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Published online 20 May 2004

Nucleic Acids Research, 2004, Vol. 32, No. 9 2819-2828
© 2004 Oxford University Press

Structural characterization of an intermolecular RNA–RNA interaction involved in the transcription regulation element of a bipartite plant virus

Richard H. Guenther, Tim L. Sit, Hanna S. Gracz1, Michael A. Dolan1, Hannah L. Townsend1, Guihua Liu1, Winnell H. Newman1, Paul F. Agris1 and Steven A. Lommel*

Department of Plant Pathology, Box 7616 and 1 Department of Molecular and Structural Biochemistry, Box 7622, North Carolina State University, Raleigh, NC 27695, USA

*To whom correspondence should be addressed. Tel: +1 919 515 6992; Fax: +1 919 515 7716; Email: steve_lommel{at}ncsu.edu
Present addresses:
Michael A. Dolan, Tripos Inc., St Louis, MO 63144, USA
Guihua Liu, Department of Chemistry, China Medical University, Shenyang, China

Received January 13, 2004; Revised March 9, 2004; Accepted April 14, 2004

The 34-nucleotide trans-activator (TA) located within the RNA-2 of Red clover necrotic mosaic virus folds into a simple hairpin. The eight-nucleotide TA loop base pairs with eight complementary nucleotides in the TA binding sequence (TABS) of the capsid protein subgenomic promoter on RNA-1 and trans-activates subgenomic RNA synthesis. Short synthetic oligoribonucleotide mimics of the RNA-1 TABS and the RNA-2 TA form a weak 1:1 bimolecular complex in vitro with a Ka of 5.3 x 104 M–1. Ka determination for a series of RNA-1 and RNA-2 mimic variants indicated optimum stability is obtained with seven-base complementarity. Thermal denaturation and NMR show that the RNA-1 TABS 8mers are weakly ordered in solution while RNA-2 TA oligomers form the predicted hairpin. NMR diffusion studies confirmed RNA-1 and RNA-2 oligomer complex formation in vitro. MC-Sym generated structural models suggest that the bimolecular complex is composed of two stacked helices, one being the stem of the RNA-2 TA hairpin and the other formed by the intermolecular base pairing between RNA-1 and RNA-2. The RCNMV TA structural model is similar to those for the Simian retrovirus frameshifting element and the Human immunodeficiency virus-1 dimerization kissing hairpins, suggesting a conservation of form and function.


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Home page
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M. Tatsuta, H. Mizumoto, M. Kaido, K. Mise, and T. Okuno
The Red Clover Necrotic Mosaic Virus RNA2 trans-Activator Is Also a cis-Acting RNA2 Replication Element
J. Virol., January 15, 2005; 79(2): 978 - 986.
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