Flaviviruses (a family of over 90 known viruses, including Zika, dengue and West Nile) are significant
human pathogens affecting 3.1 billion people annually, and most of these enveloped viruses do not have any
viable vaccines or antivirals capable of combating their spread. The primary objective of current flavivirus antiviral
design is to disrupt specific mechanisms in the flavivirus life cycle that are key for virus survival, including virus
attachment to the host cell, viral endocytosis, genome uncoating, genome replication, and virus maturation
through the Golgi. These potentially druggable mechanisms have been explored in great detail both within our
lab and elsewhere. However, little is known about the assembly mechanisms that drive infectious virus particle
formation. Because of the high structural homology among all flaviviruses, the identification of assembly
mechanisms will provide ubiquitous targets for therapeutic intervention in virus proliferation.
In other enveloped virus systems, assembly depends on the interaction of virus core proteins with lipid
membranes or membrane bound glycoproteins to produce infectious virus particles. Exploration via single
particle Cryo-EM reconstruction has shown that the nucleocapsid core (NC) of immature Zika virus, is found in
close proximity with the envelope glycoproteins on the inner side of the virus’s lipid bilayer. Due to this close
proximity, I hypothesized that the NC interacts with the envelope glycoproteins during virus assembly. I further
hypothesized and that these interactions occur between the capsid protein (CP) and transmembrane helices of
the precursor membrane (prM) protein and the envelope (E) protein while the particle is in the immature state.
Since no information currently exists on virus assembly relying on CP-prM/E interactions, these interactions have
the potential to be exploited as new drug targets capable of inhibiting the proliferation of flaviviruses.
To this end, I am investigating two independent strategies to validate the hypothesized NC-prM/E
interactions using dengue virus serotype 2 (DENV2) as a model system. The Kuhn lab is particularly adept in
mutagenesis studies using DENV2, which is why the decision was made to use this virus as our model system
instead of Zika. Firstly, amino acids within the prM/E transmembrane helices that were posited to be key in the
assembly process of DENV2 and other flaviviruses have been or will be mutated to investigate their role in
promoting particle assembly. Secondly, the ability of the DENV2 prM and E transmembrane helices to interact
with CP is being examined through the use of reconstituted prM and E proteins within SMA lipid nanoparticles.
Due to the high similarity of all flaviviruses, the techniques, results and mechanisms identified in this study can
be applied to other flaviviruses. Combining these essential experimental studies with the proposed training skills
and collaborative opportunities for effective scientific communication through writing, speaking and mentoring
will prepare me well for a future in biomedical research.