Studies On The Functions Of Unfolded Protein Response And HSP90 On SFTSV Replication

Severe fever with thrombocytopenia syndrome(SFTS)is an emerging,highly pathogenic,infectious disease caused by the infection of a newly discovered tick-borne phlebovirus,SFTS virus(SFTSV),with major clinical symptoms comprising high fever with leukopenia,thrombocytopenia and multi-organ failure.Since its first report in 2010 in China,cases have been found in several provinces in China,and subsequently reported in South Korea and Japan,and the fatality rate can be as high as 30%.SFTSV belongs to the genus Banyangvirus in the family Phenuiviridae of the order Bunyavirales.But limited information on the molecular mechanism of SFTSV infection and pathogenesis impedes the development of effective vaccines and drugs for SFTS prevention and treatment.In this thesis,we studied the interactions between SFTSV and host unfolded protein response(UPR)and heat shock protein 90(HSP90),which shed light onto the molecular mechanism of SFTSV infection.An isobaric tag for relative and absolute quantification(iTRAQ)-based quantitative proteomic analysis of SFTSV-infected HEK-293 cells was performed to explore dynamic host cellular protein responses toward SFTSV infection.The proteomic results highlighted a potential role of endoplasmic reticular stress-triggered unfolded protein response(UPR)in SFTSV infection.Further functional studies confirmed that all three major branches of the UPR,including the the activating transcription factor-6(ATF6),and the inositol-requiring protein-1(IRE1),PKR-like endoplasmic reticulum kinase(PERK)pathways,were activated by SFTSV.However,only the former two pathways play a crucial role in SFTSV infection.Furthermore,expression of SFTSV glycoprotein(GP)alone was sufficient to stimulate the UPR,whereas suppression of PERK and ATF6 notably decreased GP expression.Finally,two other newly discovered Banyangvirus,Heartland virus and Guertu virus,also stimulated the UPR in HEK-293 cells,suggesting a common mechanism shared by these genetically related Banyangvirus.SFTSV can induce inclusion bodies in cytoplasm by non-structural protein NSs during infection,and NSs plays a key role in SFTSV inhibiting IFN-I induction and signaling pathways.HSP90 is an important intracellular molecular chaperone involved in replication of many viruses.In this chapter,we found that HSP90 inhibitors affected SFTSV replication and formation of inclusion bodies induced by NSs.HSP90 inhibitors were used to treat cells overexpressing viral proteins or SFTSV infected cells for a short time,showing NSs not other viral proteins was the main target of HSP90,and HSP90 assisted expression of NSs occuring on translation level.HSP90? could interact with NSs was identified by Co-immunoprecipitation,and whereas knock down expression of HSP90? inhibited SFTSV replication.Finally,we found that the antagonistic effect of SFSTV on IFN-I induction and signaling pathway can be alleviated by HSP90 inhibitors.These results suggest that HSP90 promotes SFTSV replication by assisting the expression of NSs,which can be interfered by HSP90 inhibitors,hinting that HSP90 can be used as a target for drug development in the treatment of SFTSV infection.