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Journal of Virology, July 2007, p. 7716-7724, Vol. 81, No. 14
0022-538X/07/$08.00+0     doi:10.1128/JVI.00549-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

An RNA Stem-Loop within the Bovine Coronavirus nsp1 Coding Region Is a cis-Acting Element in Defective Interfering RNA Replication{triangledown}

Cary G. Brown,{dagger} Kimberley S. Nixon, Savithra D. Senanayake,{ddagger} and David A. Brian*

Departments of Microbiology and Pathobiology, University of Tennessee College of Veterinary Medicine, Knoxville, Tennessee 37996-0845

Received 15 March 2007/ Accepted 26 April 2007

Higher-order cis-acting RNA replication structures have been identified in the 3'- and 5'-terminal untranslated regions (UTRs) of a bovine coronavirus (BCoV) defective interfering (DI) RNA. The UTRs are identical to those in the viral genome, since the 2.2-kb DI RNA is composed of only the two ends of the genome fused between an internal site within the 738-nucleotide (nt) 5'-most coding region (the nsp1, or p28, coding region) and a site just 4 nt upstream of the 3'-most open reading frame (ORF) (the N gene). The joined ends of the viral genome in the DI RNA create a single continuous 1,635-nt ORF, 288 nt of which come from the 738-nt nsp1 coding region. Here, we have analyzed features of the 5'-terminal 288-nt portion of the nsp1 coding region within the continuous ORF that are required for DI RNA replication. We observed that (i) the 5'-terminal 186 nt of the nsp1 coding region are necessary and sufficient for DI RNA replication, (ii) two Mfold-predicted stem-loops within the 186-nt sequence, named SLV (nt 239 to 310) and SLVI (nt 311 to 340), are supported by RNase structure probing and by nucleotide covariation among closely related group 2 coronaviruses, and (iii) SLVI is a required higher-order structure for DI RNA replication based on mutation analyses. The function of SLV has not been evaluated. We conclude that SLVI within the BCoV nsp1 coding region is a higher-order cis-replication element for DI RNA and postulate that it functions similarly in the viral genome.

* Corresponding author. Mailing address: Department of Microbiology, University of Tennessee, Knoxville, TN 37996-0845. Phone: (865) 974-4030. Fax: (865) 974-4007. E-mail: dbrian{at}utk.edu

{triangledown} Published ahead of print on 2 May 2007.

{dagger} Present address: University of Tennessee, Chattanooga, Chattanooga, TN 37408.

{ddagger} Present address: Crowder College, Neosho, MO 64850.

Journal of Virology, July 2007, p. 7716-7724, Vol. 81, No. 14
0022-538X/07/$08.00+0     doi:10.1128/JVI.00549-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Gustin, K. M., Guan, B.-J., Dziduszko, A., Brian, D. A. (2009). Bovine Coronavirus Nonstructural Protein 1 (p28) Is an RNA Binding Protein That Binds Terminal Genomic cis-Replication Elements. J. Virol. 83: 6087-6097 [Abstract] [Full Text]  


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