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Journal of Virology, December 2002, p. 12654-12662, Vol. 76, No. 24
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.24.12654-12662.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

RNA Recombination in Brome Mosaic Virus: Effects of Strand-Specific Stem-Loop Inserts

R. C. L. Olsthoorn,1,{dagger} A. Bruyere,1,{ddagger} A. Dzianott,1 and J. J. Bujarski1,2*

Plant Molecular Biology Center, Department of Biological Sciences, Northern Illinois University, DeKalb, Illinois 60115,1 Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan, Poland2

Received 9 May 2002/ Accepted 18 September 2002

A model system of a single-stranded trisegment Brome mosaic bromovirus (BMV) was used to analyze the mechanism of homologous RNA recombination. Elements capable of forming strand-specific stem-loop structures were inserted at the modified 3' noncoding regions of BMV RNA3 and RNA2 in either positive or negative orientations, and various combinations of parental RNAs were tested for patterns of the accumulating recombinant RNA3 components. The structured negative-strand stem-loops that were inserted in both RNA3 and RNA2 reduced the accumulation of RNA3-RNA2 recombinants to a much higher extent than those in positive strands or the unstructured stem-loop inserts in either positive or negative strands. The use of only one parental RNA carrying the stem-loop insert reduced the accumulation of RNA3-RNA2 recombinants even further, but only when the stem-loops were in negative strands of RNA2. We assume that the presence of a stable stem-loop downstream of the landing site on the acceptor strand (negative RNA2) hampers the reattachment and reinitiation processes. Besides RNA3-RNA2 recombinants, the accumulation of nontargeted RNA3-RNA1 and RNA3-RNA3 recombinants were observed. Our results provide experimental evidence that homologous recombination between BMV RNAs more likely occurs during positive- rather than negative-strand synthesis.

* Corresponding author. Mailing address: Plant Molecular Biology Center, Department of Biological Sciences, Northern Illinois University, DeKalb, IL 60115-2861. Phone: (815) 753-0601. Fax: (815) 753-7855. E-mail: jbujarski{at}niu.edu.

{dagger} Present address: Leiden Institute of Chemistry, Gorlaeus Laboratories, Leiden University, Leiden, The Netherlands.

{ddagger} Present address: Qiagen-France, 91974 Courtaboeuf, France.

Journal of Virology, December 2002, p. 12654-12662, Vol. 76, No. 24
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.24.12654-12662.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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