Splicing factor SF3B3,a NS5-binding protein,restricts ZIKV infection by targeting GCH1

Zika virus(ZIKV),a positive-sense single-stranded RNA virus,causes congenital ZIKV syndrome in children and Guillain-Barre Syndrome(GBS)in adults.ZIKV expresses nonstructural protein 5(NS5),a large protein that is essential for viral replication.ZIKV NS5 confers the ability to evade interferon(IFN)signalling;however,the exact mechanism remains unclear.In this study,we employed affinity pull-down and liquid chromatography-tandem mass spectrometry(LC-MS/MS)analyses and found that splicing factor 3b subunit 3(SF3B3)is associated with the NS5-Flag pull-down complex through interaction with NS5.Functional assays showed that SF3B3 overexpression inhibited ZIKV replication by promoting IFN-stimulated gene(ISG)expression whereas silencing of SF3B3 inhibited expression of ISGs to promote ZIKV replication.GTP cyclohydrolase I(GCH1)is the first and ratelimiting enzyme in tetrahydrobiopterin(BH4)biosynthesis.NS5 upregulates the expression of GCH1 during ZIKV infection.And GCH1 marginally promoted ZIKV replication via the IFN pathway.Additionally,GCH1 expression is related to the regulation of SF3B3.Overexpression of the SF3B3 protein effectively reduced GCH1 protein levels,whereas SF3B3 knockdown increased its levels.These findings indicated that ZIKV NS5 binding protein SF3B3 contributed to the host immune response against ZIKV replication by modulating the expression of GCH1.

Non-Structural Protein 5 of Zika Virus Interacts with p53 in Human Neural Progenitor Cells and Induces p53-Mediated Apoptosis

Zika virus(ZIKV) infection could disrupt neurogenesis and cause microcephaly in neonates by targeting neural progenitor cells(NPCs). The tumor suppressor p53-mediated cell cycle arrest and apoptotic cell death have been suggested to be activated upon ZIKV infection, yet the detailed mechanism is not well understood. In the present study, we investigated the effects of ZIKV-encoded proteins in the activation of p53 signaling pathway and found that, among the ten viral proteins,the nonstructural protein 5(NS5) of ZIKV most significantly activated the transcription of p53 target genes. Using the immunoprecipitation-coupled mass spectrometry approach, we identified that ZIKV-NS5 interacted with p53 protein. The NS5-p53 interaction was further confirmed by co-immunoprecipitation and GST pull-down assays. In addition, the MTase domain of NS5 and the C-terminal domain of p53 were mapped to be responsible for the interaction between these two proteins. We further showed that ZIKV-NS5 was colocalized with p53 and increased its protein level in the nuclei and able to prolong the half-life of p53. Furthermore, lentivirus-mediated expression of ZIKV-NS5 in hNPCs led to an apparent cell death phenotype. ZIKV-NS5 promoted the cleavage of PARP1 and significantly increased the cell apoptosis of h NPCs.Taken together, these findings revealed that ZIKV-NS5 is a previously undiscovered regulator of p53-mediated apoptosis in hNPCs, which may contribute to the ZIKV-caused abnormal neurodevelopment.