Generation of an Attenuated, Cross-Protective Pepino mosaic virus Variant Through Alignment-Guided Mutagenesis of the Viral Capsid Protein

Authors

Godwill M. Chewachong, Ohio State UniversityFollow
Sally A. Miller, Ohio State UniversityFollow
Joshua J. Blakeslee, Ohio State UniversityFollow
David M. Francis, Ohio State UniversityFollow
Thomas Jack Morris, University of Nebraska-LincolnFollow
Feng Qu, Ohio State UniversityFollow

Document Type

Article

Date of this Version

2015

Citation

Chewachong, G. M., Miller, S. A., Blakeslee, J. J., Francis, D. M., Morris, T. J., and Qu, F. 2015. Generation of an attenuated, cross-protective Pepino mosaic virus variant through alignment-guided mutagenesis of the viral capsid protein. Phytopathology 105:126-134.

doi: 10.1094/PHYTO-01-14-0018-R

Comments

Copyright © 2015 The American Phytopathological Society. Used by permission.

Abstract

Mild variants of many viruses are able to protect infected plants from subsequent invasion by more severe variants of the same viruses through a process known as cross-protection. In the past, the cross-protective viral variants were commonly derived from mild field isolates that were sometimes genetically heterogeneous, providing variable levels of crossprotection. Here, we report a novel approach to rapidly generate crossprotective variants of the tomato-infecting Pepino mosaic virus (PepMV) independently of the availability of mild field isolates. Our approach sought to attenuate PepMV by mutating less conserved amino acid residues of the abundantly produced capsid protein (CP). These less-conserved amino acid residues were identified through multiple alignments of CPs of six potexviruses including PepMV, and were altered systematically to yield six PepMV mutants. These mutants were subsequently inoculated onto the model plant Nicotiana benthamiana, as well as tomato, to evaluate their accumulation levels, symptom severities, and cross-protection potentials. The mutant KD, in which the threonine (T) and alanine (A) residues at CP positions 66 and 67 were replaced with lysine (K) and aspartic acid (D), respectively, were found to accumulate to low levels in infected plants, cause very mild symptoms, and effectively protect both N. benthamiana and tomato against secondary infections by wild-type PepMV. These data suggest that our approach represents a simple, fast, and reliable way of generating attenuated viral variants capable of cross-protection.