Dante Terino, University of Alberta

Dante Terino, Justine Kniert, Dr. Heather Eaton, Dr. Maya Shmulevitz

Building & Escaping

Like other dsRNA viruses, reoviruses must conceal their genomes within core capsid shells during replication. To confer attachment and stability, cores must acquire an outercapsid (OC). We previously found that core vs. OC assembly is facilitated by three distinct reovirus factories that are temporospatially segregated: core-only factories at the periphery, OC proteins at lipid droplets, and core OC mixing factories where virions assemble. It remained unclear if microtubules (MTs) are involved in the formation of core-only factories from input cores, transition of core-only to OC( ) factories, or the movement of fully assembled virions at the perinuclear space. To explore MTs role in input core distribution in factories, immunofluorescence confocal microscopy (IFCM) was used to visualize fluorescently labeled reovirus cores in cells treated with the nocodazole. At 12hpi input core distribution within OC(-) factories was unaffected in nocodazole-treated cells. To measure MT role in OC(-) to OC( ) factory transition, factories and OC proteins were visualized as before. MT disruption did not affect OC(-) to OC( ) transition at 12hpi, however deposition of new virions at the perinuclear space was halted. To assess how MTs might impact core versus virion assembly rates, RNA expression (core assembly) was measured by RT-qPCR and compared to viral titers (whole virus assembly). Nocodazole treatment delayed and reduced whole virus assembly but had no effect on core assembly. Altogether these results show that MTs do not play a major role in early stages of factory formation, however they are necessary during the final stages of virion assembly and deposition.