Molecular studies on bromovirus capsid protein. III. Analysis of cell-to-cell movement competence of coat protein defective variants of cowpea chlorotic mottle virus.

To determine whether the role of coat protein (CP) in cell-to-cell movement of dicot-adapted cowpea chlorotic mottle bromovirus (CCMV) is distinct from that of monocot-adapted brome mosaic bromovirus (BMV), two reporter genes, beta-glucuronidase (GUS) and enhanced green fluorescent protein (EGFP), were substituted for the CP in a biologically active clone of CCMV RNA3 (C3). Primary leaves of Nicotiana benthamiana, Chenopodium quinoa, and cowpea were co-inoculated with wild-type (wt) CCMV RNA 1 and -2 and either C3/delta CP-GUS or C3/delta CP-EGFP and analyzed for GUS activity or the presence of green fluorescence. The visual appearance of infections caused by GUS or EGFP variants indicated that, in CCMV, epidermal cell-to-cell movement can occur without a functional CP. By contrast, inoculation of MP defective variants of C3/delta CP-GUS or C3/delta CP-EGFP resulted in subliminal infections. Additional experiments examining the infectivity of wt BMV RNA 1 and -2 and a BMV RNA3 variant bearing the EGFP in the place of CP (B3/delta CP-EGFP) confirmed previous observations that, unlike CCMV, epidermal cell-to-cell movement of BMV is dependent on the expression of a functional CP. Taken together, the results demonstrate that BMV and CCMV use different mechanisms for initial epidermal cell-to-cell spread, and the individual role played by the respective CP genes in this active process is discussed.