Capsid coding sequence is required for efficient replication of human rhinovirus 14 RNA.
Mechanisms by which the plus-sense RNA genomes of picornaviruses are replicated remain poorly defined, but existing models do not suggest a role for sequences encoding the capsid proteins. However, candidate RNA replicons (delta P1 beta gal and delta P1Luc), representing the sequence of human rhinovirus 14 virus (HRV-14) with reporter protein sequences (beta-galactosidase or luciferase, respectively) replacing most of the P1 capsid-coding region, failed to replicate in transfected H1-HeLa cells despite efficient primary cleavage of the polyprotein. To determine which P1 sequences might be required for RNA replication, HRV-14 mutants in which segments of the P1 region were removed to frame from the genome were constructed. Mutants with deletions involving the 5'proximal 1,489 nucleotides of the P1 region replicated efficiently, while those with deletions involving the 3' 1,079 nucleotides did not. Reintroduction of the 3' P1 sequence into the nonreplicating delta P1Luc construct resulted in a new candidate replicon, delta P1Luc/VP3, which replicated well and expressed luciferase efficiently. Capsid proteins provided in trans by helper virus failed to rescue the nonreplicating delta P1Luc genome but were able to package the larger-than-genome-length delta P1Luc/VP3 replicon. Thus, a 3'-distal P1 capsid-coding sequence has a previously unrecognized cis-active function related to replication of HRV-14 RNA.
McKnight
K L
KL
Department of Microbiology and Immunology, University of North Carolina at Chapel Hill 27599-7030, USA.
Lemon
S M
SM
D000595:Amino Acid Sequence; D001483:Base Sequence; D002213:Capsid / Q000235:genetics*; D003001:Cloning, Molecular; D004279:DNA, Viral; D017353:Gene Deletion; D015967:Gene Expression Regulation, Viral; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D012367:RNA, Viral / Q000096:biosynthesis*; D012093:Replicon; D012229:Rhinovirus / Q000235:genetics* / Q000502:physiology; D012260:Ribonucleases / Q000378:metabolism; D014779:Virus Replication / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics
D004279:DNA, Viral; D012367:RNA, Viral; D008156:Luciferases; D012260:Ribonucleases; D001616:beta-Galactosidase