USE OF FREE-FLOW ELECTROPHORESIS FOR THE ANALYSIS OF CELLULAR UPTAKE OF PICORNAVIRUSES

Free-flow electrophoresis is a powerful tool to separate subcellular v esicles such as early and late endosomes from plasma membranes. Using this technique, the intracellular distribution of poliovirus type 2 Sa bin (PV2) and its derived subviral particles was analyzed upon infecti on of HeLa cells. Comparison of various infection conditions showed th at maximally 30% of total cell associated PV2 was found in endosomal c ompartments with the remainder being associated with plasma membrane f ractions; 2% of viral label was recovered from the cytoplasm in form o f free virions. Sucrose gradient centrifugation analysis of the viral material recovered from the respective fractions revealed that intrace llular virus was exclusively in its native conformation. This is in sh arp contrast to human rhinovirus serotype 2 (HRV2), which is rapidly m odified to RNA-free subviral particles upon accumulation in endosomes. The data suggest that productive poliovirus uncoating can occur at th e plasma membrane whereas internalized virus is most probably aborted.