Temperature-induced SVCV-infected EPC Cells In Proteomic Analysis And Effect Of Hsp70 On SVCV Replication

Temperature is related to the growth of animals,but also related to the occurrence of diseases.Temperature as a cause of viral infection can affect the replication process of some viruses.For Spring Viraemia of Carp Virus(SVCV),the temperature of fish body is 16℃-17℃,high temperature or low temperature is not easy to onset.Few study on the effect of temperature on SVCV infection.Heat shock proteins,as important temperature-effect proteins,are involved in the infection process of many viruses,and have less research on the effects of SVCV replication.In this study,this experiment explored the effects of different temperatures on the activity of SVCV-infected carp epithelial cells(EPC)and the changes in proteomics.Through screening validation,the effects of Hsp70 replication were explored,and the results were helpful to understand the pathogenesis of SVCV.To provide new strategies for the treatment and prevention of SVCV in the next step.1.Effect of temperature on the activity and apoptosis of EPC cells infected with SVCV.The cell viability was measured by MTT method.The cell treatment temperature was 16 °C,21 °C,26 °C,31 °C and 36 °C for 12 h,24h and 36 h.Compared with 26°C,the temperature increased with the increase of time.The activity gradually decreased;after infection with the virus,the cell activity increased with time at a high temperature,and the decrease was more significant;the virus titer showed the highest titer at 31 °C.At 26 °C and 31 °C,the infection time was 24 h,and the apoptosis was measured.After SVCV infection of EPC cells,the proportion of dead cells and late apoptosis was significantly increased.The effect was higher when the temperature was increased.The expression of SVCV-G gene was observed at 31 °C.2.Temperature analysis of SVCV-infected EPC cells by proteomicsProteomics changes were determined at 26 °C and 31 °C with an infection time of 24 h.The results show: 35 differential proteins were found after the comparison between 26 ℃ SVCV and 26 ℃groups,of which 19 were up-regulated and 16 were down-regulated;786 differential proteins were found after the comparison between 31 ℃ SVCV and 31 ℃ groups,of which 383 were up-regulated and 403 were down-regulated;624 differential proteins were found after the comparison between 31 ℃ SVCV and 26 ℃ SVCV groups,of which 314 were up-regulated and 310 were down-regulated;624differential proteins were found after the comparison between 31 ℃ and 26 ℃ groups,of which 314 were up-regulated and 310 were down-regulated;220 differential proteins were found after the comparison between 31 ℃ and 26 ℃ groups,of which 93 were up-regulated and 127 were down-regulated.The main functional annotations of differential proteins were concentrated,such as regulation of insulin secretion involved in cellular response,cell redox homeostasis,spindle assembly,DNA replication initiation;differential protein enrichment pathways are mainly enriched in Protein processing in endoplasmic reticulum,Glutamatergic synapse,Melanogenesis.partial differences in Protein processing in endoplasmic reticulum Protein validation was consistent with proteomics.3.Effects of Hsp70 knockdown on SVCV replicationScreening for proteomics results,knockdown of Hsp70 gene in the Protein processing in endoplasmic reticulum;The results showed that Hsp70 knockdown significantly regulated the expression of TLR3,IRF3 and IFN-β,and did not significantly regulate TLR9,IRF7 and IFN-α;Hsp70 knockdown significantly increased the expression of SVCV-G,possibly with Hsp70 regulating TLR3 and The amount of IRF3 expression is related to the decreased expression of IFN-β.