Genome Delivery and Ion Channel Properties Are Altered in VP4 Mutants of Poliovirus

Abstract

During entry into host cells, poliovirus undergoes a receptor-mediated conformational transition to form 135S particles with irreversible exposure of VP4 capsid sequences and VP1 N termini. To understand the role of VP4 during virus entry, the fate of VP4 during infection by site-specific mutants at threonine-28 of VP4 (4028T) was compared with that of the parental Mahoney type 1 virus. Three virus mutants were studied: the entry-defective, nonviable mutant 4028T.G and the viable mutants 4028T.S and 4028T.V, in which residue threonine-28 was changed to glycine, serine, and valine, respectively. We show that mutant and wild-type (WT) VP4 proteins are localized to cellular membranes after the 135S conformational transition. Both WT and viable 4028T mutant particles interact with lipid bilayers to form ion channels, whereas the entry-defective 4028T.G particles do not. In addition, the electrical properties of the channels induced by the mutant viruses are different from each other and from those of WT Mahoney and Sabin type 3 viruses. Finally, uncoating and/or cytoplasmic delivery of the viral genome is altered in the 4028T mutants: the 4028T.G lethal mutant does not release its genome into the cytoplasm, and genome delivery is slower during infection by mutant 4028T.V 135S particles than by mutant 4028T.S or WT 135S particles. The distinctive electrical characteristics of the different 4028T mutant channels indicate that VP4 sequences might form part of the channel structure. The different entry phenotypes of these VP4 mutants suggest that the ion channels may be related to VP4's role during genome uncoating and/or delivery.Poliovirus, a member of the Picornaviridae family, encapsidates its 7,400-nucleotide positive-sensed RNA genome within an icosahedrally symmetric protein shell that is formed by 60 copies of the four capsid proteins (VP1 to VP4). VP1, VP2, and VP3 form the surface of the virion, with VP1 located at each fivefold axis and VP2 and VP3 alternately positioned around each threefold axis. VP4 in its entirety as well as the amino termini of VP1, VP2, and VP3 are buried within the interior of the capsid, lying along the inner surface of the virion shell (12).Poliovirus entry into cells is initiated by binding to the poliovirus receptor (PVR) on the cell surface. PVR binding induces a conformational transition within the virus particle that leads to formation of altered particles (termed A particles) sedimenting at 135S (versus the 160S sedimentation value of the native particle) (references 8 and 23 and references therein). This conformational transition results in relocation of VP4 and the VP1 N termini from the particle interior to the virion exterior. The appearance of VP4 and VP1 domains on the particle surface is correlated with dramatic differences in the functional behavior of the native 160S and altered 135S particles. Functionally, these PVR-induced conformational rearrangements generate 135S particles that acquire the ability to bind to liposomes, to form ion channels in lipid bilayers, and ...

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