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. 2023 Jul 3;120(28):e2302226120. doi: 10.1073/pnas.2302226120

Fig. 2.

Fig. 2.

Knockout/down of MED10b and other subunits in the middle module of Mediator complex in wild-type N. benthamiana activate plant defense against TSWV infection. (A) A photograph showing a 12-wk-old N. benthamiana plant and an Nbmed10b knockout mutant plant. (B) Western blot analysis of TSWV N protein accumulation in the TSWV-inoculated leaves of the wild-type N. benthamiana plant or Nbmed10b mutant plant at 5 dpi using an N protein–specific antibody. (C) NbMED10b-silenced (TRV-NbMED10b) or nonsilenced (TRV-GUS control) N. benthamiana plants were inoculated with TSWV infectious clone [L(+)opt+M(−)opt+SR(+)eGFP] via agro-infiltration. The infiltrated N. benthamiana plant leaves were harvested at 60 h post-agro infiltration (hpai) and imaged for eGFP fluorescence under an inverted fluorescence microscope (Scale bars, 800 μm.). (D) Western blot results showing the accumulation level of eGFP in the infiltrated leaves shown in (C) using a GFP-specific antibody at 60 hpai. (E) A diagram showing the subunits in the head, middle, and tail modules of Mediator complex in N. tabacum. (F) Subunits NbMED1, NbMED4, NbMED7, NbMED9, NbMED21, NbMED26, and NbMED31 in the middle module; subunits NbMED11 and NbMED18 in the head module; and subunits NbMED15 and NbMED23 in the tail module of Mediator complexes were individually silenced through VIGS using a TRV-based VIGS vector. These N. benthamiana plants were then inoculated individually with the TSWV infectious clone L(+)opt+M(–)opt+SR(+)eGFP via agro-infiltration. The eGFP fluorescence in the inoculated leaves was imaged at 60 h post TSWV inoculation (hpi) under an inverted fluorescence microscope (Scale bars, 800 μm). (G) Western blot analysis results showing eGFP accumulations in various assayed leaf samples shown in (F) using GFP antibody.