Poliovirus-membrane interactions during cell entry

Poliovirus initiates infection of host cells by binding to the poliovirus receptor (PVR). PVR not only serves as a docking site for the virus but also regulates entry by triggering a conformational transition in the virus. As a result of this conformational transition, an altered particle with a sedimentation coefficient of 135S (versus 160S for the native virus) is generated. The 135S particle is an intermediate in the poliovirus entry process. An identical conformational change can also be attained in vitro in absence of PVR by heating under specific conditions suggesting that this process requires energy.; The entry of poliovirus also may be regulated through the interaction of the 135S particles with the membranes. To understand the role of the VP4 protein during virus entry, mutants of VP4 at threonine 28 were studied. A non-viable point mutant, 4028T.G, fails to deliver RNA into the host cells whereas related viable mutants, 4028T.S and 4028T.V, are capable of delivering RNA albeit with altered kinetics compared to the wild type virus. The defect in these mutants is at a stage after the formation of the 135S particle. The VP4 protein from all the virus strains tested is associated with the membrane of the infected cells and remains associated with the membrane even as other viral capsid proteins are progressively lost. Examination of the interaction of the virus with lipid bilayers using electrophysiologic techniques show that unlike the WT virus, the non-viable 4028T.G mutant failed to form ion channels. The viable mutants (4028T.S and 4028T.V) were capable of forming ion channels, however, the channels formed had drastically different electrical characteristics. Thus, the ability of the virus to form ion channels in bilayers appears to correlate with its ability to deliver RNA in host cells and the channel properties may affect the kinetics of RNA delivery. Additionally, because channel properties are altered by a single amino acid substitution in the VP4 protein, the VP4 protein is most likely a constituent of the ion channel.; Although viral domains that are involved in the membrane interaction of the 135S particle have been identified, cellular components also may be important at this stage. Based on the essential role for the interaction of the myristylated VP4 protein with the membrane, the importance of cholesterol-rich, detergent-insoluble membrane (DIMs) domains in viral entry was examined. Methyl-β-cyclodextrin (MβCD) treatment prevents infection of 90% of the cells. This inhibition of infection occurs after the receptor-mediated conformational transition to the 135S particle possibly due to removal of cellular cholesterol. However, unlike other viruses that are affected by MβCD treatment, neither the poliovirus receptor, nor the capsid proteins of the infecting poliovirus are specifically localize to the DIMs. Furthermore their localization is unaffected by MβCD treatment. Therefore MβCD treatment may affect RNA delivery by qualitatively altering the interaction of the virus with the membrane or by other unknown mechanisms.