PDF(Pubmed)
Abstract：
Resistance to inhibitors of cholinesterases (ric-8 proteins) are involved in modulating G-protein function, but little is known of their potential physiological importance in the heart. In the present study, we assessed the role of resistance to inhibitors of cholinesterase 8b (Ric-8b) in determining cardiac contractile function. We developed a murine model in which it was possible to conditionally delete ric-8b in cardiac tissue in the adult animal after the addition of tamoxifen. Deletion of ric-8b led to severely reduced contractility as measured using echocardiography days after administration of tamoxifen. Histological analysis of the ventricular tissue showed highly variable myocyte size, prominent fibrosis, and an increase in cellular apoptosis. RNA sequencing revealed transcriptional remodeling in response to cardiac ric-8b deletion involving the extracellular matrix and inflammation. Phosphoproteomic analysis revealed substantial downregulation of phosphopeptides related to myosin light chain 2. At the cellular level, the deletion of ric-8b led to loss of activation of the L-type calcium channel through the β-adrenergic pathways. Using fluorescence resonance energy transfer-based assays, we showed ric-8b protein selectively interacts with the stimulatory G-protein, Gαs. We explored if deletion of Gnas (the gene encoding Gαs) in cardiac tissue using a similar approach in the mouse led to an equivalent phenotype. The conditional deletion of the Gαs gene in the ventricle led to comparable effects on contractile function and cardiac histology. We conclude that ric-8b is essential to preserve cardiac contractile function likely through an interaction with the stimulatory G-protein and downstream phosphorylation of myosin light chain 2.
摘要：
对胆碱酯酶（ric-8蛋白）抑制剂的抗性与调节G蛋白功能有关，但对其在心脏中潜在的生理重要性知之甚少。在本研究中,我们评估了对胆碱酯酶8b(Ric-8b)抑制剂的耐药性在确定心脏收缩功能方面的作用.我们开发了一种鼠模型，其中在添加他莫昔芬后，可以有条件地删除成年动物心脏组织中的ric-8b。在施用他莫昔芬后几天使用超声心动图测量，ric-8b的缺失导致收缩性严重降低。心室组织的组织学分析显示高度可变的心肌细胞大小，显著的纤维化和细胞凋亡的增加。RNA测序显示响应于涉及细胞外基质和炎症的心脏rc-8b缺失的转录重塑。磷酸化蛋白质组分析显示与肌球蛋白轻链2相关的磷酸肽的显著下调。在细胞层面,rc-8b的缺失导致通过β-肾上腺素能途径的L型钙通道的激活丧失。使用基于荧光共振能量转移的测定，我们显示了ric-8b蛋白与刺激性G蛋白选择性相互作用，Gαs.我们探索了在小鼠中使用类似方法在心脏组织中缺失Gnas（编码Gαs的基因）是否导致等效表型。心室中Gαs基因的条件性缺失导致对收缩功能和心脏组织学的可比影响。我们得出的结论是，ric-8b对于保持心脏收缩功能至关重要，可能是通过与刺激G蛋白相互作用和肌球蛋白轻链2的下游磷酸化。
