Figure 3.

Silencing of NbMAPKKKβ and NbMAPKKKγ suppresses PlAMV-Li1-induced PCD. A) Confirmation of specific MAPKKK gene silencing in gene-silenced plants. The relative quantities of NbMAPKKKα, NbMAPKKKβ, and NbMAPKKKγ mRNA in non-silenced or NbMAPKKKα-, NbMAPKKKβ-, or NbMAPKKKγ-silenced plants were measured using real-time RT-PCR. The data for each leaf tissue sample were normalized to the Nb18S rRNA gene expression level in the same sample. Data shown represent means ± standard deviation of at least three independent plants. Asterisks indicate statistically significant differences from control plants (P < 0.05). B) Typical phenotypes observed in non-silenced and NbMAPKKKβ- and NbMAPKKKγ-silenced plants. Pictures were taken 21 dpi. C) Symptoms in the PlAMV-Li1-infiltrated areas of non-silenced and NbMAPKKKβ- or NbMAPKKKγ-silenced plants (upper panels) and DAB staining of the same leaves (lower panels). Red circles indicate cell death; yellow circles indicate no symptoms. Leaves were infiltrated with a PlAMV-Li1-expressing Agrobacterium culture grown to a turbidity (OD₆₀₀) of 0.05. D) Ion leakage in the PlAMV-Li1-infiltrated areas of non-silenced and NbMAPKKKβ- or NbMAPKKKγ- silenced plants. Data shown represent means ± standard deviation of at least three independent plants. Asterisks indicate statistically significant differences from control plants (P < 0.05).