Effects of green fluorescent protein or beta-glucuronidase tagging on the accumulation and pathogenicity of a resistance-breaking Lettuce mosaic virus isolate in susceptible and resistant lettuce cultivars

The RNA genome of a resistance-breaking isolate of Lettuce mosaic virus (LM V-E) was engineered to express the jellyfish green fluorescent protein (GFP ) or beta-glucuronidase (GUS) fused to the helper-component proteinase (HC- Pro) to study LMV invasion and spread in susceptible and resistant lettuce cultivars. Virus accumulation and movement were monitored by either histoch emical GUS assays or detection of GFP fluorescence under UV light. The GFP- and GUS-tagged viruses spread systemically in the susceptible lettuce cult ivars Trocadero and Vanguard, where they induced attenuated symptoms, compa red with the wild-type virus. Accumulation of the GFP-tagged virus was redu ced but less affected than in the case of the GUS-tagged virus. Systemic mo vement of both recombinant viruses was very severely affected in Vanguard 7 5, a lettuce cultivar nearly isogenic to Vanguard but carrying the resistan ce gene mol(2). Accumulation of the recombinant viruses in systemically inf ected leaves was either undetectable (GUS-tag) or erratic, strongly delayed , and inhibited by as much as 90% (GFP-tag). As a consequence, and contrary to the parental virus, the recombinant viruses were not able to overcome t he protection afforded by the mol2 gene. Taken together, these results indi cate that GUS or GFP tagging of the HC-Pro of LMV has significant negative effects on the biology of the virus, abolishing its resistance-breaking pro perties and reducing its pathogenicity in susceptible cultivars.