Study On The Glutathionylation Modification Of ATP Synthase In Angiotensin Ⅱ-induced Mouse Cardiac Hypertrophy Model

Cardiac hypertrophy is a common pathological process of chronic hypertension,hypertrophic cardiomyopathy and other heart diseases.The mechanism of cardiac hypertrophy is very complex,which involves changes at the level of gene transcription,stimulation of the rate of protein synthesis(translation),and increased assembly of myofibrils.In the last two decades,there is increasing evidence that oxidative stress is involved in the formation of the cardiac hypertrophy and chronic heart failure.Reactive oxygen species mediated redox signaling and modification play an important role in regulating cardiac hypertrophy.However,the effect of redox modification in cardiac hypertrophy is not clear.S-glutathionylation is an oxidative post-translational modification of protein cysteine residues by the addition of glutathione,which is usually involved in the condition of oxidative stress or nitrosative stress.As a tripeptide of glutamic acid,cysteine,and glycine,GSH represents one of the most prevalent and important thiol buffers in the cell.The ratio of GSH(reduced)and its disulfide,GSSG(oxidized),contributes to the redox potential of the cell and thereby contributes to redox homeostasis.Intracellular GSH level is affected by cysteamine,one of the product of Vanin 1.S-glutathionylation can protect cysteine thiol from irreversible oxidative damage but this post-translational modification adds a tripeptide and a net negative charge that can lead to distinct structural and functional changes in the target protein.F1FO-ATPase is a key enzyme found in mitochondrial inner membranes where ATP,the perfect energy currency for the cells,was generated.It uses a proton gradient to drive ATP synthesis by allowing the passive flux of protons across the membrane down their electrochemical gradient and using the energy released by the transport reaction to release newly formed ATP from the active site of F-ATPase.In recent years,it has been reported that cysteine of ATP synthase a and y subunits underwent a process of S-glutathionylation in dysynchronic heart failure(DHF)models,which resulted in alteration in the hydrolysis activity of ATP synthase.In contrast,the S-glutathionylation of ATP synthase was reduced and ATP synthase activity was recovered in the canine model of cardiac resynchronization therapy(CRT).In addition,in the pathologic model of ischemia/reperfusion and heart failure patients,ATP synthase also underwent S-glutathionylation,implying that S-glutathionylation as an important redox modification is closely related to the progress of pathological processes in as cardiovascular diseases.However,it is not clear whether S-glutathionylation on ATP synthase participated in the pathological condition of cardiac hypertrophy.In the present study,based on the cardiac hypertrophic mouse model induced by Ang II infusion,the differentially expressed proteins were identified using quantitative proteomics approach,and the alteration of S-glutathionylation on ATP synthase was explored.The results showed that the most of the differentially expressed proteins in the Ang II-stimulated heart were located in the mitochondria,which were mainly involved in cardiac energy metabolism and ATP synthesis.Ang II stimulation caused significant remodeling of cardiac energy metabolism characterized as that cardiac glucose and lipid metabolism associated gene expressions were markedly elevated and lipid accumulation was evidenced.Moreover,Ang II stimulation induced significant increases in ROS level and the up-regulation of uncoupled NOSs and NADPH oxidases,indicating the oxidative stress in Ang II induced-mouse model of cardiac hypertrophy.Accordingly,the GSH/GSSG ratio was significantly decreased,and the S-glutathionylation of both in cardiac and mitochondria proteins were documented in Ang-II stimulated heart samples.Furthermore,S-glutathionylation of ATP synthase alpha subunit was enhanced in hypertrophic model in vitro induced by Ang II.Consequently,the activity of ATP synthase was inhibited by S-glutathionylation.In summary,in the model of Ang II induced-mouse cardiac hypertrophy,differentially expressed proteins were mainly associated with energy metabolism and oxidative stress.Oxidative stress led to the change of S-glutathionylation,which resulted in reduction of the activity of ATP synthase.From the perspective of S-glutathionylation,this study provided a new insight into the study of mechanism of energy metabolism remodeling.