Skin Proteomics Analysis Of Zebrafish In Responses To Spring Viremia Of Carp Virus (SVCV)

In this study,the zebrafish(Danio rerio)was infected with the spring viremia of carp virus(SVCV)as the experimental animal model.The clinical manifestations of the diseased fish were spotted hemorrhages on the skin,gills and fins.Histopathological observation was performed,the results showed that separation of superficial and deep layer in skin,melanocytes decreased,cystic and goblet mucous cells increased;gill lamellae basal cell proliferation,epithelial necrosis;mucosal epithelial vacuoles cells,cystic cells increased in testine;hepatic cell edema,necrosis;disappearance cell nuclei in the spleen,expansion of splenic sinus,and necrosis.Furthermore,zebrafish as model by using SVCV virus injected infection.Proteomics analysis of SVCV virus and differential protein expression of the infected zebrafish skin were performed using iTRAQ techniques.The results showed that the number of maps matched to the identified peptides was 65385,and 1996 peptides were identified.The number of unique peptides identified was 15233,and a total of 3,999 proteins were identified.A total of 320 differential proteins were identified in post-infection 24 h,among which 131 proteins were up-regulated and 189 proteins were down-regulated;181 differential proteins were identified in post-infection 96 h,of which 104 were up-regulated and 77 were down-regulated;GO and KEGG pathways statistic analysis showed the differentially expressed proteins involved in that chemokines,TLR,type I IFN,NF-?B,MAPK and complement systemsignal pathways in skin mucosa immune response process.The results showed that SVCV virus infects zebrafish for 24 h and complements(C3,C6,C9),Toll-like receptor 3(TLR3),interferon-stimulating gene 15(ISG15),nuclear factor NF-?B(NF-?B),and melanin in the skin.Tumor differentiation related protein 5(MDA5b),signal transduction and transcription activator 5b(Stat5b),B cell receptor related protein 31(Bcap31),interferon regulatory factor 7(IRF7),TNF receptor related factor-6(TRAF6),etc.Genes were up-regulated and epithelial cell adhesion molecules(Epcam),C7 and other genes were down-regulated;complements(C3,C6,C9),interferon regulatory factors(IRF2,IRF7),TLR3,NF-?B,Interferon beta(IFN?),interferon-induced Mx(MxB),retinoid-inducible protein I(RIG-I),mitochondrial antiviral signal proteins(MAVs),interleukin 10(IL-10),and Janus proteins Tyrosine kinase 2a(JAK2a)genes were up-regulated while C7 and IRF6 genes were down-regulated(p<0.01).The results of qRT-PCR showed that the expression levels of 23 genes were basically consistent with the quantitative results of iTRAQ protein,and the difference in qRT-PCR valueswas significant(p<0.01).The analysis on the iTRAQ level would help to understand the defense mechanisms of fish skin mucosal immune response to rhabdovirus.