Insight into poliovirus genome replication and encapsidation obtained from studies of 3B-3C cleavage site mutants.
A poliovirus (PV) mutant (termed GG), which is incapable of producing 3AB, VPg, and 3CD proteins due to a defective cleavage site between the 3B and 3C proteins, replicated, producing 3BC-linked RNA rather than the VPg-linked RNA produced by the wild type (WT). GG PV RNA is quasi-infectious. The yield of infectious GG PV relative to replicated RNA is reduced by almost 5 logs relative to that of WT PV. Proteolytic activity required for polyprotein processing is normal for the GG mutant. 3BC-linked RNA can be encapsidated as efficiently as VPg-linked RNA. However, a step after genome replication but preceding virus assembly that is dependent on 3CD and/or 3AB proteins limits production of infectious GG PV. This step may involve release of replicated genomes from replication complexes. A pseudorevertant (termed EG) partially restored cleavage at the 3B-3C cleavage site. The reduced rate of formation of 3AB and 3CD caused corresponding reductions in the observed rate of genome replication and infectious virus production by EG PV without impacting the final yield of replicated RNA or infectious virus relative to that of WT PV. Using EG PV, we showed that genome replication and encapsidation were distinct steps in the multiplication cycle. Ectopic expression of 3CD protein reversed the genome replication phenotype without alleviating the infectious-virus production phenotype. This is the first report of a trans-complementable function for 3CD for any picornavirus. This observation supports an interaction between 3CD protein and viral and/or host factors that is critical for genome replication, perhaps formation of replication complexes.
Oh
Hyung Suk
HS
Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, 16802, USA.
Pathak
Harsh B
HB
Goodfellow
Ian G
IG
Arnold
Jamie J
JJ
Cameron
Craig E
CE
D016679:Genome, Viral; D006801:Humans; D009154:Mutation; D017955:Poliovirus / Q000235:genetics*; D020815:Polyproteins; D012367:RNA, Viral; D014764:Viral Proteins; D019065:Virus Assembly; D014779:Virus Replication / Q000235:genetics*
D020815:Polyproteins; D012367:RNA, Viral; D014764:Viral Proteins