We have experience producing atomic and near atomic structures from data collected using DED technology and now have access to a newly installed Penn State Titan Krios microscope. The maps were generated with data collected prior to the technological advance of direct electron detection (DED). Hypothesis: Entry mechanisms offer a target for antivirals because rapidly mutating RNA viruses such as DWV will not tolerate changes that alter structures required for host entry.Our preliminary results reveal the gross, overall conformational changes the capsid goes through during entry, even though the maps are of modest (7Å) and low (20Å) resolution. Specifically we propose to:Aim 1: Solve atomic or near atomic resolution maps for each DWV structural intermediate and define the conformational change linked to genome release. The structure-function studies proposed here will elucidate DWV entry with the overarching goal of learning enough about the entry mechanisms to develop antivirals or prophylactics to protect honey bees. Our work so far has shown the conformational change between the open and closed states is linked to genome release. The second conformation is a "closed" state in which the spikes are gathered closely together and appear to form a five-fold channel. The first is the "open" state in which the spikes are positioned further away from the icosahedral five-fold symmetry axis and separated from each other. A prominent feature of the capsid is the VP1 C-terminal extension that forms a spike that takes on two different conformations. Specifically the 3-D maps illustrate conformational changes the capsid undergoes to attach to the host cell and release the genome to establish an infection. Using a structure/function approach, we have solved four DWV structures (preliminary results) that explain steps during assembly and entry. Host entry mechanisms represent a key target for antivirals however, a significant knowledge gap exists in understanding DWV entry into honey bee cells.
Goals / Objectives Deformed wing virus (DWV) is a critically important honey bee pathogen linked to colony collapse.
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