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Homolog-scanning mutagenesis reveals poliovirus receptor residues important for virus binding and replication
M E Morrison1, Y J He, M W Wien
1Department of Microbiology, Columbia University College of Physicians & Surgeons, New York, New York 10032.
Abstract:
Poliovirus initiates infection of primate cells by binding to the poliovirus receptor, Pvr. Mouse cells do not bind poliovirus but express a Pvr homolog, Mph, that does not function as a poliovirus receptor. Previous work has shown that the first immunoglobulin-like domain of the Pvr protein contains the virus binding site. To further identify sequences of Pvr important for its interaction with poliovirus, stable cell lines expressing mutated Pvr molecules were examined for their abilities to bind virus and support virus replication. Substitution of the amino-terminal domain of Mph with that of Pvr yields a molecule that can function as a poliovirus receptor. Cells expressing this chimeric receptor have normal binding affinity for poliovirus, yet the kinetics of virus replication are delayed. Results of virus alteration assays indicate that this chimeric receptor is defective in converting native virus to 135S altered particles. This defect is not observed with cells expressing receptor recombinants that include Pvr domains 1 and 2. Because altered particles are believed to be an intermediate in poliovirus entry, these findings suggest that Pvr domains 2 and 3 participate in early stages of infection. Additional mutants were made by substituting variant Mph residues for the corresponding residues in Pvr. The results were interpreted by using a model of Pvr predicted from the known structures of other immunoglobulin-like V-type domains. Analysis of stable cell lines expressing the mutant proteins revealed that virus binding is influenced by mutations in the predicted C'-C" loop, the C" beta-strand, the C"-D loop, and the D-E loop. Mutations in homologous regions of the immunoglobulin-like CD4 molecule alter its interaction with gp120 of human immunodeficiency virus type 1. Cells expressing Pvr mutations on the predicted C" edge do not develop cytopathic effect during poliovirus infection, suggesting that poliovirus-induced cytopathic effect may be induced by the virus-receptor interaction.
Insights
Poliovirus receptor (Pvr) interaction with poliovirus involves specific domains beyond initial binding. Domains 2 and 3 of Pvr are crucial for early infection stages and viral replication, influencing cytopathic effects.
Area of Science:
- Virology
- Molecular Biology
- Immunology
Background:
- Poliovirus infects primate cells via the poliovirus receptor (Pvr).
- Mouse cells have a Pvr homolog (Mph) that doesn't bind poliovirus.
- The first immunoglobulin-like domain of Pvr contains the virus binding site.
Purpose of the Study:
- To identify specific Pvr sequences critical for poliovirus interaction.
- To understand the role of different Pvr domains in viral entry and replication.
Main Methods:
- Creation and analysis of stable cell lines expressing mutated and chimeric Pvr molecules.
- Assessing poliovirus binding affinity and replication kinetics.
- Performing virus alteration assays to study viral particle conversion.
Main Results:
- A chimeric receptor with Pvr's amino-terminal domain enabled poliovirus binding but delayed replication.
- This chimera was defective in converting native virus to 135S altered particles, unlike Pvr domains 1 and 2 recombinants.
- Mutations in specific Pvr loops and strands (C -C", C" beta-strand, C"-D, D-E) affected virus binding.
- Pvr mutations on the C" edge prevented cytopathic effect.
Conclusions:
- Pvr domains 2 and 3 are essential for early poliovirus infection stages, including viral entry.
- Specific structural regions of Pvr, beyond the primary binding site, regulate viral replication and infection outcomes.
- The virus-receptor interaction may directly induce poliovirus-mediated cytopathic effects.