FIGURE 10.

Pathogenicity determination assays of the four silencing suppressors from CYVMV or CroYVMB. Pathogenicity induced by V2, C2, C4, or βC1 was determined via agroinfiltrating their respective overexpression constructs (fused with eGFP in pEarleyGate103) to N. benthamiana leaves. Five plants were infiltrated for each construct. (A) Variable leaf curl symptoms can be detected in plants infiltrated with V2-GFP, C2-GFP, or C4-GFP constructs (Ai–iii) but not βC1-GFP-infiltrated plants (Aiv). CYVMV and CroYVMB infiltrated (i.e., the positive control) plants also showed similar symptoms (Av). As expected, no symptom was observed with mock infiltrated plants (i.e., the negative control) (Avi). (B) Similar as the positive control, transcripts of all four silencing suppressor genes can be detected in systemically infected leaves of their respective infiltrated plants regardless of the symptomatic or asymptomatic phenotypes (Bi–v). The negative control did not generate any amplification (Bvi). (C) Consistently, confocal microscopy detected expression of the respective GFP-tagged suppressor proteins in all systemically infected leaves (Ci–iv). As expected, GFP expression was not observed in either positive or negative controls (Cv,vi).