On the binding and fusion of baculovirus ODV to midgut epithelial cells: distribution and orientation of pif proteins

The initial steps of infection for many enveloped viruses are binding to and fusion with cellular membranes. These processes are carried out in a concerted fashion in a complex of several viral proteins, together called the virus binding/fusion complex. Baculovirus virions exist as two phenotypes: the occlusion derived virus (ODV) and the budded virus (BV). ODVs are occluded into polyhedral occlusion bodies and are responsible for the initial infection of insect midgut epithelial cells. ODVs contain more than 10 different envelope proteins, including the four per os infectivity factors PIF1, PIF2, PIF3 and P74, which are essential for infectivity per os. PIF1, PIF2 and P74 function in virus binding while the function of PIF3 is still unknown. Up until now no fusion protein had been identified for ODV although they are believed to enter the microvillus through direct membrane fusion. P74 is C-terminally anchored in the ODV envelope with its N terminal part outside of the particle. We have employed a combination of immuno-electron microscopy and protein chemistry techniques to study the location, distribution and orientation of PIF1, PIF2 and PIF3 proteins on the ODV particles, using Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) as a experimental model. We could show that PIF1 and PIF2 proteins are present on the outside of the ODV particle and are uniformly distributed along the ODV envelope. Using in vivo crosslinking and mass spectrometry we have searched for viral and host proteins, which interact with PIF proteins. Finally, we will report on the possible role of PIF protein cleavage in the initial stage of ODV infection of midgut epithelial cells.