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J Virol, July 1998, p. 5870-5876, Vol. 72, No. 7
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Genes Required for Replication of the 15.5-Kilobase RNA Genome of a Plant Closterovirus

Valery V. Peremyslov,1 Yuka Hagiwara,1 and Valerian V. Dolja1,2,*

Department of Botany and Plant Pathology1 and Center for Gene Research and Biotechnology,2 Oregon State University, Corvallis, Oregon 97331

Received 29 December 1997/Accepted 14 April 1998

A full-length cDNA clone of beet yellows closterovirus (BYV) was engineered and used to map functions involved in the replication of the viral RNA genome and subgenomic RNA formation. Among 10 open reading frames (ORFs) present in BYV, ORFs 1a and 1b suffice for RNA replication and transcription. The proteins encoded in these ORFs harbor putative methyltransferase, RNA helicase, and RNA polymerase domains common to Sindbis virus-like viruses and a large interdomain region that is unique to closteroviruses. The papain-like leader proteinase (L-Pro) encoded in the 5'-proximal region of ORF 1a was found to have a dual function in genome amplification. First, the autocatalytic cleavage between L-Pro and the remainder of the ORF 1a product was essential for replication of RNA. Second, an additional L-Pro function that was separable from proteolytic activity was required for efficient RNA accumulation. The deletion of a large, ~5.6-kb, 3'-terminal region coding for a 6-kDa hydrophobic protein, an HSP70 homolog, a 64-kDa protein, minor and major capsid proteins, a 20-kDa protein, and a 21-kDa protein (p21) resulted in replication-competent RNA. However, examination of mutants with replacements of start codons in each of these seven 3'-terminal ORFs revealed that p21 functions as an enhancer of genome amplification. The intriguing analogies between the genome organization and replicational requirements of plant closteroviruses and animal coronavirus-like viruses are discussed.

* Corresponding author. Mailing address: Department of Botany and Plant Pathology, Oregon State University, Cordley Hall 2082, Corvallis, OR 97331. Phone: (541) 737-5472. Fax: (541) 737-3573. E-mail: doljav{at}bcc.orst.edu.

J Virol, July 1998, p. 5870-5876, Vol. 72, No. 7
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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