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Journal of Virology, November 2006, p. 10395-10406, Vol. 80, No. 21
0022-538X/06/$08.00+0     doi:10.1128/JVI.01137-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Removal of Divalent Cations Induces Structural Transitions in Red Clover Necrotic Mosaic Virus, Revealing a Potential Mechanism for RNA Release^,

Michael B. Sherman,^1* Richard H. Guenther,² Florence Tama,³ Tim L. Sit,² Charles L. Brooks,³ Albert M. Mikhailov,⁴ Elena V. Orlova,⁵ Timothy S. Baker,^1, and Steven A. Lommel²

Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907,¹ Department of Plant Pathology, North Carolina State University, Raleigh, North Carolina 27695,² Department of Molecular Biology, Scripps Research Institute, La Jolla, California 92037,³ Institute of Crystallography, Russian Academy of Sciences, Moscow 119333, Russia,⁴ School of Crystallography, Birkbeck College, London WC1E 7HX, United Kingdom⁵

Received 1 June 2006/ Accepted 9 August 2006

The structure of Red clover necrotic mosaic virus (RCNMV), an icosahedral plant virus, was resolved to 8.5 Å by cryoelectron microscopy. The virion capsid has prominent surface protrusions and subunits with a clearly defined shell and protruding domains. The structures of both the individual capsid protein (CP) subunits and the entire virion capsid are consistent with other species in the Tombusviridae family. Within the RCNMV capsid, there is a clearly defined inner cage formed by complexes of genomic RNA and the amino termini of CP subunits. An RCNMV virion has approximately 390 ± 30 Ca²⁺ ions bound to the capsid and 420 ± 25 Mg²⁺ ions thought to be in the interior of the capsid. Depletion of both Ca²⁺ and Mg²⁺ ions from RCNMV leads to significant structural changes, including (i) formation of 11- to 13-Å-diameter channels that extend through the capsid and (ii) significant reorganization within the interior of the capsid. Genomic RNA within native capsids containing both Ca²⁺ and Mg²⁺ ions is extremely resistant to nucleases, but depletion of both of these cations results in nuclease sensitivity, as measured by a significant reduction in RCNMV infectivity. These results indicate that divalent cations play a central role in capsid dynamics and suggest a mechanism for the release of viral RNA in low-divalent-cation environments such as those found within the cytoplasm of a cell.

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* Corresponding author. Present address: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch at Galveston, Galveston, TX 77555. Phone: (409) 772-6310. Fax: (409) 772-4298. E-mail: mbsherma{at}utmb.edu.

Supplemental material for this article may be found at http://jvi.asm.org/.

Published ahead of print on 18 August 2006.

Present address: Departments of Chemistry & Biochemistry and Molecular Biology, University of California—San Diego, La Jolla, CA 92093.

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Journal of Virology, November 2006, p. 10395-10406, Vol. 80, No. 21
0022-538X/06/$08.00+0     doi:10.1128/JVI.01137-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

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