Extended Data Fig. 9: Effector proteins colocalized and interacted with ATG8.

(a-c) FRET analysis of the colocalized puncta between YFP tagged avrRpm1-YFP (a), avrRpt2-YFP (b), or avrRps4-YFP (c) and cerulean tagged ATG8e in protoplasts of Arabidopsis suspension cells. The dashed circular outlines indicate examples of colocalized punctae for photobleaching. Arrows indicate the colocalized fluorescence. Separated images of each channel, both pre-bleach and post-bleach, are shown on the right side (from top to bottom: CFP, YFP, and merged). Scale bars, 10 μm. (d) Colocalization analysis of avrRpm1-YFP, avrRpt2-YFP, or avrRps4-YFP with the PVC/MVB marker RFP-Rha1 in protoplasts of Arabidopsis suspension cells. Separated images of each channel are shown on the right side (from top to bottom: YFP, RFP, and merged). Scale bars, 10 μm. (e) The pair of YFPc and YFPn-NBR1 was used as a negative control for NBR1 interaction with avrRpm1, avrRpt2, and avrRps4 in N. benthamiana plants using a BiFC assay. (f) ATG8e interaction with bacterial effectors avrRpm1, avrRpt2, and avrRps4 in N. benthamiana plants using a BiFC assay. ATG8e fused with a N-terminus of YFP (YFPn) was co-expressed with avrRpm1, avrRpt2, or avrRps4 fused with C-terminus YFP (YFPc). Reconstituted YFP fluorescence, colocalized with autophagosome marker mCherry-ATG8e, indicates a positive interaction. The pair of SH3P2-YFPc and YFPn- ATG8e was used as a positive control, while the pair of YFPc and YFPn-ATG8e was used as a negative control. The regions within the white outline are enlarged (Right) (magnification: 5×). Scale bar, 10 µm. The experiments in (d–f) were repeated independently three times with similar results.