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Journal of Virology, September 2006, p. 9336-9340, Vol. 80, No. 18
0022-538X/06/$08.00+0     doi:10.1128/JVI.00630-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

An Element of the Tertiary Structure of Peach Latent Mosaic Viroid RNA Revealed by UV Irradiation

Carmen Hernández,1 Francesco Di Serio,2 Silvia Ambrós,3 José-Antonio Daròs,1 and Ricardo Flores2*

Instituto de Biología Molecular y Celular de Plantas (UPV-CSIC), Universidad Politécnica de Valencia, Avenida de los Naranjos s/n, 46022 Valencia, Spain,1 Istituto di Virologia Vegetale del CNR, Sede di Bari, 70126 Bari, Italy,2 Instituto Valenciano de Investigaciones Agrarias, Moncada 46113, Valencia, Spain3

Received 29 March 2006/ Accepted 30 June 2006

Following UV irradiation, denaturing polyacrylamide gel electrophoresis and Northern blot hybridization revealed a cross-link in Peach latent mosaic viroid (PLMVd) plus-strand RNA. Primer extension and partial alkaline hydrolysis of the UV-irradiated PLMVd plus-strand RNA resulting from the hammerhead-mediated self-cleavage mapped the cross-link at U81 and at the 3'-terminal C289 (or at a very proximal nucleotide). Supporting this notion, in vitro-synthesized PLMVd plus-strand RNAs with short insertions/deletions at their 3' termini failed to cross-link. Because U81 and C289 are conserved in PLMVd variants and because the initiation site of PLMVd minus-strand RNA maps at a short double-stranded motif containing C289, the UV-photo-cross-linkable element of tertiary structure may be functionally significant. A second cross-linked species similar in size and sequence to the monomeric circular PLMVd form, observed in some PLMVd variants, probably derives from UV-induced ligation of the two termini resulting from self-cleavage.

* Corresponding author. Mailing address: Instituto de Biología Molecular y Celular de Plantas (UPV-CSIC), Universidad Politécnica de Valencia, 46022 Valencia, Spain. Phone: 34-96-3877861. Fax: 34-96-3877859. E-mail: rflores{at}ibmcp.upv.es.

Journal of Virology, September 2006, p. 9336-9340, Vol. 80, No. 18
0022-538X/06/$08.00+0     doi:10.1128/JVI.00630-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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