Screening And Analysis Of Differentially Expressed Phosphoproteins Of The PEDV-infected Vero E6 Cells

Porcine epidemic diarrhea virus(PEDV)is an acute and highly contagious enteric disease characterized by severe diarrhea,vomiting,dehydration,and a mortality rate of up to 90% in suckling piglets.Recently,the outbreak of PEDV has shown an increasing trend in worldwide.Phosphorylation is the most common form of transient protein modification.The virus uses the host kinase system to phosphorylate viral proteins to gain efficient infection and control host innate immunity.However,few reports on the changes of phosphorylated proteins in host cells mediated by PEDV infection.In this study,the differential expressed phosphoproteins of PEDV-infected Vero E6 cells was investigated using an isobaric tag for relative and absolute quantitation(iTRAQ)-MS/MS approach.The results showed that the numbers of identified peptides and phosphoproteins between PEDV infected group(V)and PEDV infected mock(C)were 2 385 and 1134,respectively,of which 455 phosphoproteins have quantitative information and 679 phosphoproteins have not quantitative information.A total of 109 differentially expressed phosphoproteins(DEPs)were found between the V group and C group(p<0.05,ratio>1.5),of which up phosphoproteins were 72,the rest belong to down phosphoproteins,and molecular function of the DEPs mainly focused on the binding according to Gene Ontology categories.The pathways in which the DEPs participated 113 signal pathways,of which some anti-viral pathways and phosphoproteins,such as Toll-like receptor signaling pathway,MAPK signaling pathway,PI3K-Akt signaling pathway,RIG-I-like receptor signaling pathway,mTOR signaling pathway,and heat shock protein 27,heat shock protein 70,were highlighted as potential factors in PEDV infection.Western blot results showed that the ratios of the up-regulated protein heat shock protein 27 between the V and C groups were consistent with those obtained using the quantitative phosphoproteomics analysis.In addition,PEDV infects Vero E6 Cells activated Heat Shock Protein 70 and differential expression of heat shock protein 70 directly affects the expression level of PEDV m RNA.The resulting data will provide valuable information to increaseunderstanding of PEDV infection.