A Preliminary Study On The Autophagic Response Induced By AcMNPV Infection In Sf9 Cells

Autophagy is an important protective mechanism of organism. Recent studies have shown that autophagy plays as an immune defense system and helps to clear microbial and viral pathogens by directly degrading microorganisms and preventing virus replicating. Meanwhile, pathogens develop strategies to escape from and even take advantage of autophagy immune response. As reported in mammals and Drosophila, autophagy is triggered by virus infection and contributes to immune response. But the induction mechanisms of autophagy and pathogens resistance are still unknown.Autographa californica nucleopolyhedrovirus(Ac MNPV) is a pathogen of Spodoptera frugiperda. But there has been no report about whether Ac MNPV can induce autophagical response in S. frugiperda. In this study, we used two recombinant baculovirus, GFP-Ac MNPV and Ds Red-Ac MNPV, to treat Spodoptera frugiperda(Sf9) cells and investigated if the autophagy is able to be induced. Moreover, we also examined whether and how the autophagy affects Ac MNPV. The main results are as below: 1 The virus dose results indicated that the TCID50 of GFP-Ac MNPV and Ds Red-Ac MNPV were 1.26×106 /m L and 9.35×107 /m L, respectively. These two recombinant baculovirus could proliferate in Sf9 cells after infection. 2 Lyso-Tracker Red staining showed that autolysosomes were significantly increased in Sf9 cells after 24 h infected by GFP-Ac MNPV. 3 Sf9 cells undergo autophagy after 48 h infected with Ds Red-Ac MNPV:(1)Autophagic vesicles were observed in Sf9 cells after Ds Red-Ac MNPV infected for 48h;(2)After transient overexpression of p IEX-4-EGFP-Bm Atg8 and followed by Ds Red-Ac MNPV infection for 48 h, punctates of EGFP-Bm Atg8-PE aggregation were more than in the non-treated control cells; And EGFP-ATG8-PE protein was significantly detected by western blotting in Ds Red-Ac MNPV-infected cells as in starved cells. 4 EGFP-ATG8-PE protein was also detected in UV-Ds Red-Ac MNPV-infected cells, indicating that UV-Ds Red-Ac MNPV also induced autophagy in Sf9 cells and viral capsid protein may be the factor to trigger autophagy. 5 Ds Red-Ac MNPV could continue to grow in Sf9 cells after autophagy was triggered. Furthermore, the viral titer significantly upregulated when the fusion of autophagosomes and lysosomes was prevented by chloroquine. This result implied that virus could escape from autophagy.In conclusion, we showed that Ac MNPV can induce autophagy in Sf9 cells and the capsid protein may be the factor. Moreover, we also found that Ac MNPV could escape from autophagy and keep on proliferating. But the detailed mechanism of the virus-induced autophagy needs to be further investigated.