Co-silencing of the Mirabilis antiviral protein (MAP) permits virus-induced gene silencing (VIGS) of other genes in Four O'Clock plants (Mirabilis jalapa)

Virus-induced gene silencing (VIGS) is a powerful and rapid technique for the analysis of gene function during plant growth and development. We attempted to use VIGS for the functional analysis of genes associated with floral senescence and abscission in Four O'Clock plants (Mirabilis jalapa). Plants infected with a Tobacco Rattle Virus (TRV) vector containing a fragment of the phytoene desaturase (PDS) gene did not show the expected photo-bleaching phenotype. We hypothesised that the failure of VIGS in this species might be due to the presence of the Mirabilis antiviral protein (MAP) which is known to have potent activity in preventing viral infection. Plants infected with a TRV vector containing fragments of both the PDS and MAP genes showed the typical photo-bleaching phenotype in photosynthetic tissues, and this was accompanied by reductions in the abundance of PDS and MAP transcripts. We also demonstrated reduced transcript abundance (and the expected phenotypes) when VIGS with a TRV-MAP vector was used to silence the chlorata42 (Ch42) and the proliferating cell nuclear antigen (PCNA) genes. Silencing of the 3,4-dihydroxy-L-phenylalanine 5-glucosyl transferase (DOPA5GT) gene reduced pigmentation in stems, leaves, and flowers. This is the first direct demonstration of the role of an enzyme that has been proposed to be a key step in the synthesis of betalain pigments. Co-silencing of endogenous anti-viral proteins may increase the range of taxa that are amenable to the use of VIGS for functional gene analysis.