Alteration of Intersubunit Acid-base Pair Interactions at the Quasi-threefold Axis of Symmetry of Cucumber Mosaic Virus Disrupts Aphid Vector Transmission

Bricault, Christine

Alteration of Intersubunit Acid-base Pair Interactions at the Quasi-threefold Axis of Symmetry of Cucumber Mosaic Virus Disrupts Aphid Vector Transmission

Abstract

In the atomic model of Cucumber mosaic virus (CMV), six amino acid residues form stabilizing salt bridges between subunits of the asymmetric unit at the quasi-threefold axis of symmetry. To evaluate the effects of these positions on virion stability and aphid vector transmissibility, six charged amino acid residues were individually mutated to alanine. All of the six engineered viruses, mutants D100A, K101A, K127A, D176A, D179A, and K182A, were viable and able to systemically infect Nicotiana tabacum and to locally infect Chenopodium quinoa. In order to assess the physical stability of mutants, virions were purified from plants and tested in a urea disruption assay. Of the six mutant viruses, only mutant K101A had variable stability during purification in the presence of 1.5 M sodium citrate and chloroform. Virions of the remaining five mutants were purified and exhibited wild type levels of virion stability in the presence of urea. Aphid vector transmissibility was nearly or completely eliminated in the case of mutants D100A, K101A, K127A, and D176A. The two remaining mutants were intermediate (mutant D179A) or unaffected (mutant K182A) in their aphid transmission phenotypes. For the majority of the engineered mutants, second-site mutations were observed following aphid transmission and/or following mechanical passaging. In mutant K127A, a spontaneous second-site mutation, E98V, restored transmission rates to that of the wild type. A model is discussed in which the presumptive role of acid-base pairing involved in the dynamic properties of virions in turn affects aphid vector transmission.

Date Issued

2010-08-14T18:27:37Z

Types

dissertation or thesis