Unraveling the Entry Mechanism of Baculoviruses and Its Evolutionary Implications

The entry of baculovirus budded virus into host cells is mediated by two distinct types of envelope fusion proteins (EFPs), GP64 and F protein. Phylogenetic analysis suggested that F proteins were ancestral baculovirus EFPs, whereas GP64 was acquired by progenitor group I alphabaculovirus more recently andmay have stimulated the formation of the group I lineage. This study was designed to experimentally recapitulate a possible major step in the evolution of baculoviruses. We demonstrated that the infec-tivity of an F-null group II alphabaculovirus (Helicoverpa armigera nucleopolyhedrovirus [HearNPV]) can be functionally res-cued by coinsertion of GP64 along with the nonfusogenic Fdef (furin site mutated HaF) fromHearNPV. Interestingly, HearNPV enters cells by endocytosis and, less efficiently, by direct membrane fusion at low pH. However, this recombinant HearNPV co-expressing Fdef and GP64mimicked group I virus not only in its EFP composition but also in its abilities to enter host cells via low-pH-triggered direct fusion pathway. Neutralization assays indicated that the nonfusogenic F proteins contribute mainly to binding to susceptible cells, while GP64 contributes to fusion. Coinsertion of GP64 with an F-like protein (Ac23) from group I virus led to efficient rescue of an F-null group II virus. In summary, these recombinant viruses and their entry modes are consid-ered to resemble an evolutionary event of the acquisition of GP64 by an ancestral group I virus and subsequent adaptive inacti-vation of the original F protein. The study described here provides the first experimental evidence to support the hypothesis of the evolution of baculovirus EFPs. Baculoviruses (Baculoviridae) are a family of rod-shaped vi-ruses with circular, supercoiled, double-stranded DNA ge