Functional conservation of the hydrophobic domain of polypeptide 3AB between human rhinovirus and poliovirus

In this study we exchanged portions of the poliovirus type 1 (PV1) hydrophobic domain within the membrane-associated polypeptide 3AB for the analogous sequences from human rhinovirus 14 (HRV14). The sequence exchanges were based upon a previous report in which the 22 amino acid hydrophobic region wa...

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Bibliographic Details
Published in:Virology (New York, N.Y.) N.Y.), 2003-09, Vol.314 (1), p.432-442
Main Authors: Towner, Jonathan S, Brown, David M, Nguyen, Joseph H.C, Semler, Bert L
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Amino Acid Substitution
Base Sequence
HeLa Cells
Humans
Molecular Sequence Data
Peptides - chemistry
Peptides - genetics
Peptides - metabolism
Poliovirus - genetics
Poliovirus - physiology
Protein Biosynthesis
Rhinovirus - genetics
Rhinovirus - physiology
RNA, Viral - biosynthesis
Transfection
Viral Core Proteins - chemistry
Viral Core Proteins - genetics
Viral Core Proteins - metabolism
Viral Plaque Assay
Virus Replication
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Summary:In this study we exchanged portions of the poliovirus type 1 (PV1) hydrophobic domain within the membrane-associated polypeptide 3AB for the analogous sequences from human rhinovirus 14 (HRV14). The sequence exchanges were based upon a previous report in which the 22 amino acid hydrophobic region was subdivided into two domains, I and II, the latter of which was shown to be required for membrane association (J. Biol. Chem. 271 (1996), 26810). Using these divisions, the HRV14 sequences were cloned into the complete poliovirus type 1 cDNA sequence. RNAs transcribed from these cDNAs were transfected into HeLa cell monolayers and used in HeLa cell-free translation/replication assays. The data indicated that 3AB sequences from PV1 and HRV14 are interchangeable; however, the substitutions cause a range of significant RNA replication defects, and in some cases, protein processing defects. Following transfection of RNAs encoding the domain substitutions into HeLa cell monolayers, virus isolates were harvested, and the corresponding viral RNAs were sequenced. The sequence data revealed that for the carboxy-terminal domain substitutions (domain II), multiple nucleotide changes were identified in the first, second, and third positions of different codons. In addition, the data indicated that for one of the PV1/HRV14 chimeras to replicate, compensatory mutations within poliovirus protein 2B may be required.
ISSN:0042-6822
1096-0341
DOI:10.1016/S0042-6822(03)00448-3