Elp3 Is Involved In The Mechanism Of Oxidative Stress Mediated By Yap1 And Skn7 In Saccharomyces Cerevisiae

Elongator is an important transcription extension factor in yeast.It is a histone acetyltransferase complex composed of two copies of Elp1-Elp6.It participates in many biological processes by regulating transcription,exocytosis and t RNA modification.Elp3,is the catalytic subunit of Elongator.The results of previous studies in our laboratory show that the absence of Elp3 can reduce the tolerance of yeast to H₂O₂,increase the intracellular MDA and ROS,and significantly reduce the nuclear accumulation of Yap1,the central transcription factor of yeast oxidative stress,indicating that Elp3 participates in oxidative stress of yeast through Yap1 pathway.In order to further study the relationship between Elp3 and antioxidant transcription factors in oxidative stress response of yeast and explore the mechanism of its influence on the nuclear localization of Yap1,the double or triple deletion mutants of Elp3 and transcription factors Yap1,Skn7 and the overexpression plasmids of Yap1,Skn7 and Yap1 nuclear localization related proteins Trx2,Gpx3 and Ybp1 were constructed in this experiment.The tolerance,physiological and biochemical indexes of deletion and overexpression strains under H₂O₂ stress were detected to determine the effect of Elp3 on oxidative stress.The production of ROS can induce oxidative sensitivity and oxidative stress in yeast.Therefore,the sensitivity and survival rate of elp3?yap1?and elp3?skn7?and elp3?yap1?skn7?cells treated with H₂O₂ were measured.It was found that the co-deletion of Elp3 and Yap1 or Skn7 increased the oxidative sensitivity of yap1?and skn7?and reduced the survival rate.MDA and ROS can reflect the degree of cell oxidative damage.Consistenting with the results of sensitivity experiment,the level of MDA and ROS in Yap1 and Skn7 deletion mutants was significantly increased by co-deletion of Elp3,which indicates that Elp3 directly or indirectly affects the oxidative stress mediated by Yap1 and Skn7.Moreover,overexpression of Yap1,Skn7,Trx2,Gpx3 and Ybp1 in WT can significantly reduce MDA and ROS levels,but overexpression of five transcription related proteins in elp3?strain can not change MDA and ROS levels,further indicating that Elp3 is indeed related to antioxidant transcription response mediated by Yap1and Skn7.Yap1,as an antioxidant transcription factor,exists in the cytoplasm under normal conditions.When induced by H₂O₂,Yap1 enters the nucleus and activates the transcription of antioxidant target genes.Yap1 requires oxidative folding before entering the nucleus,and its protein modification process involves the assistance of Trx2-Gpx3 and Ybp1 proteins.The results showed that the overexpression of Trx2,Gpx3and Ybp1 in WT strains could improve the nuclear localization ratio of Yap1,but had no significant change in elp3?strains.It is suggested that Elp3 are involved in Yap1 mediated oxidative stress by influencing the process of Yap1 oxidative folding assisted by Trx2,GPx3 and Ybp1.The results of transcriptional activity showed that the transcriptional activity of Yap1 and Skn7 in elp3?was significantly higher than that of WT.This result seems to be contrary to the decrease of nuclear accumulation of Yap1 in elp3?.Further experiments showed that,unlike in WT,overexpression of Trx2,Gpx3 and Ybp1 in elp3?strain did not enhance the transcription activity of Yap1.It is speculated that the loss of Elp3 may hinder the process of the Yap1 entering the nucleus assisted by Trx2,Gpx3 and Ybp1,results in a significant reduction of Yap1 nuclear accumulation,thus switch on other compensatory pathways to activate the transcription activity of less Yap1 in the nucleus and reducs the damage to cells caused by the rapid accumulation of ROS,so as to remedy the oxidative sensitive phenotype caused by the loss of Elp3.