RECOMBINANT VARICELLA ZOSTER VIRUS WITH DUAL FLUORESCENT REPORTERS TAGGING DISTINCT VIRION COMPONENTS TO STUDY AXONAL TRANSPORT OF VIRAL PROTEINS IN HUMAN NEURONS

The human herpesvirus varicella zoster virus (VZV) causes chickenpox, and when reactivated, herpes zoster. Rapid retrograde axonal transport of VZV delivers infecting nucleocapsids from peripheral skin and mucosal tissues to ganglionic sensory neuron somata where they establish latency. Anterograde axonal transport delivers virions or their components to the periphery where virus is released to infect cells in reactivation disease. The herpesvirus virion is composed of a DNA-containing nucleocapsid, a glycoprotein-studded lipid envelope and the tegument, an amorphous intermediate layer containing proteins involved in the decision between latent and lytic infection as well as in assembly and axonal transport.

We established a model for studying axonal transport of VZV using human neurons derived from embryonic stem cells in microfluidic chambers. Using this model and a newly generated series of dual fluorescent protein-tagged recombinant VZV we ask: which components of the virion are 1) rapidly transported upon peripheral infection to the neuronal soma, and (2) transported together to the periphery upon reactivation.

VZV nucleocapsids were labelled by tagging ORF23 with GFP or mRFP, and key tegument proteins were simultaneously tagged with the other color fluorescent protein. Analyses of highly purified viral preparations confirmed the presence of both labeled proteins in individual virions. In retrograde transport studies, the transcriptional activator tegument proteins ORF10 and 62 did not co-transport with nucleocapsids. However both proteins were associated with nucleocapsids in the soma and the axon of productively infected neurons. These results demonstrate that entering retrogradely transporting nucleocapsids and assembling anterograde transported virions show differential association with these tegument protein transcriptional activators. Ongoing studies are evaluating co-transport of additional tegument proteins with nucleocapsids.