泛素-蛋白酶体系统(UPS)是通过与泛素结合而导致底物蛋白选择性降解的重要机制。由于心肌细胞的自我更新能力非常有限,因为它们容易由于持续的机械和代谢压力而导致蛋白质损伤,UPS在心脏生理学和病理生理学中起着关键作用。虽然蛋白酶体活性的改变有助于各种心脏病变,如心力衰竭和缺血/再灌注损伤(IRI),影响其活动的环境线索仍然未知,他们是这项工作的重点。Ciechanover小组最近的一项研究表明,培养的癌细胞系中的氨基酸(AA)饥饿调节蛋白酶体细胞内定位和活性,我们在人类诱导多能干细胞衍生的心肌细胞中测试了两个假设(iPSC-CM,CMs):(i)AA饥饿导致CMs中的蛋白酶体易位,与在培养的癌细胞系中观察到的相似;(ii)亚细胞蛋白酶体区室化的操作与心律失常形式的电生理异常有关,通过改变细胞内Ca2+处理介导。主要发现是:(i)使CMs饥饿到AA导致蛋白酶体从细胞核转位到细胞质,同时补充芳香氨基酸酪氨酸(Y),色氨酸(W)和苯丙氨酸(F)(YWF)抑制蛋白酶体募集;(ii)缺乏AA的治疗引起心律失常;(iii)在核蛋白酶体隔离(-AAYWF)后观察到的心律失常被KB-R7943阻断,后者是钠钙交换剂NCX逆转模式的抑制剂;(iv)与AA饥饿有关的离体大鼠心脏的灌注等级与AA饥饿有关。总的来说,我们的新发现描述了一种新发现的机制,该机制将UPS与CM和整个心脏的心律失常发生联系起来.
The ubiquitin-proteasome system (UPS) is an essential mechanism responsible for the selective degradation of substrate proteins via their conjugation with ubiquitin. Since cardiomyocytes have very limited self-renewal capacity, as they are prone to protein damage due to constant mechanical and metabolic stress, the UPS has a key role in cardiac physiology and pathophysiology. While altered proteasomal activity contributes to a variety of cardiac pathologies, such as heart failure and ischemia/reperfusion injury (IRI), the environmental cues affecting its activity are still unknown, and they are the focus of this work. Following a recent study by Ciechanover\'s group showing that amino acid (AA) starvation in cultured cancer cell lines modulates proteasome intracellular localization and activity, we tested two hypotheses in human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs, CMs): (i) AA starvation causes proteasome translocation in CMs, similarly to the observation in cultured cancer cell lines; (ii) manipulation of subcellular proteasomal compartmentalization is associated with electrophysiological abnormalities in the form of arrhythmias, mediated via altered intracellular Ca2+ handling. The major findings are: (i) starving CMs to AAs results in proteasome translocation from the nucleus to the cytoplasm, while supplementation with the aromatic amino acids tyrosine (Y), tryptophan (W) and phenylalanine (F) (YWF) inhibits the proteasome recruitment; (ii) AA-deficient treatments cause arrhythmias; (iii) the arrhythmias observed upon nuclear proteasome sequestration(-AA+YWF) are blocked by KB-R7943, an inhibitor of the reverse mode of the sodium-calcium exchanger NCX; (iv) the retrograde perfusion of isolated rat hearts with AA starvation media is associated with arrhythmias. Collectively, our novel findings describe a newly identified mechanism linking the UPS to arrhythmia generation in CMs and whole hearts.