Requirement for a viral trans-acting factor encoded by brome mosaic virus RNA-2 provides strong selection in vivo for functional recombinants.
Interaction of specific nucleotide sequences with trans-acting proteins is intrinsic to replication of viral as well as eucaryotic genomes. Brome mosaic virus RNA-2 encodes one of the two viral proteins known to be essential for replication. (R. French, M. Janda, and P. Ahlquist, Science 231:1294-1297, 1986; P. A. Kiberstis, L. S. Loesch-Fries, and T. C. Hall, Virology 112:804-808, 1981). Transfection of barley protoplasts with wild-type transcripts of brome mosaic virus RNA-1 and RNA-3 and serial dilutions of RNA-2 transcripts possessing unaltered coding sequences but bearing mutations that greatly incapacitated replication of RNA-2 revealed that trace amounts of RNA-2 are sufficient to support replication of the viral genome. In six replicate experiments containing RNA-2 transcripts devoid of the 3' 200 nucleotides that encompass the tRNA-like structure containing the minus-strand promoter, detectable levels of progeny RNA-1 and RNA-3 and subgenomic RNA-4 were present. This showed that viral p2 protein translated from the supplied RNA-2 functioned in trans to support replication of RNA-1 and RNA-3. However, in two similar experiments, progeny RNA-2 with electrophoretic mobility indistinguishable from that of wild-type RNA-2 was seen at 24 h postinoculation. Northern hybridization (RNA blot) analysis confirmed the presence of the tRNA-like 3' terminus on these progeny RNAs, indicating that recombinational restoration of the deleted sequence had occurred. This suggests that, under certain circumstances, RNA recombination may be a rapid and frequent phenomenon.
Rao
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Biology Department, Texas A&M University, College Station 77843-3258.
Hall
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TC
D001483:Base Sequence; D005814:Genes, Viral; D001467:Hordeum; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000235:genetics* / Q000502:physiology; D009154:Mutation; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D012367:RNA, Viral / Q000235:genetics* / Q000302:isolation & purification; D011995:Recombination, Genetic; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics; D014779:Virus Replication
D012367:RNA, Viral; D015534:Trans-Activators; D014764:Viral Proteins