Human rhinovirus HRV14 uncoats from early endosomes in the presence of bafilomycin.

Determination of infectious progeny virus and in vivo labelling with [(35)S]methionine followed by immunoprecipitation demonstrates that the major receptor group human rhinovirus HRV14 is able to infect HeLa cells in the presence of the V-ATPase inhibitor bafilomycin A1. However, host cell shut off is delayed and viral yield is decreased in the presence of the drug. Uncoating can thus take place under conditions that prevent endosomal acidification indicating that it is catalysed by the viral receptor alone.

Effect of bafilomycin A1 and nocodazole on endocytic transport in HeLa cells: implications for viral uncoating and infection.

Bafilomycin A1 (baf), a specific inhibitor of vacuolar proton ATPases, is commonly employed to demonstrate the requirement of low endosomal pH for viral uncoating. However, in certain cell types baf also affects the transport of endocytosed material from early to late endocytic compartments. To characterize the endocytic route in HeLa cells that are frequently used to study early events in viral infection, we used 35S-labeled human rhinovirus serotype 2 (HRV2) together with various fluid-phase markers.

Uptake of poliovirus into the endosomal system of HeLa cells.

To understand the topology and mechanism of poliovirus uncoating, the question of whether intact virions can be endocytosed by the host cell was studied by a combination of various techniques. In order to prevent alteration of the virus to subviral particles, Hela cells were infected at 26 degrees C. At this temperature the majority of cell-associated virions remained at the plasma membrane, whereas a smaller amount accumulated in vesicles having the same mobility (upon free-flow electrophoresis) and migration behaviour on Nycodenz density gradients as early and late endosomes.

Use of free-flow electrophoresis for the analysis of cellular uptake of picornaviruses.

Free-flow electrophoresis is a powerful tool to separate subcellular vesicles such as early and late endosomes from plasma membranes. Using this technique, the intracellular distribution of poliovirus type 2 Sabin (PV2) and its derived subviral particles was analyzed upon infection of HeLa cells. Comparison of various infection conditions showed that maximally 30% of total cell associated PV2 was found in endosomal compartments with the remainder being associated with plasma membrane fractions; 2% of viral label was recovered from the cytoplasm in form of free virions.

Major and minor receptor group human rhinoviruses penetrate from endosomes by different mechanisms.

Intercellular adhesion molecule 1 and the low-density lipoprotein receptor are used for cell entry by major and minor receptor group human rhinoviruses (HRVs), respectively. Whereas minor-group viruses, exemplified by HRV2, transfer their genomic RNA to the cytoplasm through a pore in the endosomal membrane (E. Prchla, C.

Virus-mediated release of endosomal content in vitro: different behavior of adenovirus and rhinovirus serotype 2.

Endosomal penetration by nonenveloped viruses might be accomplished by either local breakdown of the endosomal membrane (e.g., adenovirus) or formation of a membrane-spanning pore by capsid proteins.

Uncoating of human rhinovirus serotype 2 from late endosomes.

The internalization pathway and mechanism of uncoating of human rhinovirus serotype 2 (HRV2), a minor-group human rhinovirus, were investigated. Kinetic analysis revealed a late endosomal compartment as the site of capsid modification from D to C antigenicity. The conformational change as well as the infection was prevented by the specific V-ATPase inhibitor bafilomycin A1.