Mutations in Viral Movement Protein Alter Systemic Infection and Identify an Intercellular Barrier to Entry into the Phloem Long-Distance Transport System

Abstract

Viral systemic infection of a plant host involves two processes, cell-to-cell movement and long-distance transport. Molecular determinants associated with these two processes were probed by investigating the effects that alanine scanning mutations in the movement protein (MP) of red clover necrotic mosaic virus (RCNMV) had on viral infection in the plant hostsNicotiana edwardsonii, Vigna unguiculata(cowpea), and the experimental plantNicotiana benthamiana.Plants were inoculated with RCNMV expressing wild-type and mutant forms of the MP. Immunocytochemical studies at the light and electron microscope levels were performed on these plants, using a polyclonal antibody raised against the RCNMV capsid protein to identify the cells/tissues that RCNMV could infect. These experiments demonstrated that one cellular boundary at which the RCNMV MP functions to facilitate entry into the phloem long-distance transport system is located at the interfaces between the bundle sheath and phloem parenchyma cells and the companion cell–sieve element complex. Interestingly, inNicotiana tabacum,a host that only allows a local infection, RCNMV cell-to-cell movement was found to be blocked at this same intercellular boundary. Four mutants that were able to systemically infectN. benthamianawere partially or completely defective for systemic infection ofN. edwardsoniiand cowpea, which indicated that these MP mutants exhibited host-specific defects. Thus, the roles of the RCNMV MP in cell-to-cell movement and in long-distance transport appear to be genetically distinct. These results are discussed in terms of the mechanism by which RCNMV enters the phloem to establish a systemic infection.