Members of the poxvirus family have been investigated for their applications as vaccines and expressionvectors, and more recently intently studied because of their use as potential biological weapons. Vacciniavirus, the prototypic member, evolves through multiple forms in which membrane morphogenesis plays a keyrole. Even though viruses generally usurp host factors for its use, it remains unclear whether key hosttransport factors that are well known to act in membrane morphogenesis of host cellular compartmentparticipate in early vaccinia membrane morphogenesis, a process critical in acquiring viral infectivity. Wehave now identified a role for coatomer, a host protein complex well characterized to form Coat Protein I(COPI) transport vesicles. However, insights into how it participates in early viral morphogenesis revealsthat the virus bypasses key regulatory mechanisms that form host COPI vesicles, but instead interacts withtwo proteins, the viral K7 and the host KDEL receptor (KDELR) through critical di-Iysine residues on theseinteracting proteins. Thus, to gain further insight into how coatomer and its two interacting proteinscontribute to viral replication, we propose two major aims. In one aim headed by Victor Hsu with proposedfunding from NERCE, perturbation of coatomer will be examined for its effect in potentially accumulatingparticular viral forms. Moreover, the KDELR will be examined with respect to its distribution on viral formsand also whether its ligands have a role in viral replication. As another aim headed by Stuart Isaacs withproposed funding from MARCE, the viral K7 protein will be examine for effects upon its deletion and alsowhen a mutant form that cannot interact with coatomer is expressed.This collaborative effort is responsive in multiple ways to the request for application under the 'NewOpportunities' initiative by the two respective Regional Centers of Excellence. First, we will be gaininginsights into a Category A pathogen, for which we will not only elucidate novel mechanisms by which a hostprotein functions dUring viral replication, but also identify potential key target(s) for the future design ofintervention against the most abundant infectious form of the virus. Second, funding of this proposal willallow one of the main investigators (Victor Hsu) to collaborate with a poxvirus expert (Stuart Isaacs), andthus, applying outside expertise in mechanisms of vesicular transport to critical issues in poxvirus biology.As viruses generally commandeer host mechanisms for their interaction with the host rather than inventingcompletely novel ones, the complementary expertise of the collaboration will likely enhance the elucidation ofhow vaccinia virus interacts with its host. Third, potential funding of this proposal will likely set the stage foran eventual more comprehensive application in the future, such as through the NIH R01 mechanism.
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