Study On Interaction Of Japanese Enceplalitis Virus NS1 Protein With Host HAX1 Protein

Japanese encephalitis(JE),referred to as JE,a zoonosisinfectious diseasewith human central nervous system which is a major public health threat to infectious diseases.causative agentis the Japanese encephalitis virus(Japanese encephalitis virus,JEV)which is belongs to flaviviridae.Now,the JE mainly report to be endemic in a largearea of Asia and transmitted primarilyby C ulex species,particularly Culex tritaeniorhynchus.The natural ecological cycle of Japanese encephalitis virus mainly involves virus circulation betweenmosquitoes and avian species.Pigs are the most important intermediate host and amplifying host of encephalitis virus.Infected with JE will have a huge impact on the productionperformance of pigs and will bring huge economic losses to the pig industry.In the life cycle of virus,there is a wide range of interactions between viruses and host protein and resolving the interaction between the host protein and the viral protein is critical to exploring the virus life cycle.comprehensive understandingthe life cycle of the virus can provide a reference for the prevention and treatment of diseases.The NS1 protein of encephalitis virus,a kind of protein which is related to membrane function with a different forms in intracellular and extracellular,is the only protein which can be secreted into extracellular.NS1 protein has complement-binding activity which can stimulate the host to produce protective antibody response.Many researchers have speculated that JEV NS1 protein mayplays an important role in the life cycle.Such as immune invasion,escape,viral replication and translation.However,its specific function remains unclear.In this study we focused on JEV NS1 protein,and NS1 gene was cloned into eukaryotic expression vector,at the same time the eukaryotic expression vector of host HAX1 protein was also constructed.Transfection of two plasmids into 293 T cells and carried out by immunoprecipitation.The results showed that the BJB NS1 protein interacted with the host HAX1 protein.Our further findings suggestted that NS1 interacts with HAX1 203-279 amino acids.Confocal results showed that the NS1 protein and HAX1 protein were co-located in the cytoplasm,and the proteins interacting complex were accumulated in the cytoplasm.Furthermore,interference the expression of HAX1 by RNAi showed that the proliferation of JEV without Significant change.Forthermore,in order to explore the interaction between HAX1 and JEV NS1 in the JEV life cyclewe also used a new generation of gene editing technology CRISPR / Cas9 to screen o ut the knockout HAX1 cell line.In summary,our study confirmed the interaction between NS1 and HAX1 whicht he interaction region is 203-279 amino acids of HAX1.Confocal results showed that the NS1 protein and HAX1 protein were co-locatedand accumulated in the cytoplasm.Furthermore,we screen out the knockout HAX1 cell line by CRISPR/Cas9.