AcMNPV P74 is cleaved at R325 and R334 by proteinases of both OB and BBMV to expose a potential fusion peptide for oral infection.

Baculoviruses enter insect midgut epithelial cells via a set of occlusion-derived virion (ODV) envelope proteins called per os infectivity factors (PIFs). P74 of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), which was the first identified PIF, is cleaved by an endogenous proteinase embedded within the occlusion body during per os infection, but the target site(s) and function of the cleavage have not yet been ascertained. Here, based on bioinformatics analyses, we report that cleavage was predicted at an arginine and lysine-rich region in the middle of P74. A series of recombinant viruses with site-directed mutants in this region of P74 were generated. R325 or R334 was identified as primary cleavage site. In addition, we showed that P74 is also cleaved by brush border membrane vesicles (BBMV) of the host insect at R325 or R334, instead of R195, R196, and R199, as previously reported. Simultaneous mutations in R195, R196, and R199 lead to instability of P74 during ODV release. Bioassays showed that mutations at both R325 and R334 significantly affected oral infectivity. Taken together, our data show that both R325 and R334 of AcMNPV P74 are the primary cleavage site for both occlusion body endogenous proteinase and BBMV proteinase during ODV release and are critical for oral infection. IMPORTANCE: Cleavage of viral envelope proteins is usually an important trigger for viral entry into host cells. Baculoviruses are insect-specific viruses that infect host insects via the oral route. P74, a per os infectivity factor of baculoviruses, is cleaved during viral entry. However, the function and precise cleavage sites of P74 remain unknown. In this study, we found that R325 or R334 between the N- and C-conserved domains of P74 was the primary cleavage site by proteinase either from the occlusion body or host midgut. The biological significance of cleavage seems to be the release of the potential fusion peptide at the N-terminus of the cleaved C-terminal P74. Our results shed light on the cleavage model of P74 and imply its role in membrane fusion in baculovirus per os infection.

Infectious Virions of Bombyx Mori Latent Virus Are Incorporated into Bombyx Mori Nucleopolyhedrovirus Occlusion Bodies.

The Bombyx mori latent virus (BmLV) belongs to the unassigned plant virus family Tymoviridae and contains a positive-sense, single-stranded RNA genome. BmLV has infected almost all B. mori-derived cultured cell lines through unknown routes. The source of BmLV infection and the BmLV life cycle are still unknown. Here, we examined the interaction between BmLV and the insect DNA virus Bombyx mori nucleopolyhedrovirus (BmNPV). Persistent infection with BmLV caused a slight delay in BmNPV propagation, and BmLV propagation was enhanced in B. mori larvae via co-infection with BmNPV. We also showed that BmLV infectious virions were co-occluded with BmNPV virions into BmNPV occlusion bodies. We propose a new relationship between BmLV and BmNPV.