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Journal of Virology, December 2009, p. 12057-12067, Vol. 83, No. 23
0022-538X/09/$08.00+0     doi:10.1128/JVI.01151-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Identification of Proteins from Prunus persica That Interact with Peach Latent Mosaic Viroid{triangledown}

Audrey Dubé, Martin Bisaillon, and Jean-Pierre Perreault*

RNA Group/Groupe ARN, Département de Biochimie, Faculté de Médecine et des Sciences de la Santé, Université de Sherbrooke, Sherbrooke, Quebéc J1H 5N4, Canada

Received 5 June 2009/ Accepted 10 September 2009

Peach latent mosaic viroid (PLMVd) is a small, single-stranded, circular RNA pathogen that infects Prunus persica trees. As with all other known viroids, the PLMVd genome does not encode any proteins. Consequently, it must interact with host cellular factors in order to ensure its life cycle. With the objective of identifying cellular proteins that interact with PLMVd, Northwestern hybridizations were performed using partially purified peach leaf extracts. Mass spectrometric analysis of the detected RNA-protein complexes led to the identification of six putative RNA-binding proteins. One of these was found to be elongation factor 1-alpha (eEF1A), and because of its known involvement in the replication and translation of various RNA viruses, further characterizations were performed. Initially, the existence of this interaction received support from an experiment that immunoprecipitated the eEF1A from a crude extract of infected peach leaves, coupled with reverse transcription-PCR detection of the PLMVd. Subsequently, eEF1A interaction with PLMVd strands of both polarities was confirmed in vitro by electrophoresis mobility shift assays, fluorescence spectroscopy, and the prediction of an altered PLMVd RNase mapping profile in the presence of the protein. The potential contribution of eEF1A to the molecular biology of PLMVd, including for viroid replication, is discussed.

* Corresponding author. Mailing address: RNA Group/Groupe ARN, Département de Biochimie, Faculté de Médecine et des Sciences de la Santé, Université de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada. Phone: (819) 564-5315. Fax: (819) 564-5340. E-mail: Jean-Pierre.Perreault{at}USherbrooke.ca

{triangledown} Published ahead of print on 16 September 2009.

Journal of Virology, December 2009, p. 12057-12067, Vol. 83, No. 23
0022-538X/09/$08.00+0     doi:10.1128/JVI.01151-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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