Extended Data Fig. 10: The autophagic degradation of effectors was impaired in vsr mutant plant.
(a) Colocalization analysis of GFP/YFP fused effectors and mCherry-ATG8e. Five-day-old transgenic plants expressing autophagosome marker mCherry-ATG8e and the DEX::avrRpm1-GFP, DEX::avrRpt2-GFP, or DEX::avrRps4-YFP were treated with DEX (+) for 24 h, with (+) or without 0.5 μM Conc A for 6 h, followed by confocal analysis. The regions within the white outline are enlarged. Arrows and arrowheads indicate colocalized punctae in cytosol or vacuole, respectively. Scale bar, 10 μm. (b) VSR mutant impair the autophagy degradation of avrRps4-YFP. Confocal images of cotyledon cells from Arabidopsis seedlings of the vsr1vsr6vsr7 transformed with DEX::avrRps4-YFP after DEX induction for 24 h, with (+) or without 0.5 μM Conc A for 6 h, followed by confocal analysis. Note that the vsr1vsr6vsr7 triple mutants accumulated YFP and mCherry positive puncta (arrows) in the cytosol after ConcA treatment, whereas WT Conc A treated plants displayed mostly intravacuolar localization of the puncta (arrowheads). Scale bar, 10 μm. (c) Quantification of the number of avrRps4-YFP punctae in vacuole when plants were treated with DEX (+) and Conc A (+). Data are presented as means ± SD of 20 individual cells from three independent experiments. ****P < 0.0001 in two-tailed unpaired Student’s t-test. Scale bars, 10 μm. The experiments in (a and b) were repeated independently three times with similar results.
