α 抑制使 SR 氧化还原状态正常化，增加 Ca 瞬变幅度和 SR Ca 含量，并减少 TAB VMs 在β-肾上腺素刺激（异丙肾上腺素）下的致心律失常自发性 Ca 波。Sham VMs 中的 Ero1α 过表达则产生相反的效果。Ero1α 抑制可减轻 TAB 心脏在异丙肾上腺素挑战下发生的 Ca 依赖性室性心动过速。在 TAB VMs 和表达人 RyR2 的人胚肾 293 细胞中的实验表明，Ero1α 介导的 SR Ca 通道活性增加涉及内腔蛋白 ERp44（内质网蛋白 44）与 RyR2 复合物的解离。定点突变和分子动力学模拟表明，ERp44 与 RyR2 的一种新型氧化还原敏感关联由内腔半胱氨酸 4806 介导。TAB VMs 中的 ERp44-RyR2 关联通过 Ero1α 抑制得到恢复，但不能通过还原剂二硫苏糖醇（dithiothreitol）恢复，因为低氧化会阻止 RyR2 和 ERp44 之间形成共价键。
结论
已经确定了 RyR2、ERp44 和 Ero1α 之间内腔相互作用的新轴。Ero1α 抑制通过稳定 RyR2-ERp44 复合物，从而减少肥厚心脏中的自发性 Ca 释放和 Ca 依赖性心动过速，同时不引起 SR 中的低氧化应激，具有有前景的治疗潜力。
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